Lignocellulosic Biorefining Technologies Book

Lignocellulosic Biorefining Technologies
Lignocellulosic Biorefining Technologies
Edited by
Avinash P. Ingle
Department of Biotechnology
Engineering School of Lorena
University of São Paulo
Lorena, São Paulo, Brazil
Anuj Kumar Chandel

Silvio Silvério da Silva

This edition first published 2020
© 2020 John Wiley & Sons Ltd
All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or
transmitted, in any form or by any means, electronic, mechanical, photocopying, recording or otherwise,
except as permitted by law. Advice on how to obtain permission to reuse material from this title is available
at http://www.wiley.com/go/permissions.
The right of Avinash P. Ingle, Anuj Kumar Chandel, and Silvio Silvério da Silva to be identified as authors
of the editorial material in this work has been asserted in accordance with law.
Registered Offices
John Wiley & Sons, Inc., 111 River Street, Hoboken, NJ 07030, USA
John Wiley & Sons Ltd, The Atrium, Southern Gate, Chichester, West Sussex, PO19 8SQ, UK
Editorial Office
The Atrium, Southern Gate, Chichester, West Sussex, PO19 8SQ, UK
For details of our global editorial offices, customer services, and more information about Wiley products
visit us at www.wiley.com.
Wiley also publishes its books in a variety of electronic formats and by print‐on‐demand. Some content
that appears in standard print versions of this book may not be available in other formats.
Limit of Liability/Disclaimer of Warranty

While the publisher and authors have used their best efforts in preparing this work, they make no
representations or warranties with respect to the accuracy or completeness of the contents of this work and
specifically disclaim all warranties, including without limitation any implied warranties of merchantability
or fitness for a particular purpose. No warranty may be created or extended by sales representatives, written
sales materials or promotional statements for this work. The fact that an organization, website, or product
is referred to in this work as a citation and/or potential source of further information does not mean that
the publisher and authors endorse the information or services the organization, website, or product may
provide or recommendations it may make. This work is sold with the understanding that the publisher
is not engaged in rendering professional services. The advice and strategies contained herein may not be
suitable for your situation. You should consult with a specialist where appropriate. Further, readers should
be aware that websites listed in this work may have changed or disappeared between when this work was
written and when it is read. Neither the publisher nor authors shall be liable for any loss of profit or any other
commercial damages, including but not limited to special, incidental, consequential, or other damages.
Library of Congress Cataloging‐in‐Publication Data

Names: Ingle, Avinash P., 1981– editor. | Chandel, Anuj Kumar, editor. |
Silva, Silvio Silvério da, editor.
Title: Lignocellulosic biorefining technologies / edited by Avinash P.
Ingle, Anuj Kumar Chandel, Silvio Silvério da Silva.
Description: First edition. | West Sussex, UK : Wiley-Blackwell, 2020. |
Includes bibliographical references and index.
Identifiers: LCCN 2019036678 (print) | LCCN 2019036679 (ebook) | ISBN
9781119568827 (hardback) | ISBN 9781119568810 (adobe pdf) | ISBN
9781119568834 (epub)
Subjects: LCSH: Lignocellulose–Biotechnology. | Biomass–Industrial
applications. | Biomass energy.
Classification: LCC TP248.65.L54 L545 2020 (print) | LCC TP248.65.L54
(ebook) | DDC 662/.88–dc23
LC record available at https://lccn.loc.gov/2019036678
LC ebook record available at https://lccn.loc.gov/2019036679
Cover Design: Wiley
Cover Images: High corn crops ©Polarpx/Shutterstock, Timber in a field ©basel101658/Shutterstock,
A field with straw bales ©donfiore/Shutterstock, Petrochemical industry ©leungchopan/Shutterstock
Set in 9.5/12.5pt STIXTwoText by SPi Global, Pondicherry, India
Printed and bound by CPI Group (UK) Ltd, Croydon, CR0 4YY
10 9 8 7 6 5 4 3 2 1
v
Contents
List of Contributors vii
1 Biorefining of Lignocellulose into Valuable Products 1

Avinash P. Ingle, Anuj Kumar Chandel, and Silvio Silvério da Silva
2 Bulk and Specialty Chemicals from Plant Cell Wall Chemistry 7

Luciana Ferrand, Florencia Vasco, and Juliana Gamboa‐Santos
3 C
haracterization of Lignocellulosic Biomass and Processing for Second-Generation
Sugars Production 29
Guadalupe Bustos Vázquez, Adrián Gonzalez Leos, Luis V. Rodríguez-Duran,
and Rodolfo Torres de Los Santos
4 Production of Biohydrogen from Lignocellulosic Feedstocks 47

Sheetal Radhakrishnan, Shiv Prasad, Sandeep Kumar, and Dhanya Subramanian
5 Recent Advances in the Production of Biodiesel Using Lignocellulosic Biomass 69

Rahul Bhagat, Harris Panakkal, Indarchand Gupta, and Avinash P. Ingle
6 Bioelectricity Production from Lignocellulosic Biomass 87

Samar Das, Shayaram Basumatary, Pankaj Kalita, Vinayak Kulkarni,
Pranab Goswami, Akhil Garg, and Xiongbin Peng
7 B
iopolymers from Lignocellulosic Biomass: Feedstocks, Production Processes,
and Applications 125
Grazielle Machado, Fernando Santos, Rogério Lourega, Jaqueline Mattia, Douglas Faria,
Paulo Eichler, and Angenor Auler
8 Sustainable Production of Biosurfactants and Their Applications 159

Paulo Ricardo Franco Marcelino, Fernanda Gonçalves, Itzcóatl Muñoz Jimenez,
Bruna Curry Carneiro, Bruno Bosquiroli Santos, and Silvio Silvério da Silva
vi Contents
9 L
ignocellulose as a Renewable Carbon Source for Microbial Synthesis
of Different Enzymes 185
Peyman Abdeshahian, Abudukeremu Kadier , Pankaj Kumar Rai, and Silvio Silvério da Silva
10 P
roduction of Organic Acids Via Fermentation of Sugars Generated
from Lignocellulosic Biomass 203
Lourdes Zumalacárregui de Cárdenas and Beatriz Zumalacárregui de Cárdenas
11 Valorization of Lignin Into Value-Added Chemicals and Materials 247

Ruly Teran Hilares, Lucas Ramos, Muhammad Ajaz Ahmed, Avinash P. Ingle, Anuj Kumar
Chandel, Silvio Silvério da Silva, Jeon Woon Choi, and Julio Cesar dos Santos
12 C
onversion of Lignocellulosic Biomass Through Pyrolysis to Promote a Sustainable
Value Chain for Brazilian Agribusiness 265
Genyr Kappler, Débora Machado de Souza, Carlos Alberto Mendes Moraes,
Regina Célia Espinosa Modolo, Feliciane Andrade Brehm, Paulo Roberto Wander,
and Luís António da Cruz Tarelho
13 I ntegrated Process of Biomass Thermochemical Conversion to Obtain Pyrolytic

Sugars for Biofuels and Bioproducts 285
Victor Haber Perez, Nathalia Ribeiro Ferreira da Silva, Euripedes Garcia Silveira Junior,
Diego Cunha Rocha, Oselys Rodriguez Justo, Geraldo Ferreira David, Diana Catalina
Cubides Roman, Valdemar Lacerda, Jr, and Manuel Garcia-Perez
14 L
ife Cycle Analysis of Lignocellulosic Conversion into Fuels, Energy,
and Chemicals 313
Mahdi Mazuchi
15 T
echnoeconomic Analysis of Biorefinery Processes for Biofuel and Other Important
Products 333
Harikishan R. Ellamla and Srinivas Appari
Index 353
vii
List of Contributors
Peyman Abdeshahian Shayaram Basumatary

Department of Biotechnology, Engineering Centre for Energy, Indian Institute of
School of Lorena, University of São Paulo, Technology Guwahati, Guwahati, Assam,
Lorena, São Paulo, Brazil India
Department of Microbiology, Masjed
Soleiman Branch, Islamic Azad University, Rahul Bhagat
Masjed Soleiman, Iran Department of Biotechnology,
Government Institute of Science,
Muhammad Ajaz Ahmed Aurangabad, Maharashtra, India
Graduate School of International
Agricultural Technology, Institute of Bruno Bosquiroli Santos
Green‐Bio Science and Technology, Department of Biotechnology, Engineering
Seoul National University, Pyeongchang, School of Lorena (EEL), University of São
Republic of Korea Paulo (USP), Lorena, São Paulo, Brazil
Guadalupe Bustos Vázquez

Feliciane Andrade Brehm
Department of Biotechnology, University
Graduate Program of Mechanical
Autonomous of Tamaulipas, Unidad
Engineering, University of Vale do Rio
Académica Multidisciplinaria Mante,
dos Sinos – UNISINOS, São Leopoldo,
Cd. Mante, Tamaulipas, México
RS, Brazil
Jeon Woon Choi

Srinivas Appari Graduate School of International
Department of Chemical Engineering, Agricultural Technology, Institute of
Birla Institute of Technology and Science Green‐Bio Science and Technology,
Pilani, Pilani, Rajasthan, India Seoul National University, Pyeongchang,
Republic of Korea
Angenor Auler
Study Center in Biorefinery, State Diana Catalina Cubides Roman
University of Rio Grande do Sul, Porto Federal University do Espírito Santo,
Alegre, Rio Grande do Sul, Brazil Vitoria, Espírito Santo, Brazil
viii List of Contributors
Bruna Curry Carneiro Douglas Faria

Department of Biotechnology, Engineering Polytechnic School, Pontifical Catholic
School of Lorena (EEL), University of São University of Rio Grande do Sul, Porto
Paulo (USP), Lorena, São Paulo, Brazil Alegre, Rio Grande do Sul, Brazil
Luís António da Cruz Tarelho

Luciana Ferrand
Department of Environment and Planning/
Instituto de Biotecnología y Biología
Centre for Environmental and Marine
Molecular, Universidad Nacional de La
Studies (CESAM), University of Aveiro
Plata, La Plata, Argentina
(UA), Aveiro, Portugal
Samar Das Geraldo Ferreira David

Centre for Energy, Indian Institute of Federal University do Espírito Santo,
Technology Guwahati, Guwahati, Assam, Vitoria, Espírito Santo, Brazil
India
Juliana Gamboa-Santos
Julio Cesar dos Santos
Centro de Investigación y Desarrollo en
Department of Biotechnology, Engineering
Criotecnología de Alimentos, Universidad
School of Lorena, University of São Paulo,
Nacional de La Plata, La Plata, Argentina
Diego Cunha Rocha Manuel Garcia-Perez

State University of Northern Rio de Biological System Engineering,
Janeiro, Campos dos Goytacazes, Rio de Washington State University, Pullman,
Janeiro, Brazil WA, USA
Paulo Eichler Euripedes Garcia Silveira Junior

Polytechnic School, Pontifical Catholic State University of Northern Rio de
University of Rio Grande do Sul, Porto Janeiro, Campos dos Goytacazes, Rio de
Alegre, Rio Grande do Sul, Brazil Janeiro, Brazil
Harikishan R. Ellamla
Akhil Garg
South African Institute for Advanced
Department of Mechatronics Engineering,
Materials Chemistry (SAIAMC),
Shantou University, Shantou, Guangdong,
University of Western Cape, Cape Town,
China
South Africa
Regina Célia Espinosa Modolo Fernanda Gonçalves

Graduate Program of Civil Engineering, Department of Biotechnology, Engineering
University of Vale do Rio dos Sinos – School of Lorena (EEL), University of São
UNISINOS, São Leopoldo, RS, Brasil, Paulo (USP), Lorena, São Paulo, Brazil
São Leopoldo, Rio Grande do Sul, Brazil
and Adrián Gonzalez Leos
Graduate Program of Mechanical Department of Biotechnology, University
Engineering, University of Vale do Rio Autonomous of Tamaulipas, Unidad
dos Sinos – UNISINOS, São Leopoldo, Académica Multidisciplinaria Mante,
RS, Brazil Cd. Mante, Tamaulipas, México
List of Contributors ix
Pranab Goswami Sandeep Kumar

Centre for Energy, Indian Institute of Centre for Environment Science and Climate
Technology Guwahati, Guwahati, Assam, Resilient Agriculture, Indian Agricultural
India Research Institute, New Delhi, India
Indarchand Gupta Anuj Kumar Chandel

Department of Biotechnology, Department of Biotechnology, Engineering
Government Institute of Science, School of Lorena, University of São Paulo,
Aurangabad, Maharashtra, India Lorena, São Paulo, Brazil
Victor Haber Perez Pankaj Kumar Rai

State University of Northern Rio de Department of Biotechnology, Invertis
Janeiro, Campos dos Goytacazes, Rio de University, Bareilly, Uttar Pradesh (UP),
Janeiro, Brazil India
Avinash P. Ingle Valdemar Lacerda Jr

Department of Biotechnology, Engineering Federal University do Espírito Santo,
School of Lorena, University of São Paulo, Vitoria, Espírito Santo, Brazil
Rogério Lourega
Abudukeremu Kadier
Polytechnic School, Pontifical Catholic
Department of Chemical and Process
University of Rio Grande do Sul, Porto
Engineering, Faculty of Engineering and
Alegre, Rio Grande do Sul, Brazil
Built Environment, National University of
Malaysia, Bangi, Selangor, Malaysia
Grazielle Machado
Research Centre for Sustainable Process
Polytechnic School, Pontifical Catholic
Technology (CESPRO), Faculty of
University of Rio Grande do Sul, Porto
Engineering and Built Environment,
Alegre, Rio Grande do Sul, Brazil
National University of Malaysia, Bangi,
Selangor, Malaysia
Débora Machado da Souza
Pankaj Kalita Graduate Program of Mechanical
Centre for Energy, Indian Institute of Engineering, University of Vale do Rio
Technology Guwahati, Guwahati, Assam, dos Sinos – UNISINOS, São Leopoldo,
India RS, Brazil
Genyr Kappler Paulo Ricardo Franco Marcelino

Graduate Program of Civil Engineering, Department of Biotechnology,
University of Vale do Rio dos Sinos – Engineering School of Lorena (EEL),
UNISINOS, São Leopoldo, RS, Brasil, University of São Paulo (USP), Lorena,
São Leopoldo, Rio Grande do Sul, Brazil São Paulo, Brazil
Vinayak Kulkarni Jaqueline Mattia

Centre for Energy, Indian Institute of Study Center in Biorefinery, State
Technology Guwahati, Guwahati, Assam, University of Rio Grande do Sul, Porto
India Alegre, Rio Grande do Sul, Brazil
x List of Contributors
Mahdi Mazuchi Nathalia Ribeiro Ferreira da Silva

Iranian Sugarcane and By‐product State University of Northern Rio de
Research and Training Institute, Ahvaz, Janeiro, Campos dos Goytacazes, Rio de
Iran Janeiro, Brazil
Carlos Alberto Mendes Moraes Luis V. Rodríguez-Duran

Graduate Program of Civil Engineering, Department of Biotechnology, University
University of Vale do Rio dos Sinos – Autonomous of Tamaulipas, Unidad
UNISINOS, São Leopoldo, RS, Brasil, Académica Multidisciplinaria Mante,
São Leopoldo, Rio Grande do Sul, Brazil Cd. Mante, Tamaulipas, México
and
Graduate Program of Mechanical Oselys Rodriguez Justo
Engineering, University of Vale do Rio Estácio de Sá University, Campos dos
dos Sinos – UNISINOS, São Leopoldo, Goytacazes, Rio de Janeiro, Brazil
RS, Brazil
Fernando Santos
Itzcóatl Muñoz Jimenez Study Center in Biorefinery, State
Department of Biotechnology, Engineering University of Rio Grande do Sul, Porto
School of Lorena (EEL), University of São Alegre, Rio Grande do Sul, Brazil
Paulo, Lorena (USP), São Paulo, Brazil
Silvio Silvério da Silva
Harris Panakkal Department of Biotechnology, Engineering
Department of Biotechnology, School of Lorena (EEL), University of São
Government Institute of Science, Paulo (USP), Lorena, São Paulo, Brazil
Aurangabad, Maharashtra, India
Dhanya Subramanian
Xiongbin Peng Centre for Environmental Science and
Department of Mechatronics Engineering, Technology, Central University of Punjab,
Shantou University, Shantou, Guangdong, Bathinda, Punjab, India
China
Ruly Teran Hilares
Shiv Prasad Department of Biotechnology, Engineering
Centre for Environment Science and School of Lorena, University of São Paulo,
Climate Resilient Agriculture, Indian Lorena, São Paulo, Brazil
Agricultural Research Institute, New Delhi,
India Rodolfo Torres de Los Santos
Department of Biotechnology, University
Sheetal Radhakrishnan Autonomous of Tamaulipas, Unidad
Regional Research Station, ICAR‐Central Académica Multidisciplinaria Mante,
Arid Zone Research Institute, Bikaner, Cd. Mante, Tamaulipas, México
Rajasthan, India
Florencia Vasco
Lucas Ramos Centro de Investigación y Desarrollo
Department of Biotechnology, Engineering en Criotecnología de Alimentos,
School of Lorena, University of São Paulo, Universidad Nacional de La Plata, La
Lorena, São Paulo, Brazil Plata, Argentina
List of Contributors xi
Paulo Roberto Wander Lourdes Zumalacárregui de Cárdenas

Graduate Program of Civil Engineering, Chemical Engineering Department,
University of Vale do Rio dos Sinos – Chemical Engineering Faculty,
UNISINOS, São Leopoldo, RS, Brasil, Technological University of Havana,
São Leopoldo, Rio Grande do Sul, Brazil Havana, Cuba
Beatriz Zumalacárregui de Cárdenas

Chemical Engineering Department,
Chemical Engineering Faculty,
Technological University of Havana,
Havana, Cuba
1
Biorefining of Lignocellulose into Valuable Products

Avinash P. Ingle, Anuj Kumar Chandel, and Silvio Silvério da Silva
Department of Biotechnology, Engineering School of Lorena, University of São Paulo, Lorena, São Paulo, Brazil
Lignocellulosic biomass is a major stakeholder in biorefineries. The transformation of

biomass into a wide range of fuels, materials, and valuable chemicals is the overall goal of
a biorefinery (de Jong and Jungmeier 2015; Chandel et al. 2018). In principle, a biorefin-
ery would exploit hybrid technologies encompassing various fields including bioengi-
neering, agriculture, and polymer chemistry. In a specific biorefinery, the feedstock is
fractionated into valuable constituents through extraction, hydrolysis, fermentation, and
controlled pyrolysis for the production of fuels, energy, and high‐value products such as
organic acids, biopigments, biosurfactants, etc. (Erickson et al. 2012; Isikgor and Becer
2015; Lee et al. 2019).
The concept of biomass fractionation into its main components offers myriad benefits to
the bioprocessing industries harnessing the various feedstocks. Nevertheless, the advance-
ment and exploitation of lignocellulosic biomass fractionation technologies are still in their
infancy in terms of technoeconomic viability (FitzPatrick et al. 2010; Chandel et al. 2018).
Hence, fundamental and applied research will be critically required in this field in the
coming decades.
Currently, the price of biomass is a major impeding factor in the economic production of
fuels and value‐added products/chemicals that accounts for up to 40–60% of the overall
price (Chandel et al. 2019). For the overall economization of the process with simplicity,
process integration is an important necessity. The second‐generation biomass together
with waste feedstocks are promising sources for the production of value‐added products
owing to their surplus availability, cost‐effectiveness and non‐requirement of land with no
competition with food crops (Chandel and Silveira 2017). However, the exploitation of lig-
nocellulosic biomasses to generate fuels and valuable chemicals is challenging because of
the complexity of pretreatment followed by enzymatic hydrolysis via the synergistic action
of cellulases to yield cellulosic sugars which are considered as renewable building blocks
(Chandel et al. 2019). Cellulosic sugars can be further converted into a plethora of bio‐
based products such as alcohols, organic acids, alkenes, lipids, etc. (Lee et al. 2019).
Lignocellulosic Biorefining Technologies, First Edition. Edited by Avinash P. Ingle,

Anuj Kumar Chandel, and Silvio Silvério da Silva.
© 2020 John Wiley & Sons Ltd. Published 2020 by John Wiley & Sons Ltd.
2 Lignocellulosic Biorefining Technologies
However, the production potential of biochemicals from these agro‐residues have not yet
been investigated at large scale under biorefinery conditions (Sanford et al. 2016).
Considering the enormous potential of different lignocellulosic biomass types in
biorefinery, as mentioned above, we have attempted in this book to explore the high‐value
biorefining products which can be created using a variety of lignocellulosic biomass as
renewable and economically viable resources.
The book contains 15 chapters. Chapter 1 by Ingle et al., which is also the book pref-
ace, presents the overall impact of lignocellulose biorefineries in a nutshell in addition
to summarizing each chapter’s content. Chapter 2 by Ferrand and colleagues is broadly
focused on the various bulk and specialty chemicals present in the plant cell wall.
Special emphasis has been given to important aspects such as structure, function, and
chemical composition of the plant cell wall. In addition, all the promising valuable
chemicals and bioactive compounds present the plant cell wall are discussed in detail.
Chapter 3 by Bustos et al. provides details about the different components present in
the lignocellulosic biomass and their characterization; different approaches available
for processing lignocellulosic biomass into second‐generation sugars are also dis-
cussed. Sheetal et al. in Chapter 4 focus on the possibilities of utilization of lignocel-
lulosic feedstocks for the production of biohydrogen. It is well known that hydrogen
energy, particularly biohydrogen, is gaining a lot of interest as a sustainable and renew-
able alternative to fast‐depleting fossil fuels. It also provides an additional benefit of
not emitting any greenhouse gases (GHG), which provides an incentive to all countries
struggling to meet the GHG limitations as per the Paris agreement to combat climate
change to adopt this clean fuel. Various recent studies have proved that lignocellulosic
biomass sources can be used as an alternate feedstock for biohydrogen production as
they are abundant, cheap, and eco‐friendly.
Chapter 5 by Bhagat and co‐authors emphasizes the role of a variety of lignocellulosic
biomass in the production of another form of clean energy – biodiesel. In this chapter, the
authors briefly discuss the major constituents of lignocellulosic biomass, and the composi-
tion and structure of each of the components present in the biomass have been explained.
Most importantly, recent advances in the production of biodiesel using lignocellulosic
biomass through different fermentation approaches have been discussed. It is forecast that
the demand for renewable energy for transportation is likely to grow by 19% till 2023
whereas approximately a 15% rise in biofuel production is expected in the same time period.
In this context, biodiesel can be seen as a renewable energy source that can be used for
partial or even total replacement of diesel.
Chapter 6 by Kalita et al. covers the production of bioelectricity from lignocellulosic bio-
mass. Electricity is a vital form of energy which plays a significant role in defining human
development. On one side, industrial and economic developments are transforming our
way of living and therefore demand for electricity demand is continuously growing.
However, on the other side, this continuous increase in energy demand leads to fossil fuel
depletion and environmental degradation. Therefore, to resolve these issues, more empha-
sis is now being place on harvesting sustainable energy from different renewable energy
sources like solar, biomass, wind, etc. In this context, the authors have discussed the vari-
ous options which can be used in the conversion of lignocellulosic biomass into valuable
fuels through thermochemical conversion technologies.
Biorefining of Lignocellulose into Valuable Products 3
As mentioned earlier, biohydrogen, biodiesel, and bioelectricity are imporant high‐value

products commonly produced from lignocellulosic feedstocks through different biorefin-
ing strategies. Lignocellulosic materials can also be potentially utilized in the production of
other valuable products with applications in numerous sectors like food and agriculture,
pharmaceutics, biomedicine, cosmetics, etc. In view of this, various chapters in the present
book have a special emphasis on different lignocellulosic biorefining products. In Chapter 7,
Machado et al. provide a special focus on the production of biopolymers using different
lignocellulosic materials as important sources of carbon, nitrogen, etc. The authors report
that production of biopolymers from renewable resources like lignocellulosic feedstocks
has been increasing due to environmental, political and economic concerns about conven-
tional plastics utilization. A wide range of biopolymers with many application possibilities
can be produced from lignocellulosic biomass, allowing the replacement of many conven-
tional plastics. Therefore, in this chapter, various key aspects including types and proper-
ties of biopolymers, different approaches used for biopolymer production, applications of
biopolymers, and the advantages and challenges of obtaining biopolymers from lignocel-
lulosic biomass are critically discussed.
In the same vein, Marcelino and co‐authors explain the importance of lignocellulosic
biomass in the production of biosurfactants in Chapter 8. Biosurfactants are amphipathic
molecules synthesized by microorganisms (bacteria, yeasts, and filamentous fungi) using a
variety of lignocellulosic feedstocks as a source of carbon and nitrogen which are essential
for the growth of these microorganisms. Biosurfactants have attracted a great deal of atten-
tion across the world due to their unique and novel surface‐active and/or emulsifying, anti-
microbial, and antitumor properties. Important topics including the types, structure, and
functions of biosurfactants, fermentation approaches used for biosurfactant production
and applications of biosurfactants are critically discussed in Chapter 8.
Chapter 9 by Abdeshahian et al. is focused on the role of lignocellulosic materials in the
production of different enzymes. In this chapter, the authors propose that lignocellulosic
biomass or wastes from various sources like agriculture, forest, industries, etc. can be
employed as promising raw materials in the production of value‐added products such as
enzymes using various biorefining technologies. Generally, the production of enzymes is
carried out by microorganisms through the fermentation process in which organic carbon
contents are provided in the form of lignocellulose which is utilized as a nutrient source by
fermenting organisms. The utilization of lignocellulosic materials could provide a cost‐
effective raw material for enzyme production which in turn reduces the enzyme produc-
tion cost.
Considering the vital role of organic acids in various biological processes, it is the need of
the hour to produce different organic acids using simple, efficient, economic, and environ-
mentally sustainable approaches. In this context, lignocellulosic biomass can be used as
inexpensive and renewable sources for the production of organic acids. Therefore, de
Cárdenas and co‐authors in Chapter 10 discuss the production of organic acids using ligno-
cellulosic feedstocks. Moreover, recent advances in the production of different acids using
lignocellulosic biomass are discussed in this chapter along with important related factors.
Chapter 11 by Terán‐Hilares et al. covers recent advances in the valorization of lignin
into value‐added products. Lignin is one of the most abundant macromolecules on Earth.
It is a complex fraction in biomass composed of various aromatic building blocks which are
cross‐linked with different carbon and ether linkages. Lignin has broad scope for valorization
to aromatics, polymers, and other value‐added materials. However, in spite of the attractive-
ness of lignin as a natural source for the production of a wide range of products, there are vari-
ous technologic barriers which limit its ubiquitous use at the industrial level. Considering all
these concerns, Chapter 11 focuses on the potential of lignin in the creation of various high‐
value products. In addition, the chemistry of lignin, various approaches for its processing, and
economic and environmental concerns associated with lignin valorization are discussed.
Chapter 12 by Moraes and co‐authors provides a detailed explanation of pyrolysis and
carbonization, which are commonly used methods for the processing of biomass to develop
by‐products for energy and agriculture. Special emphasis has been given to the thermo-
chemical conversion of agricultural or lignocellulosic biomass to obtain a solid product like
biochar.
Chapter 13 by Perez et al. is about the integrated processes for thermochemical conver-
sion of biomass to produce pyrolytic sugars like levoglucosan required for biofuels and
other important bioproducts by fermentation. In this chapter, the authors discuss the inte-
grated processes of a biomass thermochemical conversion plant as a subprocess of an
autonomous bioethanol plant. Further, they emphasize that such technologic alternatives
can be inexpensive, eco‐friendly, and attractive in a country like Brazil where surplus
amounts of sugarcane bagasse are available.
Chapter 14 by Mazuchi is focused on a life cycle analysis of lignocellulosic conversion
into value‐added products. In order to study the impacts of different materials, products,
and processes on environmental sustainability, a life cycle assessment is required so vari-
ous aspects of such an assessment have been discussed in this chapter.
The last chapter, by Ellamla, covers the most important issues associated with the biore-
finery industries. It provides details about the technoeconomic analysis of biofuel produc-
tion and other important products. As far as biorefinery industries are concerned, a
technoeconomic analysis is essential to assess the feasibility of integrating lignocellulosic
biomass into various biorefinery products like biofuels and other high‐value compounds.
Lignocellulosic Biorefining Technologies is a collection of articles elucidating recent
advances in the utilization of a variety of lignocellulosic feedstocks for the production of
high‐value products including important forms of bioenergy and other industrially impor-
tant biorefining compounds like biopolymers, biosurfactants, enzymes, organic acids, etc.
The text in each chapter is supported by clear, informative tables and figures. Each chapter
contains relevant references to published articles, which offer a large amount of primary
information and further links to a nexus of data and ideas.
All the chapters in this book have been written by one or more specialists, experts in their
field, and are highly informative and detailed. In this way, we would like to offer a rich
guide for researchers, undergraduate or graduate students of various disciplines such as
agriculture, food science, biotechnology, biofuel and bioenergy industries, and allied sub-
jects. In addition, this book will be useful for those working in various industries, regula-
tory bodies, and global fuel and energy organizations.
The editors are very grateful to all the contributors for their outstanding efforts to provide
state‐of‐the‐art information on the subject matter of their respective chapters. Their efforts
will certainly enhance and update the knowledge of readers about lignocellulosic biorefin-
ing technologies. We express our sincere thanks to the publishers and authors whose
Biorefining of Lignocellulose into Valuable Products 5
research has been cited in the book. We are also thankful to Rebecca Ralf, Sindhuja
Sethuraman, and the team at John Wiley and Sons Ltd. for their generous cooperation and
efforts in producing this book.
Among the editors, Avinash P. Ingle is very grateful to the Research Council for the State
of Sao Paulo (FAPESP), Brazil, for providing financial assistance (Process No. 2016/22086‐2)
in the form of a postdoctoral fellowship. Anuj Kumar Chandel is grateful to the Brazilian
Federal Agency for the Support and Evaluation of Graduate Education (USP‐CAPES) for a
visiting researcher and professor fellowship. Silvio Silvério da Silva is grateful to the
Brazilian National Council for Scientific and Technological Development (CNPq) (Process
No. 303943/2017‐3) and FAPESP (Process No. 2016/10636-8) for providing support for research.
We hope that the book will be useful for all readers looking for information on the latest
research and advances in the field of lignocellulosic biorefining technologies.
References
Chandel, A.K. and Silveira, M.H.L. (2017). Sugarcane Bio‐Refinery: Technologies,

Commercialization, Policy Issues and Paradigm Shift. Amsterdam, The Netherlands: Elsevier.
Chandel, A.K., Garlapati, V.K., Singh, A.K. et al. (2018). The path forward for lignocellulose
biorefineries: bottlenecks, solutions, and perspective on commercialization. Bioresource
Technology 264: 370–381.
Chandel, A.K., Albarelli, J.Q., dos Santos, D.T. et al. (2019). Comparative analysis of key
technologies for cellulosic ethanol production from Brazilian sugarcane bagasse at the
commercial‐scale. Biofuels, Bioproducts and Biorefining 13 (4): 994–1014.
Erickson, B., Nelson, J.E., and Winters, P. (2012). Perspective on opportunities in industrial
biotechnology in renewable chemicals. Biotechnology Journal 7: 176–185.
FitzPatrick, M., Champagne, P., Cunningham, M.F., and Whitney, R.A. (2010). A biorefinery
processing perspective: treatment of lignocellulosic materials for the production of value‐
added products. Bioresource Technology 101: 8915–8922.
Isikgor, F.H. and Becer, C.R. (2015). Lignocellulosic biomass: a sustainable platform for the
production of bio‐based chemicals and polymers. Polymer Chemistry 6: 4497–4559.
de Jong, E.D. and Jungmeier, G. (2015). Biorefinery concept in comparison to petrochemical
refineries. In: Industrial Biorefineries and White Biotechnology (eds. A. Pandey, R. Hofer,
C. Larroche, et al.), 3–33. Amsterdam, The Netherlands: Elsevier.
Lee, S.Y., Kim, H.U., Chae, T.U. et al. (2019). A comprehensive metabolic map for production
of bio‐based chemicals. Nature Catalysis 2: 18–33.
Sanford, K., Chotani, G., Danielson, N., and Zahn, J.A. (2016). Scaling up of renewable
chemicals. Current Opinion in Biotechnology 38: 112–122.
7
Bulk and Specialty Chemicals from Plant

Cell Wall Chemistry
Luciana Ferrand1, Florencia Vasco2, and Juliana Gamboa‐Santos2
1
Instituto de Biotecnología y Biología Molecular, Universidad Nacional de La Plata, La Plata, Argentina
2
Centro de Investigación y Desarrollo en Criotecnología de Alimentos, Universidad Nacional de La Plata, La Plata, Argentina
2.1 Introduction
Plant cell walls are the most abundant renewable resource on our planet, representing 70%
of the annual biomass production by land plants worldwide (Pauly and Keegstra 2008). The
woody material of plant cell walls comprises three main types of carbon‐based poly-
mer – cellulose, hemicellulose, and lignin – which collectively are called lignocellulosic
biomass (LG) (Sanderson 2011). However, not only the woody material of plants is being
used as a natural source of cellulosic and noncellulosic materials, but also leaves, branches,
flowers, algae, seeds, fruits and vegetables, among others (Asgher et al. 2013; Bernaerts
et al. 2018). LG biomass can even come from organic wastes or by‐products from other
industries, contributing to the ecologic cycle that is being demanded around the globe
(Ekman et al. 2013; Ofori‐Boateng and Lee 2013; Faris et al. 2015; Petkowicz et al. 2017;
Ravindran and Jaiswal 2016; Chrysikou et al. 2018).
Despite agricultural practices and various agro‐based industries generating huge
amounts of LG biomass every year (about 933 million tons), only 2% of this resource is
utilized by humans (Pauly and Keegstra 2008; De Bhowmick et al. 2018). This means that
a considerable amount of such LG materials is sadly discarded. However, LG biomass is
gaining increasing research interest and special importance because of its renewable nature
and the great amount of lost resources (Lyu et al. 2018; Oyola‐Rivera et al. 2018; Mei et al.
2019; Luo et al. 2019). Furthermore, LG materials can potentially be converted into differ-
ent high‐value products such as biofuels, value‐added chemicals, and cheap energy sources
for microbial fermentation and enzyme production (Anwar et al. 2014; Naseem et al. 2016;
Cao et al. 2018; Ma et al. 2018). Since a great amount of these materials are generated from
atmospheric CO2, water, and sunlight through biological photosynthesis, biomass is con-
sidered to be the only sustainable source of organic carbon on Earth. Considering the
depletion of petroleum resources, the intensive utilization of fossil fuels and the awareness
of global warming, LG biomass appears to be the perfect substitute for petroleum for the

production of fuels and fine chemicals with net zero carbon emission. It also has a low
content of nitrogen, sulfur and ash, which results in lower emissions of NO, SO2 and soot,
compared to conventional fuels (Isikgor and Becer 2015; Dhyani and Bhaskar 2018).
From the first generation of biofuels obtained from energy crops to the second generation
originated from many types of biomass, many challenges have been overcome. Nowadays,
in Europe, more than 220 LG biorefineries are in operation, of which approximately 180 are
first generation, while the remainder belong to the second generation (Hassan et al. 2019).
Therefore, biorefineries which allow both bioethanol production and co‐production of
biochemicals are still in expansion, also being an environmentally friendly system to help
mitigate global climate change (Chrysikou et al. 2018).
The aim of this chapter is to explore the different value‐added chemicals that could be
extracted from cell wall polymers and used as platform chemicals or feedstock to obtain
high‐interest chemicals, pharmaceutical, biomedical or food products. To this end, we will
briefly survey the basics of plant cell wall structure and function. After a short description
of the chemical composition of the plant cell wall, a detailed overview of the valuable
chemicals that can be obtained from LG biomass is given.
This chapter could help chemists, biotechnologists, agronomists, and food technologists
to overview the LG biomass reutilization potential in order to obtain the high value added
from block units to bioactive products.
2.2 Plant Cell Wall Structure and Function
The plant cell wall is a complex matrix of polysaccharides that provides support and
strength, essential for plant cell survival. Polysaccharides perform a great diversity of func-
tions during plant life, including:
i) supporting the cell membrane and preventing it from bursting
ii) expanding under turgor pressure at a controlled rate and direction, facilitating and
regulating cell growth
iii) cooperating with adjacent cells under similar turgor pressure to build a mechanically
competent 3D tissue with every cell wall maintained in tension (Jarvis 2011)
iv) protecting against other organisms and environmental stresses.
The cell wall of higher plants is not a uniform structure and is composed of three major
layers (Figure 2.1):
i) the middle lamella or lamellum, which is a thin layer of approximately 50 nm thickness
sandwiched between the primary cell walls of neighboring cells. The lamellum is the
first formed layer that cements the cell walls of two adjacent cells and is composed of
calcium and magnesium pectates (Zamil and Geitmann 2017)
ii) the primary cell wall, which lies down the middle lamella, surrounds and protects the
cell, contributing to the wall structural integrity, cell adhesion, and signal transduction
(McCann and Carpita 2008);
iii) the secondary cell wall, located between the primary wall and the plasma membrane, is
produced only in some cells once cell elongation ceases. Secondary walls may be depos-
ited in a number of layers (S1, S2, S3) and are mainly found in tracheary elements
Bulk and Specialty Chemicals from Plant Cell Wall Chemistry 9
Pectin
Cellulose
Hemicellulose
Lignin
S3 S2 S1
Middle lamella
Primary wall
Secondary
wall
Cell membrane
Cytoplasm
Figure 2.1 Plant cell wall structure and composition.
“tracheids in seedless vascular plants and gymnosperms and vessels in angiosperms”

and fibers in the primary xylem and the secondary xylem (wood) (Zhong and Ye 2015).
2.3 Plant Cell Wall Chemical Composition
The plant cell wall is a complex network of cellulosic, hemicellulosic, and pectic polysac-
charides and proteins (Altartouri and Geitmann 2015) whose composition depends on
plant species and cell type (Davison et al. 2013). However, from a general point of view,
they all have a conservative composition involving two phases: a fibrillar phase, which acts
as the skeleton and is composed mainly of cellulose microfibrils, and a matrix phase, which
contains a high proportion of noncellulosic polysaccharides that vary in their chemical
structures. Structural proteins, glycoproteins, and phenolic components, including lignin,
may also be present in the wall matrix (Harris 2006).
The matrix phase of primary walls is almost completely hydrated (65% water), con-
sisting of hemicelluloses and pectins, with some structural proteins (Brett and Waldron
1996). Primary cell walls are usually classified as type I or type II, according to their
polysaccharide composition. Type I walls contain cellulose, xyloglucan as the main
hemicellulose, and abundant amounts of pectin. These primary walls are present in
dicots and noncommelinoid monocots. Type II walls are present in grasses (Poales) cell
walls and contain a higher amount of cellulose with negligible percentages of pectin
and proteins. The major hemicellulose in type II walls is arabinoxylan (Carpita and
McCann 2000).
Secondary walls contain cellulose and arabinoxylan and glucomannans as the major
hemicelluloses (Brett and Waldron 1996). In secondary walls, the water in the matrix phase
is largely replaced by lignin, making them nearly impenetrable to solutes and enzymes.
2.3.1 Cellulosic Polysaccharides

Cellulose is a major structural component of plant cell walls, which is responsible for
mechanical strength. It is a linear polymer of D‐anhydro‐glucopyranose molecules joined
by β‐1,4‐glycosidic bonds (Figure 2.2) (Alexandridis et al. 2018). Due to their linearity and
regular structure, the cellulose molecules associate with each other over large regions,
forming bundles of polycrystalline fibers. The degree of polymerization (DP) of cellulose
largely depends on the origin and pretreatments applied to the raw LG biomass. DP defines
the solubility in water; oligosaccharides having a DP above 13 will probably present some
difficulties in forming homogeneous water solutions while oligosaccharides with a DP over
30 become similar to cellulose. Due to the huge amount of hydroxyl groups, cellulose pre-
sents an intra‐ and intermolecular hydrogen bonding network, which makes the crystalline
structure robust. To solubilize cellulose polymer, the majority of hydroxyl bonds should
break simultaneously. Thus, at mild temperatures, cellulose is insoluble not only in water
but also in most typical solvents. Cellulose is one of the most important biomass resources
due to its abundance in nature, with the peculiarity of not being included in the human
OH OH
O O COOMe
O O HO
O
HO O O
HO
OH
OH OH
(a) (c)
H H H OH
HO O O OH OH OH
H HO
HO H OH HO H OH OH
H OH H H H
H
(i) (ii)
H OH OH
HO MeO MeO OMe
HO O H O
H H OH
HO H OH HO H H OH OH
H OH H H OH OH (i) (ii) (iii)
(iii) (iv)
(b) (d)
Figure 2.2 Chemical structure of the basic constituents of lignocellulosic biomass. Building blocks
of (a) cellulose: β‐d‐glucopyranose; (b) hemicellulose: (i) β‐D‐xylopyranose, (ii) β‐D‐mannopyranose,
(iii) β‐d‐glucopyranose, (iv) ɑ‐d‐galactopyranose; (c) pectin; (d) lignin: (i) p‐coumaryl alcohol, (ii)
coniferyl alcohol, (iii) synapyl alcohol.
food chain. Cellulose applications involve paper products, textiles, polymer composites,
and chemical precursors of pharmaceutics, food, drinks, and coatings (Deng et al. 2015b).
2.3.2 Noncellulosic Polysaccharides

Noncellulosic polysaccharides present in plant cell matrix encompass mainly hemicellu-
loses (d‐xylans, d‐glucans, d‐mannans, etc.) and pectins (d‐galacturonans).
2.3.2.1 Hemicellulose
Hemicellulose is the second most abundant polysaccharide after cellulose in plant cell
walls, accounting for 15–30% of lignocellulosic biomass by weight (Kapu and Trajano
2014). Although hemicelluloses are found in both primary and secondary cell walls of both
monocotyledonous and dicotyledonous plant tissues, greater amounts of hemicellulose in
wood and woody biomass than in herbaceous and agricultural biomass have been reported
(Vassilev et al. 2012). Among the three main components in biomass, hemicellulose is a
promising material to produce value‐added chemicals. Hemicellulose consists of a short,
highly branched polymer of five‐ and six‐carbon polysaccharide units, such as xylan, man-
nan, β‐glucans, and xyloglucans (Cao et al. 2018; Luo et al. 2019). Compared to cellulose
and lignin, hemicellulose has a lower DP (100–200 U). Hemicellulose is more unstable than
cellulose and therefore degrades more easily when subjected to heat treatment. Although
there are many studies on the conversion of cellulose and lignin, those about hemicellulose
conversion are limited.
2.3.2.2 Pectic Polysaccharides

Pectin is one of the most important cell wall polysaccharides, mainly those belonging to
vegetables and fruits, and is linked covalently to form a gel matrix, interspacing the cellu-
lose‐hemicellulose network in the primary cell wall. It is probably the most complex mac-
romolecule found in nature, as it can be composed of as many as 17 different monosaccharide
units containing more than 20 different linkages. Historically, the compound was dubbed
“pectic acid” coming from the Greek word πηӼƬες, whose translation is coagulated mate-
rial. The name was coined by the French scientist Henri Braconnot, who first isolated pec-
tin from vegetables in 1825 (Yanakieva et al. 2012; Chan et al. 2017). Although a long time
has passed since the discovery of pectin, its structural and chemical properties are still the
subject of study around the world due to the heterogeneity of this polymer.
Regarding the basic structure, pectin is a heteropolysaccharide predominantly contain-
ing galacturonic acid (GalA) residues, an oxidized form of D‐galactose (Figure 2.2). The
GalA units are linked by ∞‐1/4 galacturonosyl linkages, which are usually interrupted by
ʟ‐rhamnose units bearing side‐chains, which cause a discontinuity in the linear structure
of the poly‐(GalA) chain. Some of the carboxyl groups on the continuous poly‐(GalA) chain
of pectin are esterified by methyl groups which generate variation in the degree of methyl
esterification (DE). This results in classification as high‐methoxyl (HM) pectins, in which
DE is greater than 50%, or low‐methoxyl (LM) pectins, being lower than 50%.
Homogalacturonans (HG), rhamnogalacturonan‐I (RGI), and rhamnogalacturonan‐II
(RGII) are the three main structural classes of pectin. The deposition and the way in which
these domains are joined to one another are still a matter of debate. Currently, it is thought
that the polysaccharide regions have covalent bonds and are ionically cross‐linked with
other pectin strands to form networks that branch throughout the primary cell walls.
The biosynthesis of pectin is known to be complex and our understanding is currently
characterized by much speculation (Chan et al. 2016; Adetunji et al. 2017). The diverse
properties of pectin at the microstructural and macromolecular levels form the basis for its
various food and nonfood applications, including reported health‐promoting benefits and
bioactivities. The major sources of commercial pectins are citrus wastes (pulp and peel:
85%), and apple pomace (14%), while some specific products may be extracted from sugar-
beet pulp (0.5%) (Ciriminna et al. 2015). This is due to the availability of these biomasses
and the quality presented by their pectins. Citrus peel and apple pomace are available in
large amounts as remainders from juice and essential oil production while sugarbeet pulp
is obtained from the sugar industry.
A schematic representation of the composition of the structural elements of cellulose,
hemicellulose, lignin, and pectin is given in Figure 2.2.
2.3.3 Proteins and Glycoproteins

There are two groups of cell wall proteins: enzymes and structural proteins. Cell wall
enzymes have a broad range of functions, including wall degradation, polymer turnover,
wall remodeling, and extension associated with plant growth, development, maturation,
and senescence (Waldron et al. 2003). The most abundant structural proteins are the
hydroxyproline‐rich glycoproteins which comprise several groups, including extensins,
arabinogalactan proteins, proline/hydroxyproline‐rich glycoproteins, and solanaceous lec-
tins (Cassab 1998; Sommer‐Knudsen et al. 1998). It has been proposed that they act as an
attachment for lignin and link pectic components (Waldron et al. 2003).
2.3.4 Phenolic Compounds

Two main classes of phenolic compounds are found in plant cell walls: lignin, the most
abundant polymer in secondary walls, and simple phenolic esters, such as ferulic acid,
which are attached to primary wall polysaccharides (Brett and Waldron 1996).
Lignin is the main component of lignocellulosic biomass, accounting for 15–30% by
weight and 40% by energy (Zhang 2018). Less than 2% of the 1.5–1.8 billion tons of indus-
trial lignin waste is used annually, of which lignosulfonate is mainly used as an additive in
building materials (Cao et al. 2018). The major source of lignin (kraft lignin and lignosul-
fonates) is the pulp and paper industry (Mahmood 2014). For the sustainable production of
biofuels and bulk chemicals, lignin is considered the second most abundant renewable
resource, after cellulose (Agarwal et al. 2018).
In plants, lignin embeds the polysaccharide network of cellulose and hemicellulose like
an adhesive that keeps the lignocellulose matrix together, providing strength, rigidity,
impermeability, and structural reinforcement to cell walls. The lignin content depends on
the plant taxonomy, being most abundant in conifer wood (30%), and 10–15% in grass. All
wall layers may accumulate phenolic polymers, namely lignins made of cytosol‐synthe-
sized hydroxycinnamoyl alcohols (Voxeur et al. 2015). Despite lignin monomer composi-
tion and linkage distribution being extremely variable between species and tissues, only
three types of substituted phenols connected by C─C or C─O bonds form the 3D irregular
polymer of lignin (Zhang 2018): p‐cumaryl alcohol, coniferyl alcohol, and synapyl alcohol
(noncondensate structure) (Figure 2.2) (Cao et al. 2018). Lignin can be classified as soft-
wood lignin, hardwood lignin, and grass lignin, depending on its origin. Softwood lignin is
formed by coniferyl alcohol and trace amounts of sinapyl alcohol‐derived units. Hardwood
lignin contains both coniferyl alcohol and sinapyl alcohol but in different ratios compared
to softwood lignin. Grass lignin contains mainly structural elements derived from
p‐coumaryl alcohol (Naseem et al. 2016).
Lignin can be separated from lignocellulose by physical, chemical, and biological meth-
ods (Hu et al. 2018). The physical method is to break the connection between lignin and
cellulose/hemicellulose structures by steam explosion or mechanical grinding (high tem-
perature and pressure), via which lignin with high purity can be obtained (Cao et al. 2018).
The biological method is carried out by enzymes under milder conditions, with the aim of
destroying the chemical connection between lignin and carbohydrate. Due to high separa-
tion efficiency and mild reaction conditions, chemical methods are widely used for indus-
trial production (Cao et al. 2018). Moreover, the production of fine chemical products from
lignin can help to reduce the consumption of fossil resources and constitutes one of the
environmentally friendly approaches of researchers.
Lignin depolymerization is the most complex, and its utilization for value‐added chemicals is
a severe challenge compared to cellulose and hemicelluloses. Despite being described as a ran-
dom construction of aromatic monomers, the scaling of lignin products is the limiting step to
obtain valuable chemicals such as phenols, aldehydes, carboxylic acids, alkanes, and arenes (Mei
et al. 2019). However, great effort should be directed to this because lignin is the most abundant
renewable natural aromatic biopolymer, which can be a sustainable candidate feedstock to pro-
duce aromatic chemicals and, in particular, to replace those ultimately derived from petroleum.
2.4 Valuable Chemicals Obtained from

Lignocellulosic Biomass
Lignocellulosic biomass is the most abundant renewable material in the world for the pro-
duction of biofuels (Cai et al. 2017). Bioethanol production via LG feedstock conversion
under a biorefinery system appears to be promising in mitigating global climate change
involving also the co‐production of biochemicals. Thus, in a biorefinery, a wide spectrum
of valuable chemical products can be obtained by combining biochemical conversion tech-
nologies. LG biomass from crops, paper residues, wood, and solid wastes constitutes a
potential sustainable resource for bio‐based fuels (Ekman et al. 2013).
As the first step in LG conversion to bioethanol, a pretreatment to remove contaminants
and reduce moisture is carried out. Dilute acid pretreatment followed by enzymatic hydrol-
ysis converts LG biomass to fermentable sugars. The fermentation of hexoses and pentoses
into bioproducts is then purified by physical treatments, such as distillation or filtration
(Borrion et al. 2012; Chrysikou et al. 2018). On the other hand, by using extraction and
processing methods, a broad range of functionalized molecules can be released from plants
waste: lipids, hemicellulose, bioactives/nutraceuticals, pectin, starch, phytochemicals,
phenols, nanoparticles, biodiesel, and activated carbon (Ravindran and Jaiswal 2016).
Phenols and phenolics Gluconic acid

Levulinic acid Lactic acid
LIGNIN Carboxylic acids
Arenes Organic acids
Formic acid
Polyols
Pyrulic acid
Alkanes Aldehydes CELLULOSE
Bioactive Lipids
Xylose
compounds
Flavonoids
Gums
Polysaccharides Dietary fibers Glucose
Furan
derivatives HEMICELLULOSE
PECTIN
Furfural Hydroxy- Biofuels
methylfurfural Xylan
Biofuels
Figure 2.3 Some valuable chemicals obtained from lignocellulosic biomass.
Figure 2.3 shows some of the most common value‐added products obtained from ligno-
cellulose sources, while a brief description of the most valuable products obtained from LG
biomass follows.
2.4.1 Sugar Alcohols

Producing water‐soluble carbohydrates from LG biomass requires cleaving ether bonds in
hemicellulose and cellulose chains. Also, the further degradation of the pentose and hex-
ose sugars (xylose, glucose) to insoluble products needs to be carefully inspected. Despite
being a high‐cost process, the hydrolysis of cellulose to glucose is known to be catalyzed by
cellulase enzymes under mild conditions. Mineral acids, such as H2SO4 and HCl, can also
catalyze the hydrolysis of cellulose to glucose. During the hydrolysis, the β‐1,4‐glycosidic
bonds in cellulose are selectively activated by the Brønsted acids in the aqueous medium.
Relatively high yields (30–70%) of glucose can be achieved by using concentrated H2SO4
(Deng et al. 2014). However, the problems of corrosion, nonrecyclability, and large amounts
of mineral acid wastes make this procedure unattractive. Lutherbacher et al. (2014) pro-
posed the use of biomass‐derived γ‐valerolactone (GVL) to promote the thermocatalytic
saccharification through complete solubilization of the biomass, including the lignin frac-
tion. The overall process, including pretreatment and enzymatic hydrolysis, could be cost‐
competitive for ethanol production. Oyola‐Rivera et al. (2018) recently reported sugar
yields up to 90%, 94%, and 88% for bagasse, plantain peel, and spent barley from brewery
production, respectively.
Among the different value‐added products obtained, xylose and glucose are the most
abundant in the deconstruction of hardwood and softwood (Lutherbacher et al. 2014).
These sugars can be converted to many platform molecules for the production of fuels and
specialty chemicals using heterogeneous catalysts.
2.4.2 Polyols
Due to its structure, kraft lignin formed by phenyl propanol and aryl‐alkyl ether bonding
can be a good source of polyols. The multiple hydroxyl groups present in the lignin’s struc-
ture are essential raw materials for polyurethane production. Also, for polyolefins, polyeth-
ylene terephthalate (PET) and polycarbonate production, the plastics can be either replaced
or enriched with bio‐based components (Brodin et al. 2017). Considering sustainability
concerns and the fact that petroleum products are commonly used in the polyurethane
industry, bio‐based polyols and lignopolyols could be an environmentally friendly solution
(Mahmood 2014). Although a bioplastic is characterized by being produced from a renew-
able source, bioplastics are not necessarily biodegradable. As an example, biopolyethylene
(BioPe) is similar to the fossil‐based polyethylene and thus is not biodegradable. Hence,
plastic biodegradability is determined by the chemical structure rather than origin (Brodin
et al. 2017).
2.4.3 Furfural
Furfural (FF), identified by the US Department of Energy (DoE) as one of the top 12 value‐
added products, has a word market of around 300 000 tons per year. FF is a typical product
which could be obtained from hemicellulose in raw biomass and is also a key platform
chemical produced in LG biorefineries. The main advantage of FF, as for other block chain
compounds, is that it could further be transformed to fuels and useful chemicals. A wide
range of products can be derived using FF as starter material, as it is an essential intermedi-
ate product in the oil refining, plastics, pharmaceutical, and agrochemical industries.
Biofuels can be derived from furfuryl acetate, GVL, levulinic acid, 1,5‐pentanediol, bicyclo-
pentane, and furfuryl alcohol resins (diesel/kerosene). On the other hand, as value‐added
products for the chemical industry, FF can lead to the following derivatives: furan, furfural
resins, tetrahydrofurfuryl alcohol, 2‐methyl tetrahydrofuran, 2‐methyl furan, furoic acid,
succinic acid, and 5‐hydroxy 2(5H)‐furanone, among others (Luo et al. 2019).
2.4.4 Hydroxymethylfurfural
Hydroxymethylfurfural (HMF) is considered a versatile key value‐added chemical (or
platform molecule) which receives much attention in the petroleum and chemical indus-
tries. HMF has a high market value up to USD 300 per kg, depending on the final chemical
quality. Currently, commercial production of HMF relies on syrups extracted from energy
crops (Yu and Tsang 2017). HMF has excellent chemical reactivity that enables the synthe-
sis of diverse value‐added chemical products. In this respect, HMF has been identified as
a primary building block for the production of furanic polyesters, polyamides, and
polyurethanes analogous to those derived from the petroleum polymer industry
(Pagán‐Torres et al. 2012).
Six‐ and five‐carbon carbohydrate derived from biomass need to be transformed into
intermediates before being used for biofuel or chemical production (Abou‐Yousef et al.
2013). The conversion of biomass‐derived carbohydrate into furan derivatives such as HMF
and FF is frequently the first step. However, the decomposition behavior of the feedstock
depends on the interactions between the cellulose, hemicellulose, and lignin. Thus, diverse
HMF yields (2–60%) depend on the substrate composition and conversion systems (Yu and
Tsang 2017). Among the carbohydrate sources employed for HMF preparation, fructose is
the most popular due to ease of conversion and high selectivity through a simple dehydra-
tion reaction. Glucose, cellulose, starch, sucrose, and inulin are also used as starting
substances for HMF production (Agarwal et al. 2018). Hydrogenation reactions result in
synthesis of the following furan derivatives: 2,5 dimethylfuran, which is a bioderived trans-
portation fuel, 2,5‐bis(hydroxymethyl)furan (BHMF) (used in manufacture of polyure-
thane foam), 2,5‐dimethyltetrahydrofuran (used in polyester preparation) and others. Also,
HMF acts as an oxidative precursor to prepare FDCA (2,5‐furandicarboxylic acid), an alter-
native intermediate product in PET manufacturing and nylon preparations (van Putten
et al. 2013). Also, 2,5‐diformylfuran (DFF) finds application in the synthesis of diamine
and Schiff bases (Agarwal et al. 2018).
The complexity of cellulose structure causes complete insolubility in aqueous and most
common organic solvents. However, in the presence of ionic solutions at suitable condi-
tions, the conversion of glucose to HMF takes place. This is due to the specific properties of
ionic liquids, such as very low vapor pressure, no flammability, high thermal and chemical
stability, and efficient solvent power for organic and inorganic substances. The solubility of
cellulose in ionic liquids is related to its anion, which can dissolve cellulose by disrupting
its hydrogen bonds. Thus, the conversion reaction of cellulose to produce HMF passes
through three consecutive steps: hydrolysis, isomerization, and dehydration (Abou‐Yousef
et al. 2013). The hydrolyzation to oligosaccharides is mediated by ionic acids and then to
glucose in a short time. Glucose should be isomerized into fructose followed by dehydra-
tion of fructose molecules. Conversion of lignocellulose in ionic liquids into the furanic
compounds HMF and FF has produced yields up to 52% and 31%, respectively (Zhang and
Zhao 2010).
2.4.5 Organic Acids

The high oxygen/carbon ratio feature of cellulose makes it an ideal precursor for the pro-
duction of organic acids (Li et al. 2018). Under suitable conditions, the lignin phenyl ring
can be oxidized to quinones or be cleaved. Because of the instability of benzoquinone struc-
tures obtained, they can be oxidized to yield aromatic ring cleavage products, such as dicar-
boxylic acids (Ma et al. 2018). The conversion of lignin to open chain organic acids has
become an area of interest in chemical research. Carboxylic acids also can be derived from
lignin co‐existing with other products, such as phenols and aldehydes (Mei et al. 2019).
The conversion of carbohydrates into organic acids such as levulinic, gluconic, and for-
mic acid is a promising way to use renewable biomass since it is more cost‐effective than
other biomass transformations, such as hydrogenation or hydrogenolysis of polyols, which

require high amounts of hydrogen resources. Lactic acid or 2‐hydroxypropionic acid is
another versatile α‐hydroxyl carboxylic acid with a variety of applications from food to
pharmaceutical industries. Moreover, it can serve as a key building block for production of
various chemical commodities.
Via oxidative dehydrogenation, pyruvic acid can be obtained and, with a dehydration
step, acryl acid. Esterification of lactic acid with alcohols gives rise to alkyl lactates used as
green solvents. Lactic acid can also be converted into 1,2‐propanediol and 2,3‐pentanedione
through hydrogenation and condensation/decarboxylation, respectively. Lactic acid from
biomass has been used for amino acid production since it has been employed for the syn-
thesis of high value‐added analine. In addition, lactic acid can be applied to synthesis of
polylactic acid that is considered as one of the most promising biodegradable plastics use-
ful in the geotextile industry, agricultural film, packaging, and thermoplastics for 3D
printing, among other applications (Brodin et al. 2017; Li et al. 2019).
2.4.6 Phenolic Compounds

Phenols are important platform chemicals widely used as biofuel and key precursors of
plastics, cosmetics, and pharmaceutical products (Rinaldi et al. 2016; Luo et al. 2019). The
structure of lignin is one of the key factors affecting its depolymerization into phenolic
monomers. For more than 20 years, researchers have been extending their understanding
of depolymerization processes in order to break down the complex biopolymer matrix into
monophenolic compounds (Sun et al. 1995; Chen et al. 2018) but it is still a huge challenge.
One of the main problems in the production of phenolic monomers is the repolymerization
reaction, with a bigger impact in the final yields. Also, the higher the molecular weight of
lignin, the lower the depolymerization yields.
2.4.7 Aldehydes and Ketones

One of the most extended routes into the valorization of lignin involves its oxidative depo-
lymerization into highly functionalized monomers (Gomes et al. 2018). Among them, van-
illin, widely used as a flavor compound in food, constitutes one of the major molecular
compounds manufactured on an industrial scale from biomass. Vanillin has broad applica-
tions in our daily life such as food flavoring and perfume, fine chemistry, and pharmaceu-
ticals. Annual vanillin demand is growing by 2–4% in Europe and the USA, being 10% in
China. Since the global demand reaches 16 000 tons, novel economical methods for pro-
ducing vanillin are of great interest (Luo et al. 2016).
The production of vanilla via lignin depolymerization has attracted interest due to the low
cost, high reserve and unique aromatic structure of lignin (Agarwal et al. 2018). It is esti-
mated that over 15% of the industrial production of vanillin is derived from lignin but the
vast majority is currently synthesized from a raw material from the petrochemical industry
called guaiacol. Guaiacol‐based processes are relatively cost‐efficient, whereas lignin‐based
processes are sustainable due to their widely available biomass resource (mainly paper fac-
tories). However, vanillin separation is more complex and cost‐ineffective from the lignin
oxidation process. Huge amounts of caustic soda wastes (160 kg/kg vanillin) are generated
during this process, with the additional environment concerns. Moreover, the price of
lignin‐based vanillin is consistently higher than that of guaiacol‐based vanillin (Luo
et al. 2016).
Although vanillin is the star product derived from lignocellulose, other aldehydes and
ketones are of interest to the chemical, pharmaceutical, and food industries. Many research-
ers are focusing on the most efficient techniques to obtain high‐purity monomers from
lignocellulose. Recently, Gomes et al. (2018) reported a successful fractionation method for
alkali lignin mixtures by adsorption with SP700 resin based on a two‐eluent technique with
deionized water and ethanol. Several families of compounds, namely the acids, aldehydes
and ketones, were recovered and quantified: p‐hydroxybenzaldehyde, vanillic acid, vanil-
lin, syringic acid, syringaldehyde, acetovanillone, and acetosyringone.
Mansur et al. (2013) studied the recovering of ketones as high‐value chemicals from
woody biomass for char production, by using the pyroligneous acid (a by‐product from
slow pyrolysis) in water.
So, lignin has great potential to be employed as starting material for the production of
value‐added materials such as vanilla and others, reducing the utilization of petrochemical
derivatives.
2.4.8 Bioactive Compounds

Considering that the global natural antioxidants market is expected to be over USD 4 billion
by 2022 (Lauberts et al. 2017), the utilization of lignin to obtain antioxidant compounds is
a prominent focus of research. As a polyphenol source, lignin naturally possesses a certain
degree of antioxidant activity (Zhao et al. 2018).
Moreover, fruit processing waste, that represents more than 0.5 billion tons of waste
worldwide, is an incalculable source of bioactive compounds. A novel biorefinery approach
could aim to produce valuable chemicals from fruit processing waste. The peel, pomace
and seed fraction of fruits and vegetables could potentially contain bioactive compounds as
diverse as pectin, lipids, flavonoids, and dietary fibers. Seeds are rich in proteins and also
can contain bioactive lipids and polyphenols while peels are a rich source of dietary fibers
(Banerjee et al. 2017). Bioactive peptides obtained by hydrolysis of proteins were found to
exhibit various pharmacologic properties ranging from antihypertensive to antiinflamma-
tory (Shavandi et al. 2018). Additionally, since bioactive polysaccharides show a range of
bioactivities such as anticancer and antiinflammatory effects, bioactive polysaccharide
research has become an area of substantial interest (Bernaerts et al. 2018).
With respect to hemicellulose, its structure comprises xyloglucans, xylans, mannans, and
glucomannans (Figure 2.2). Xylan upon controlled hydrolysis produces xylooligosaccha-
rides (XOS) which are oligomers of β‐1,4‐linked xylose residues with various constituents
including acetyl, phenolic, and uronic acids (Banerjee et al. 2017). Oligosaccharides such as
XOS belong to a class of dietary fibers. The dietary fibers have aroused great interest because
they reduce the concentration of cholesterol in blood, glucose levels after meals, and the
response to insulin. These effects are mainly due to a natural fiber component known as
β‐D‐glucan, present in oats and barley husks. Along with XOS, inulin, fructooligosaccha-
rides (FOS), galactooligosaccharides (GOS), and maltooligosaccharides (MOS) represent a
class of nonstarch polysaccharide‐based dietary fibers. Fermentation of oligosaccharides
produces short‐chain fatty acids, which provide a range of health benefits (gut and
microbiota health, antioxidant and antitumor activity, metabolic control of glucose and
lipid profile and immunomodulation) (Chimtong et al. 2016; Mohanty et al. 2018;
Katsimpouras et al. 2018). Xylan, an important constituent of hemicellulose, can serve
in medical applications such as drug delivery, inflammatory bowel disease, hydrogels,
and tissue engineering (Banerjee et al. 2017).
2.4.9 Gums and Hydrocolloids

Previously, the potential of LG biomass has been suggested to provide highly useful ingre-
dients for the medical and food industries. Cellulose and its derivatives are used in diagnos-
tic and biomedical applications such as filler in solid dosage formulations, component
immobilization (Li et al. 2018), drug delivery systems (Gopinath et al. 2018; Jacob et al.
2018), and hydrogels (Huang et al. 2018), among others, which exploit their water retention
and biodegradability properties (Lin and Dufresne 2014). In the pharmaceutical industry,
polysaccharides find application as polymers. Chitosan and carrageenan are typical prod-
ucts utilized as edible films, thickening and emulsifying agents, and cosmetic additives
(Elsabee and Abdou 2013). For food applications, a large variety of soluble and insoluble
gums are obtained from cellulose, which are frequently used as ingredients in innumerable
foodstuffs. Among them, microcrystalline cellulose (MCC), carboxylmethylcellulose
(CMC), methylcellulose (MC), and hydroxypropyl methylcellulose (HPMC) have special
properties; they are used as aroma conveyors, antifouling agents (MCC), stabilization of
foams, emulsions, protein dispersions, pectin gels and starch against heat (colloidal MCC),
improving adhesion, replacing fats and oils, controling the growth of ice crystals in ice
cream, surfactants, fat replacers and fat absorption reducers in fried foods (MC and HPMC)
(Tanti et al. 2016; Luo et al. 2017; Oh and Lee 2018).
2.4.10 Pectins
Food sectors such as dairy, bakery, nutraceutical and functional foods (Rababah et al. 2015;
Schmidt et al. 2015; Zhuang et al. 2015; Li and Nie 2016) as well as pharmaceutical domains
like drug delivery (Liu et al. 2007) are interested in other polysaccharides from cell walls,
like pectins. Among the most studied functional and physicochemical properties of pectin
in different conventional and unconventional sources are water affinity properties (viscos-
ity increase and gelification) (Sousa et al. 2015; Hosseini et al. 2016; Nascimento et al. 2016;
Chan et al. 2017; Rasidek et al. 2018), surface properties such as emulsifying and emulsion
stabilizing (Ngouémazong et al. 2015) and elaboration of edible biodegradable films and
coatings (Oliveira et al. 2016; Penhasi 2017; Moreira et al. 2016; Rossi Márquez et al. 2017;
Muñoz‐Labrador et al. 2018). Health‐promoting benefits have also been reported: reduc-
tion of postprandial glycemic responses, maintenance of normal blood cholesterol concen-
trations, and antioxidant and antiinflammatory functions. It has been reported that pectin
with a low degree of esterification can inhibit tumor growth, induce apoptosis, suppress
metastasis, and modulate immune responses (Katzenmaier et al. 2014; Louis et al. 2014;
Almeida et al. 2015; Tan et al. 2018). Table 2.1 summarizes some interesting research about
chemical platform compounds, lignocellulosic sources, and feedstock of plant materials.
Table 2.1 High‐value chemicals obtained from lignocellulosic biomass.
Chemical compounds Lignocellulosic source/ feedstock References
Furfural/hydroxy‐methylfurfural Hemicellulose/Pinus pinaster Peleteiro et al.

(2016)
Organic acids (levulinic acid) Cellulose/corncob residue Li et al. (2014)
Glucose Cellulose/sugarcane bagasse Dussan et al.
(2014)
Xylan Hemicellulose/sugarcane Vallejos et al.
bagasse (2015)
Polyols Lignin/residues of oil palm Faris et al. (2015)
Furfural Hemicellulose/corncobs Deng et al.
(2015a)
Furfural Hemicellulose/Eucalyptus Peleteiro et al.
globulus wood (2016)
Pectin (high methoxyl) Pectic polysaccharides/ Petkowicz et al.
watermelon (2017)
Pectin (low methoxyl) Pectic polysaccharides/sunflower Sobol et al.
seed head (2017)
Phenolics Lignin/sugarcane bagasse Pinheiro et al.
(2017)
Arenes compounds Lignin/abedul Shao et al. (2017)
Carboxylic acids and aromatics Lignin/corn residue Lyu et al. (2018)
compounds
Aldehydes compounds Lignin/oil palm leaves Hazwan‐Hussin
et al. (2018)
Furfural/xylose/glucose Biomass/corn stover, hard‐ and Lutherbacher
softwood et al. (2014)
Sugar alcohols Biomass/bagasse, plantain peel Oyola‐Rivera
and barley et al. (2018)
Furan derivatives Cellulose Abou‐Yousef
et al. (2013)
Phenolics Lignin Luo et al. (2019)
Organic acids Cellulose Li et al. (2018)
Phenolics Lignin Cheng et al.
(2012)
Phenolics Lignin/black liquor (paper Chen et al.
industry) (2018)
Aldehydes (vanillin/ Lignin/wheat straw Sun et al. (1995)
syringaldehyde)
Aldehydes (vanillic acid/vanillin/ Lignin/tobacco stalks Gomes et al.
syringic acids/syringaldehyde) (2018)
Aldehydes (vanillin) Lignin Luo et al. (2016)
Ketones Pyroligneus acid/woody biomass Mansur et al.
(2013)
Polysaccharides Microalgal biomasses Bernaerts et al.
(2018)
2.5 Conclusions and Final Remarks
Concern about global warming is promoting the replacement of carbon or fossil products
as well as the study of sustainable alternatives for the treatment, processing, extraction and
conversion of lignocellulose into monomers or intermediate compounds useful for the
chemical, pharmacologic, biomedical, and food industries. Every day, new ways of obtain-
ing platform chemicals are discovered, their sources being more diverse and original. In
this chapter, a wide range of highly valuable products derived from lignocellulosic biomass
were reviewed. Examples of chemical platforms for several industries and some of their
applications were discussed. The reuse of by‐products and the revalorization of waste are
emerging as part of the solution to meet the ecological problems facing the planet.
Bioutilization and conversion of agro‐industrial waste into useful products constitutes a
superior and eco‐friendly strategy for proper waste management.
The future offers even more promise. As an example, we need technology for the better
exploitation of LG biomass based on the cost‐effective production of important industrial
enzymes, like cellulases and hemicellulases, through fermentation of agro‐industrial waste
from microorganisms such as fungi. The importance of enzyme production also concerns
their utilization for the production of fuel ethanol from LG biomass through cellulose
hydrolysis.
Another aspect to consider is that recycling nutrient‐rich sources could reduce the
organic waste sent to landfill while producing specialty chemical products. In addition,
some chemical compounds present in by‐products, such as coffee by‐products, are of
eco‐toxicologic concern, as in the case of tannins, caffeine, and chlorogenic acid. These
compounds are thought to be harmful to aquatic organisms and also have negative
effects on seed germination and plant growth. Bioremediation for detoxifying coffee
by‐products can produce instead other valuable products such as enzymes. Thus, suc-
cessive steps of treatment or processing driven by environmental concern can lead to
the discovery of high added‐value products from LG biomass sources, not taken into
account previously.
Related to biomaterials, the field of biodegradable bioplastics for the replacement of
packaging is one of the most promising areas for ecologic research. However, to develop
efficient products that completely replace the nonbiodegradable products that we con-
sume daily will be a challenge. The bioplastic market represents a small percentage of
the total plastics market but it constitutes a future promising area for research and
development. Polylactic acid is one of the most promising elements for creating bio‐
based plastics. Its characteristics make it biocompatible with the human body and also
it may be reabsorbed. Other classes of bio‐based polyesters, such as polyhydroxyal-
kanoates, can be produced via biotechnologic routes by applying different microorgan-
isms. Depending on the processing conditions, the carbon source, the synthesis
microorganisms and the use of additives, a variety of polymers can be obtained with
different properties and characteristics, which increases the field of research in the
biotechnologic area. At the same time, the replacement of products which are deriva-
tives of the oil industry (nonrenewables, toxics) is also of vital importance. Based on
this need, one current research area is the ecologic wood adhesives coming from ligno-
cellulosic materials.
Also, new trends in biotechnology research aspire to modify specific plant targets in
order to obtain a lignocellulosic biomass according to our requirements. Plants can be
designed by engineering to improve their structures or genetic pathways and thus enhance
biomass composition.
References
Abou‐Yousef, H., Hassan, E.B., and Steele, P. (2013). Rapid conversion of cellulose to 5‐
hydroxymethylfurfural using single and combined metal chloride catalysts in ionic liquid.
Journal of Fuel Chemistry and Technology 41 (2): 214–222.
Adetunji, L.R., Adekunle, A., Orsat, V., and Raghavan, V. (2017). Advances in the pectin
production process using novel extraction techniques: a review. Food Hydrocolloids
62: 239–250.
Agarwal, B., Kailasam, K., Sangwan, R.S., and Elumalai, S. (2018). Traversing the history of
solid catalysts for heterogeneous synthesis of 5‐hydroxymethylfurfural from carbohydrate
sugars: a review. Renewable and Sustainable Energy Review 82: 2408–2425.
Alexandridis, P., Ghasemi, M., Furlani, E.P., and Tsianou, M. (2018). Solvent processing of
cellulose for effective bioresource utilization. Current Opinion in Green and Sustainable
Chemistry 14: 40–52.
Almeida, E.A., Facchi, S.P., Martins, A.F. et al. (2015). Synthesis and characterization of pectin
derivative with antitumor property against Caco‐2 colon cancer cells. Carbohydrate Polymers
115: 139–145.
Altartouri, B. and Geitmann, A. (2015). Understanding plant cell morphogenesis requires
real‐time monitoring of cell wall polymers. Current Opinion in Plant Biology 23: 76–82.
Anwar, Z., Gulfraz, M., and Irshad, M. (2014). Agro‐industrial lignocellulosic biomass a key to
unlock the future bio‐energy: a brief review. Journal of Radiation Research and Applied
Sciences 7 (2): 163–173.
Asgher, M., Ahmad, Z., and HMN, I. (2013). Alkali and enzymatic delignification of sugarcane
bagasse to expose cellulose polymers for saccharification and bio‐ethanol production.
Industrial Crops and Products 44: 488–495.
Banerjee, J., Singh, R., Vijayaraghavan, R. et al. (2017). Bioactives from fruit processing wastes:
green approaches to valuable chemicals. Food Chemistry 225: 10–22.
Bernaerts, T.M.M., Gheysen, L., Kyomugasho, C. et al. (2018). Comparison of microalgal
biomasses as functional food ingredients: focus on the composition of cell wall related
polysaccharides. Algal Research 32: 150–161.
Borrion, A.L., McManus, M.C., and Hammond, G.P. (2012). Environmental life cycle
assessment of bioethanol production from wheat straw. Biomass Bioenergy 47: 9–19.
Brett, C.T. and Waldron, K.W. (1996). Physiology and biochemistry of plant cell walls. In:
Topics in Plant Physiology, vol. 2, 114–256. The Netherlands: Springer.
Brodin, M., Vallejos, M., Opedal, N.T. et al. (2017). Lignocellulosics as sustainable resources for
production of bioplastics – a review. Journal of Cleaner Production 162: 646–664.
Cai, J., He, Y., Yu, X. et al. (2017). Review of physicochemical properties and analytical
characterization of lignocellulosic biomass. Renewable and Sustainable Energy Reviews 76:
309–322.
Cao, L., Yu, I.K.M., Liu, Y. et al. (2018). Lignin valorization for the production of renewable
chemicals: state‐of‐the‐art review and future prospects. Bioresource Technology 269: 465–475.
Carpita, N. and McCann, M. (2000). The cell wall. In: Biochemistry and Molecular Biology of
Plants (eds. B.B. Buchanan, W. Gruissem and R.L. Jones), 52–108. Rockville, MD: American
Society of Plant Physiologists.
Cassab, G.I. (1998). Plant cell wall proteins. Annual Review of Plant Physiology and Plant
Molecular Biology 49: 281–309.
Chan, S.Y., Choo, W.S., Young, D.J., and Loh, X.J. (2017). Pectin as a rheology modifier: origin,
structure, commercial production and rheology. Carbohydrates Polymeres 161: 118–139.
Chen, C., Jin, D., Ouyang, X. et al. (2018). Effect of structural characteristics on the
depolymerization of lignin into phenolic monomers. Fuel 223: 366–372.
Cheng, S., Wilks, C., Yuan, Z. et al. (2012). Hydrothermal degradation of alkali lignin to bio‐
phenolic compounds in sub/supercritical ethanol and water–ethanol co‐solvent. Polymer
Degradation and Stability 97: 839–848.
Chimtong, S., Saenphoom, P., Karageat, N., and Somtua, S. (2016). Oligosaccharide production
from agricultural residues by non‐starch polysaccharide degrading enzymes and their
prebiotic properties. Agriculture and Agricultural Science Procedia 11: 131–136.
Chrysikou, L.P., Bezergianni, S., and Kiparissides, C. (2018). Environmental analysis of a
lignocellulosic‐based biorefinery producing bioethanol and high‐added value chemicals.
Sustainable Energy Technologies and Assessments 28: 103–109.
Ciriminna, R., Chavarría‐Hernández, N., Inés Rodríguez Hernández, A., and Pagliaro, M.
(2015). Pectin: a new perspective from the biorefinery standpoint. Biofuels, Bioproducts and
Biorefining 9 (4): 368–377.
Davison, B.H., Park, J., Davis, M.F., and Donohoe, B.S. (2013). Plant cell walls: basics of
structure, chemistry, accessibility and the influence on conversion. In: Aqueous Pretreatment
of Plant Biomass for Biological and Chemical Conversion to Fuels and Chemicals (ed. C.E.
Wyman), 23–38. UK: Wiley.
De‐Bhowmick, G., Sarmah, A.K., and Sen, R. (2018). Lignocellulosic biorefinery as a model for
sustainable development of biofuels and value added products. Bioresource Technology 247:
1144–1154.
Deng, A., Ren, J., Li, H. et al. (2015a). Corncob lignocellulose for the production of furfural by
hydrothermal pretreatments and heterogeneous catalytic process. RSC Advances 5:
60264–60272.
Deng, W., Zhang, Q., and Wang, Y. (2015b). Catalytic transformation of cellulose and its
derived carbohydrates into chemicals involving C‐C bond cleavage. Journal of Energy
Chemistry 24 (5): 595–607.
Dhyani, V. and Bhaskar, T. (2018). A comprehensive review on the pyrolysis of lignocellulosic
biomass. Renewable Energy 129: 695–716.
Dussan, K.J., Silva, D.D.V., Moraes, E.J.C. et al. (2014). Dilute‐acid hydrolysis of cellulose to
glucose from sugarcane bagasse. Chemical Engineering Transactions 38: 433–438.
Ekman, A., Wallberg, O., Joelsson, E., and Börjesson, P. (2013). Possibilities for sustainable
biorefineries based on agricultural residues – a case study of potential straw‐based ethanol
production in Sweden. Applied Energy 102: 299–308.
Elsabee, M.Z. and Abdou, E.S. (2013). Chitosan based edible films and coatings: a review.
Materials Science and Engineering C 33 (4): 1819–1841.
Faris, A.H., Mohamad‐Ibrahim, M.N., Abdul‐Rahim, A. et al. (2015). Preparation and

characterization of lignin polyols from the residues of oil palm empty fruit bunch. Bio
Resources 10 (4): 7339–7352.
Gomes, E.D., Mota, M.I., and Rodrigues, A.E. (2018). Fractionation of acids, ketones and
aldehydes from alkaline lignin oxidation solution with SP700 resin. Separation and
Purification Technology 194: 256–264.
Gopinath, V., Saravanan, S., Al‐Maleki, A.R. et al. (2018). A review of natural polysaccharides
for drug delivery applications: special focus on cellulose, starch and glycogen. Biomedicine
and Pharmacotherapy 107: 96–108.
Harris, P.J. (2006). Primary and secondary plant cell walls: a comparative overview. New
Zealand Journal of Forestry Science 36 (1): 36–53.
Hassan, S.S., Williams, G.A., and Jaiswal, A.K. (2019). Lignocellulosic biorefineries in Europe:
current state and prospects. Trends in Biotechnology 37 (3): 231–234.
Hazwan‐Hussin, M., Samad, N.A., Latif, N.H.A. et al. (2018). Production of oil palm (Elaeis
guineensis) fronds lignin‐derived non‐toxic aldehyde for eco‐friendly wood adhesive.
International Journal of Biological Macromolecules 113: 1266–1272.
Hosseini, S.S., Khodaiyan, F., and Yarmand, M.S. (2016). Optimization of microwave assisted
extraction of pectin from sour orange peel and its physicochemical properties. Carbohydrate
Polymers 140: 59–65.
Hu, J., Zhang, Q., and Lee, D.J. (2018). Kraft lignin biorefinery: a perspective. Bioresource
Technology 247: 1181–1183.
Huang, X., Xie, F., and Xiong, X. (2018). Surface‐modified microcrystalline cellulose for
reinforcement of chitosan film. Carbohydrate Polymers 201: 367–373.
Isikgor, F.H. and Becer, C.R. (2015). Lignocellulosic biomass: a sustainable platform for the
production of bio‐based chemicals and polymers. Polymer Chemistry 25: 4497–4559.
Jacob, J., Haponiuk, J.T., Thoma, S., and Gopi, S. (2018). Biopolymer based nanomaterials in
drug delivery systems: a review. Material Today Chemistry 9: 43–55.
Jarvis, M.C. (2011). Plant cell walls: Supramolecular assemblies. Food Hydrocolloids 25 (2):
257–262.
Kapu, N.S. and Trajano, H.L. (2014). Review of hemicellulose hydrolysis in softwoods and
bamboo. Biofuels, Bioproducts and Biorefining 8: 857–870.
Katsimpouras, C., Dedes, G., Bistis, P. et al. (2018). Acetone/water oxidation of corn Stover for
the production of bioethanol and prebiotic oligosaccharides. Bioresource Technolology 270:
208–215.
Katzenmaier, E.M., André, S., Kopitz, J., and Gabius, H.J. (2014). Impact of sodium butyrate on
the network of adhesion/growth‐regulatory galectins in human colon cancer in vitro.
Anticancer Research 34 (10): 5429–5438.
Lauberts, M., Sevastyanova, O., Ponomarenko, J. et al. (2017). Fractionation of technical lignin
with ionic liquids as a method for improving purity and antioxidant activity. Industrial Crops
and Products 95: 512–520.
Li, J., Jiang, Z., Hu, L., and Hu, C. (2014). Selective conversion of cellulose in corncob residue
to levulinic acid in an aluminum trichloride‐sodium chloride system. Chem Sus Chem 7 (9):
2482–2488.
Li, J.M. and Nie, S.P. (2016). The functional and nutritional aspects of hydrocolloids in foods.
Food Hydrocolloids 53: 46–61.
Li, L., Wang, F., Shao, Z. et al. (2018). Chitosan and carboxymethyl cellulose‐multilayered
magnetic fluorescent systems for reversible protein immobilization. Carbohydrate Polymers
201: 357–366.
Li, S., Deng, W., Li, Y. et al. (2019). Catalytic conversion of cellulose‐based biomass and
glycerol to lactic acid. Journal of Energy Chemistry 32: 138–151.
Lin, N. and Dufresne, A. (2014). Nanocellulose in biomedicine: current status and future
prospect. European Polymer Journal 59: 302–325.
Liu, L.S., Fishman, M.L., and Hicks, K.B. (2007). Pectin in controlled drug delivery‐a review.
Cellulose 14 (1): 15–24.
Louis, P., Hold, G.L., and Flint, H.J. (2014). The gut microbiota, bacterial metabolites and
colorectal cancer. Nature Reviews Microbiology 12 (10): 661–672.
Luo, J., Melissa, P., Zhao, W. et al. (2016). Selective lignin oxidation towards vanillin in phenol
media. Chemistry Select 1 (15): 4596–4601.
Luo, N., Wang, M., Li, H. et al. (2017). Visible‐light‐driven self‐hydrogen transfer hydrogenolysis
of lignin models and extracts into phenolic products. ACS Catalysis 7 (7): 4571–4580.
Luo, Y., Li, Z., Li, X. et al. (2019). The production of furfural directly from hemicellulose in
lignocellulosic biomass: a review. Catalysis Today 319: 14–24.
Lutherbacher, J.S., Rand, J.M., Alonso, D.M. et al. (2014). Nonenzimatic sugar production from
biomass using biomass‐derived gamma‐valerolactone. Science 343: 277–280.
Lyu, G., Yoo, C.G., and Pan, X. (2018). Alkaline oxidative cracking for effective
depolymerization of biorefining lignin to mono‐aromatic compounds and organic acids with
molecular oxygen. Biomass Bioenergy 108: 7–14.
Ma, R., Guo, M., and Zhang, X. (2018). Oxidative valorization of lignin. In: Lignin Valorization:
Emerging Approaches, Energy and Environment Series (ed. G.T. Beckham), 128–158. Royal
Society of Chemistry.
Mahmood, N. (2014). Production of polyols via direct hydrolysis of Kraft lignin: optimization
of process parameters. Journal of Science & Technology for Forest Products and Processes 4
(2): 44–51.
Mansur, D., Yoshikawa, T., Norinaga, K. et al. (2013). Production of ketones from pyroligneous
acid of woody biomass pyrolysis over an iron‐oxide catalyst. Fuel 103: 130–134.
McCann, M.C. and Carpita, N.C. (2008). Designing the deconstruction of plant cell walls.
Current Opinion in Plant Biology 11: 314–320.
Mei, Q., Shen, X., Liu, H., and Han, B. (2019). Selectively transform lignin into value‐added
chemicals. Chinese Chemical Letters 30 (1): 15–24.
Mohanty, D., Misra, S., Mohapatra, S., and Sahu, P.S. (2018). Prebiotics and synbiotics: recent
concepts in nutrition. Food Bioscience 26: 152–160.
Moreira, M.R., Álvarez, M.V., Martín‐Belloso, O., and Soliva‐Fortuny, R. (2016). Effects of
pulsed light treatments and pectin edible coatings on the quality of fresh‐cut apples: a
hurdle technology approach. Journal of the Science of Food and Agriculture 97 (1): 261–268.
Muñoz‐Labrador, A., Moreno, R., Villamiel, M., and Montilla, A. (2018). Preparation of citrus
pectin gels by power ultrasound and its application as edible coating in strawberries. Journal
of the Science of Food and Agriculture 98 (13): 4866–4875.
Nascimento, G.E., Simas‐Tosin, F.F., Iacomini, M. et al. (2016). Rheological behavior of high
methoxyl pectin from the pulp of tamarillo fruit (Solanum betaceum). Carbohydrate
Polymers 139: 125–130.
Naseem, A., Tabasum, S., Zia, K.M. et al. (2016). Lignin‐de‐ rivatives based polymers, blends
and composites: a review. International Journal of Biological Macromolecules 93: 296–313.
Ngouemazong, E.D., Christiaens, S., Shpigelman, A. et al. (2015). The emulsifying and
emulsion‐stabilizing properties of pectin: a review. Comprehensive Reviews in Food Science
and Food Safety 14: 705–718.
Ofori‐Boateng, C. and Lee, K.T. (2013). Sustainable utilization of oil palm wastes for bioactive
phytochemicals for the benefit of the oil palm and nutraceutical industries. Phytochemistry
Reviews 12 (1): 173–190.
Oh, I.K. and Lee, S. (2018). Utilization of foam structured hydroxypropyl methylcellulose for
oleogels and their application as a solid fat replacer in muffins. Food Hydrocolloids 77:
796–802.
Oliveira, T.S., Zea‐Redondo, L., Moates, G.K. et al. (2016). Pomegranate peel pectin films as
affected by montmorillonite. Food Chemistry 198: 107–112.
Oyola‐Rivera, O., González‐Rosario, A.M., and Cardona‐Martínez, N. (2018). Catalytic
production of sugars and lignin from lignocellulosic biomass wastes using dilute sulfuric
acid in gamma‐valerolactone. Biomass Bioenergy 119: 284–292.
Pagán‐Torres, Y.J., Wang, T., Gallo, J.M.R. et al. (2012). Production of 5‐hydroxymethylfurfural
from glucose using a combination of Lewis and brønsted acid catalysts in water in a biphasic
reactor with an alkylphenol solvent. ACS Catalysis 2 (6): 930–934.
Pauly, M. and Keegstra, K. (2008). Cell‐wall carbohydrates and their modification as a resource
for biofuels. Plant Journal 54 (4): 559–568.
Peleteiro, S., Santos, V., Garrote, G., and Parajo, J.C. (2016). Furfural production from
eucalyptus wood using acid ionic liquid. Carbohydrate Polymers 146: 20–25.
Penhasi, A. (2017). Preparation and characterization of in situ ionic cross‐linked pectin films:
II. Biodegradation and drug diffusion. Carbohydrate Polymers 157 (10): 651–659.
Petkowicz, C.L.O., Vriesmann, L.C., and Williams, P.A. (2017). Pectins from food waste
extraction, characterization and properties of watermelon rind pectin. Food Hydrocolloids
65: 57–67.
Pinheiro, F.G.C., Soares, A.K.L., Santaella, S.T. et al. (2017). Optimization of the extraction
with acetosolv of lignin from sugar cane bagasse for the production of phenolic resin.
Rababah, T.M., Alúdatt, M.H., and Brewer, S. (2015). Jam processing and impact on
composition of active compounds. In: Processing and Impact on Active Components in Food,
1e (ed. P. Victor), 681–687. USA: Academic Press.
Rasidek, M., Azwani, N., Firdhaus, M. et al. (2018). Rheological flow models of banana peel
pectin jellies as affected by sugar concentration. International Journal of Food Properties 21:
2087–2099.
Ravindran, R. and Jaiswal, A.K. (2016). Exploitation of food industry waste for high‐value
products. Trends in Biotechnololy 34 (1): 58–69.
Rinaldi, R., Jastrzebski, R., Clough, M.T. et al. (2016). Paving the way for lignin valorisation:
recent advances in bioengineering, biorefining and catalysis. Angewandte Chemie‐Int. Ed. 55
(29): 8164–8215.
Rossi Márquez, G., di Pierro, P., Mariniello, L. et al. (2017). Fresh‐cut fruit and vegetable
coatings by transglutaminase crosslinked whey protein/pectin edible films. Food Science and
Sanderson, K. (2011). A chewy problem lignocellulose. Nature 474: S12–S14.

Schmidt, U.S., Koch, L., Rentschler, C. et al. (2015). Effect of molecular weight reduction,
acetylation and esterification on the emulsification properties of citrus pectin. Food
Biophysics 10: 217–227.
Shao, Y., Xia, Q., Dong, L. et al. (2017). Selective production of arenes via direct lignin
upgrading over a niobium‐based catalyst. Nature Communications 8: 16104.
Shavandi, A., Bekhit, A.E.D.A., Saeedi, P. et al. (2018). Polyphenol uses in biomaterials
engineering. Biomaterials 167: 91–106.
Sobol, I.V., Donchenko, L.V., Rodionova, L.Y. et al. (2017). Peculiarities of analytical
characteristics of pectins extracted from sunflower hearts. Asia Journal of Pharmaceutics 11
(1): S97–S100.
Sommer‐Knudsen, J., Bacic, A., and Clarke, A.E. (1998). Hydroxyproline‐rich plant
glycoproteins. Phytochemistry 47: 483–497.
Sousa, A.G., Nielsen, H.L., Armagan, I. et al. (2015). The impact of rhamnogalacturonan‐I side
chain monosaccharides on the rheological properties of citrus pectin. Food Hydrocolloids 47:
130–139.
Sun, R., Lawther, J.M., and Banks, W.B. (1995). The effect of alkaline nitrobenzene oxidation
conditions on the yield and components of phenolic monomers in wheat straw lignin and
compared to cupric(II) oxidation. Industrial Crops and Products 4 (4): 241–254.
Tan, H., Chen, W., Liu, Q. et al. (2018). Pectin oligosaccharides ameliorate colon cancer by
regulating oxidative stressand inflammation‐activated signaling pathways. Frontiers in
Immunology 9: 1504.
Tanti, R., Barbut, S., and Marangoni, A.G. (2016). Hydroxypropyl methylcellulose and
methylcellulose structured oil as a replacement for shortening in sandwich cookie creams.
Food Hydrocolloids 61: 329–337.
Vallejos, M.E., Felissia, F.E., Kruyeniski, J., and Area, M.C. (2015). Kinetic study of the
extraction of hemicellulosic carbohydrates from sugarcane bagasse by hot water treatment.
van Putten, R.J., van der Waal, J.C., de Jong, E. et al. (2013). Hydroxymethylfurfural, a versatile
platform chemical made from renewable resources. Chemical Reviews 113: 1499–1597.
Vassilev, S.V., Baxter, D., Andersen, L.K. et al. (2012). An overview of the organic and inorganic
phase composition of biomass. Fuel 94: 1–33.
Voxeur, A., Wang, Y., and Sibout, R. (2015). Lignification: different mechanisms for a versatile
polymer. Currunt Opinion in Plant Biolology 23: 83–90.
Waldron, K.W., Parker, M.L., and Smith, A.C. (2003). Plant cell walls and food quality.
Comprehensive Reviews in Food Science and Food Safety 2 (4): 128–146.
Yanakieva, I., Kussovski, V., and Kratchanova, M. (2012). Isolation, characterization and
modification of citrus pectins. Journal of BioScience and Biotechnology 1 (3): 223–233.
Yu, I.K.M. and Tsang, D.C.W. (2017). Conversion of biomass to hydroxymethylfurfural: a
review of catalytic systems and underlying mechanisms. Bioresource Technolology 238:
716–732.
Zamil, M.S. and Geitmann, A. (2017). The middle lamella‐more than a glue. Physical Biology
14 (1): 015004.
Zhang, J. (2018). Catalytic transfer hydrogenolysis as an efficient route in cleavage of lignin
and model compounds. Green Energy and Environment 3 (4): 328–334.
Zhang, Z. and Zhao, Z.K. (2010). Microwave‐assisted conversion of lignocellulosic biomass

into furans in ionic liquid. Bioresource Technology 101 (3): 1111–1114.
Zhao, L., Ouyang, X., Ma, G. et al. (2018). Improving antioxidant activity of lignin by
hydrogenolysis. Industrial Crops and Products 125: 228–235.
Zhong, R. and Ye, Z.H. (2015). Secondary cell walls: biosynthesis, patterned deposition and
transcriptional regulation. Plant Cell Physiology 56 (2): 195–214.
Zhuang, Y., Sterr, J., Kulozik, U., and Gebhardt, R. (2015). Application of confocal Raman
microscopy to investigate casein micro‐particles in blend casein/pectin films. International
Journal of Biological Macromolecules 74: 44–48.
29
Characterization of Lignocellulosic Biomass

and Processing for Second-Generation Sugars Production
Guadalupe Bustos Vázquez, Adrián Gonzalez Leos, Luis V. Rodríguez-Duran,
and Rodolfo Torres de Los Santos
Department of Biotechnology, University Autonomous of Tamaulipas, Unidad Académica Multidisciplinaria Mante, Cd. Mante,
Tamaulipas, México
3.1 Introduction
In recent decades, the greenhouse effect has been concerning the scientific community.
Tropical deforestation and burning of fossil fuels and vegetation, including pruning wastes
which are usually burned in the field, releasing carbon dioxide to the atmosphere, are
increasing the production of greenhouse gases (Bustos et al. 2007; Paramanandham and
Ronald Ross 2015). This is important when we consider the large amount of pruning wastes
generated worldwide.
During some agro‐industrial processes, by‐products are generated which, when not recy-
cled or processed, may cause various environmental problems. Their elimination usually
poses a management problem for the producing companies. However, these materials are
especially attractive sources of various compounds (such as sugars, pigments, food fiber,
protein, polyphenols, lignin, etc.) and can be potentially useful when they are transformed
by appropriate reactions in products of higher added value (Moldes et al. 2002;
Paramanandham and Ronald Ross 2015). The use of these compounds would thus revalor-
ize the waste fraction, and would provide useful compounds for the food, medical, and
chemical fields.
The present chapter focuses on the use of lignocellulosic biomass to produce biofuels and
the great potential this holds for the food industry.
3.2 Lignocellulosic Biomass
Biomass is defined as the renewable organic matter of plant, animal or natural origin or
artificial transformation thereof (Tandon 2015). According to its origin, biomass is classi-
fied as livestock, agricultural, forestry, urban, and industrial. From energy, industrial, and

agricultural points of view, biomass can be transformed into a wide variety of important
liquid, solid or gaseous products.
Lignocellulosic biomass is the most abundant organic material on Earth constituted fun-
damentally of cellulose, hemicelluloses, and lignin (of macromolecular character, the
structural materials of the cell wall) (Figure 3.1), (Reddy and Yang 2005; Amador et al.
2012; Saini et al. 2015; Madadi and Abbas 2017). The rest of biomass, called the nonstruc-
tural materials, includes extracts, proteins, starch, and ashes. Fan et al. (1982) describs the
common chemical fractions as follows.
3.2.1 Cellulose
Cellulose, (C6H10O5)n, is a linear homopolymer consisting of units of β‐glucose linked by
bonds 1–4, which has a degree of polymerization that varies between 1000 and 10 000 units
(Chesson and Forsberg 1997; Saini et al. 2015). The breakage of this molecule requires the
presence of catalysts (acids or enzymes). The cellulose performs a supporting function in
OH OH OH
O O O
OH OH OH
p-Coumaryl alcohol Coniferyl alcohol Sinapyl alcohol
H G S
S
G
H H
S G
S
Macrofibril S
G G
Plant
H H H
S G
Plant cell
G H
Macrofibril G
Cell Lignin
wall
Lignin
Hemicellulose 10–20 nm
Pentose
Hexose
n-3 n-3
n-3
Crystalline
cellulose n-3
Glucose n-3
Hydrogen
Cellodextrin bond
Figure 3.1 Structure of lignocellulose. Source: Adapted from Madadi and Abbas (2017); an open
access article, distributed under the terms of the Creative Commons Attribution License.
Characterization of Lignocellulosic Biomass and Processing for Second-Generation Sugars Production 31
the plant, acting as a skeleton. The physicochemical properties of cellulose depend on its
origin and degree of polymerization. Linear cellulose chains are associated in subunits of
approximately 3–4 nm in length that establish intermolecular hydrogen bonds forming
microfibrils. These join, forming bundles, to give rise to alternate sequences of amorphous
zones and crystalline zones. The porosity is variable depending on the arrangement of the
fibers in the different tissues.
When hydrogen bonds are broken along the bundle of chains, amorphous regions that
allow their hydration and better accessibility to the enzymatic attack are formed. The
microfibrils have variable amounts of amorphous and crystalline components which
depend on the degree of polymerization, the extension of the hydrogen bonds and finally
the source of cellulose from which they derive (Scheller and Ulvskov 2010; Alvira et al.
2010; Amador et al. 2012; Arevalo‐Gallegos et al. 2017).
3.2.2 Hemicellulose
Hemicelluloses are polymers of different monosaccharides: hexoses (glucose, mannose,
galactose) or pentoses (xylose, arabinose, rhamnose) (Amador et al. 2012). The function of
hemicellulose in plants is to facilitate the chemical bonds of cellulose with lignin, acting as
a binding agent between the fibers (Saini et al. 2015; Quiroz et al. 2016). In turn, within
hemicelluloses, two fractions are distinguished: γ‐cellulose (constituted by sugars other
than glucose) and β‐cellulose (constituted by glucose units). β‐cellulose shows differences
with respect to cellulose in its configuration (branched chains instead of linear), in its
lower rate of polymerization (between 200 and 300 units), and in its lack of crystallization.
Hemicelluloses can be hydrolyzed to sugars more easily than cellulose, although the reac-
tion must also be catalyzed by acids or enzymes (Nigam 2001).
Among the hemicelluloses we can find two other chemical fractions.
●● Polyuronides or pectic substances, which are polymers that possess uronic acids (arising
from the oxidation of sugars) as constituent units.
●● Acetyl groups, which form a fraction associated with hemicelluloses from the esterifica-
tion of functional groups. Acetyl groups are hydrolyzed by treatment with strong acids to
acetic and formic acids which, by saponification with alkali, give rise to the correspond-
ing salts of these acids (Wilkie 1983; Scheller and Ulvskov 2010).
3.2.3 Lignin
The chemical structure of lignin corresponds to the function it performs in the plant.
Lignin has a polymeric structure, three‐dimensional and amorphous, consisting of oxygen-
ated phenylpropane units, linked together by ether or carbon–carbon bonds, with variable
polymerization rates. It has hydroxyl and methoxyl groups in its structure, in quantities
dependent on the material being considered (Hamelinck et al. 2005). The biological func-
tions of lignin include agglomerating cellulose fibers, giving them greater rigidity, protect-
ing them from external attacks (both environmental and microbiological), and giving them
hydrophobic character. Due to these functions, it presents great resistance to chemical and
biological attacks, being one of the most resistant natural polymers. All the methods used
to isolate it cause changes in its structure.
The fact that the chemical nature of lignin differs greatly from cellulose and hemicelluloses
determines its different reactivity (Watkins et al. 2015). Thus, the typical reactions of hydroly-
sis of polysaccharides hardly affect lignin, being little reactive against acids and enzymes,
while it is more sensitive than polysaccharides to oxidation reactions or the action of certain
organic solvents. The interest of the effective attack of lignin lies in the fact that it allows
obtaining of products derived from it, at the same time that the polysaccharide–lignin com-
plex is broken down, an essential step to obtain good yields in the use of cellulose.
3.3 Analysis and Characterization of
One of the first steps for the development of processes for second‐generation sugars pro-
duction is the compositional analysis of the lignocellulosic biomass (Karimi and Taherzadeh
2016). Several methods have been developed for the analysis of lignocellulose. Most are
based on the fractionation of structural carbohydrates through a series of physical, chemi-
cal or enzymatic treatments. Then, the fractions obtained are analyzed by gravimetric,
volumetric or chromatographic methods. The composition of the fractions obtained by
each of these methods can differ significantly, which makes it difficult to compare the
results obtained by different methodologies.
A classic method for the analysis of lignocellulosic biomass is the determination of crude
fiber. This is part of the proximal chemical analysis and is accepted by the Association of
Official Agricultural Chemists (AOAC 2005a). For the determination of crude fiber, the
sample is defatted and then subjected successively to acid (1.25% H2SO4) and alkaline
hydrolysis (1.25% NaOH). The residue is dried, weighed, and the ash content determined.
Crude fiber is composed mainly of cellulose and lignin since hemicellulose, pectins, and
hydrocolloids are digested during acid and alkaline treatments.
Ven Soest (1963, 1967) developed the Acid Detergent Fiber (ADF) and Neutral Detergent
Fiber (NDF) methods for the fractionation and compositional analysis of plant‐based mate-
rials. NDF includes cellulose, hemicellulose and lignin while ADF includes only cellulose
and lignin. Hemicellulose contents may be calculated by the subtraction of ADF from NDF
values. Several modifications were made to improve the NDF method resulting in variabil-
ity among laboratories. Mertens (2002) conducted a collaborative study to standardize the
amylase‐treated NDF (aNDF) method. This modified technique was further adopted as an
official method by the Association of Official Agricultural Chemists (AOAC 2005b).
Southgate (1969) developed a method for the analysis of nonavailable carbohydrates that
provides values for the main components of the plant cell wall (water‐soluble polysaccha-
rides, hemicellulose, cellulose, and lignin). In this method, the sample is washed succes-
sively with methanol, water, and ether. The insoluble residue is then subjected to enzymatic
digestion with a commercial preparation of amylase followed by dilute acid (5% H2SO4)
and concentrated acid (72% H2SO4) hydrolysis. Simple sugars are measured by colorimetric
techniques. Englyst et al. (1982) modified the Southgate method and used gas chromatog-
raphy for the analysis of released sugars to improve the specificity of this technique.
However, the Englyst method does not measure lignin and uses indirect measurement‐by‐
difference techniques to estimate certain fractions (Dhingra et al. 2012).
Currently, one of the most widely used methods for the analysis of the structural carbo-
hydrates of lignocellulosic biomass is known as the quantitative saccharification procedure
which was standardized by scientists at the National Renewable Energy Laboratory (NREL)
(Li and Xu 2013). This procedure involves a strong acid solution (72% H2SO4) in primary
hydrolysis, followed by dilution with water and a secondary high‐temperature (120 °C)
hydrolysis. This hydrolyzes the polymeric carbohydrates into soluble monosaccharides.
Monomers in the hydrolysate solution are measured by high‐performance liquid chroma-
tography (HPLC), and lignin is determined gravimetrically from lignin‐rich residue (Sluiter
et al. 2010).
3.4 Lignocellulosic Biomass Utilization
In recent years, interest in the use of lignocellulosic biomass has grown due to the large
amount of agro‐industrial waste that is a cheap source of renewable raw material which
could be used in fermentation processes (Soni et al. 2018).
3.4.1 Current Agro-industrial Products and Lignocellulose

Materials in Mexico
Lignocellulosic biomass refers to crops, crop residues or forestry biomass. It has been esti-
mated that lignocellulose biomass in the world accounts for 60 billion tons per year.
Commonly, lignocellulosic biomass is present in four main sources: agricultural residues,
forest residues, energy crops, and cellulosic wastes (Jouzani and Taherzadeh 2015).
Lignocellulosic biomass is composed of the carbohydrates cellulose and hemicellu-
lose; since it contains around 70% of sugar, it makes a good substrate for hydrogen pro-
duction and other biofuels. Actually, lignocellulosic biomass is considered as the future
feedstock for ethanol production because potential characteristics including natural
abundance, renewability, recyclability, and year‐round global accessibility make resid-
ual biomass an eco‐attractive and petro‐alternative candidate (Cardona et al. 2010;
Arevalo‐Gallegos et al. 2017). Table 3.1 shows the chemistry composition and generated
volumes derived from the main agricultural products in Mexico such as cane bagasse,
rice husk, stalk and agave bagasse, all of which have a high potential for second‐
generation sugars production.
Mexico has an average annual volume of 52.6 million tons of sugar cane, the mainstay of
the national sugar industry (Secretaría de Agricultura y Desarrollo Rural [SAGARPA]
2016). The most important residue derived from the processing of sugar cane is bagasse,
which represents 30% of ground cane (Quiroz et al. 2016) and is one of the major lignocel-
lulosic residues in tropical countries (Cardona et al. 2010).
Agave plants are endemic to the arid regions of northern Mexico and the southern United
States. They are used for producing tequila and mezcal. Agave bagasse has been widely
studied in Mexico in recent years as biomass for animal feed, fiberboard production,
bioethanol, organic fertilizer, and activated carbon because of its high availability and
potential (Carrillo‐Nieves et al. 2019).
Table 3.1 Agricultural products and their agro-industrial wastes having high potential
for the production of second-generation sugars (Soltani et al. (2015) and SAGARPA (2016)).
Production in Main type of

Mexico agro-industrial Estimated waste
Main product (thousand tons) wastes volume Current uses of waste
Sugarcane 52 636 Cane bagasse 28–32% integral Food for pig and other animals
bagasse Bioethanol
Agave 1846 Agave bagasse 22% of agave Animal feed Fiberboard
production Bioethanol
Organic fertilizer Activated
carbon
Rice 10 165 Rice husk 28% of husk Rubber, insulator, absorben
and pigment
Corn 25.7 Stalk 27.5% from the Bioethanol, silage production
total plant and compost
Coffee beans 28 007 Coffee bean waste 90% of fruit Poultry food
Coffee as a means of lead and
cadmium absorption
Orange 4215 Orange peel 47.68% from the Essential oil
total fruit Pectin
Thanks to the variety of soils, climates and ecosystems, Mexico has ideal conditions for
growing a wide variety of products and rendering lignocellulosic biomass with a high
potential for second‐generation sugars production. The agricultural residues produced are
directly linked to the farming practices of each region and the technologies employed for
cultivating, transportation, storage, and processing of those residues (Carrillo‐Nieves et al.
2019). However, for the use of lignocellulosic materials, a pretreatment must be performed,
which is an essential step in the pyrolysis of lignocellulosic biomass. The effects of different
pretreatment processes on lignocellulose composition and sugar yield have been exten-
sively investigated (Chen et al. 2017). Though several pretreatment regimes are available,
biological pretreatment seems to be promising, with the added benefit of being an eco‐
friendly process with no inhibitor generation (Sindhu et al. 2016).
3.5 Obtaining Fermentative Media
3.5.1 Media Rich in Glucose

The degradation of cellulose chains requires catalysis by acids or enzymes. Both possibili-
ties can be achieved through the following processes.
3.5.1.1 Acid Hydrolysis

Cellulose is a polymer of glucose that can be depolymerized through hydrolysis in mono-
mers; these can be used in biorefining of sugars to produce fuels with high energy density
and other chemical products (Huang et al. 2008; Alonso et al. 2010; Geboers et al. 2011;
Brandt et al. 2013; Wijayapala 2014).
The mechanism of the reaction of hydrolysis of polysaccharides to monosaccharides, in

media catalyzed by acids, is produced by breakage of the bond in which the exocyclic
oxygen participates, with the formation of a cyclic carbon ion. For the study of acid
hydrolysis processes of polysaccharides are usually distinguished between those using
concentrated or dilute acids (Harris et al. 1984; Wright et al. 1985; Helm 1987; Kumar
et al. 2009; Pilath et al. 2010; Tian et al. 2010; El‐Zawawy et al. 2011; Amador et al. 2012;
Chandler et al. 2012).
The disadvantage of the hydrolysis of cellulose with dilute acids is that, due to the exist-
ence of sugars in the hydrolysis medium, degradation products toxic for microorganisms
are formed, which may prevent subsequent transformations via fermentation (Wright
et al. 1985; Chandel et al. 2007, 2010, 2011; Alvira et al. 2010; Gírio et al. 2010; Amador
et al. 2012).
The hydrolysis of cellulose with concentrated acids can proceed rapidly at moderate tem-
perature, avoiding degradation reactions and allowing almost quantitative yields in glucose
in short reaction times (Amador et al. 2012; Barrier et al. 1986; Galbe and Zacchi 2002,
2007; Wright et al. 1985). The drawback of this mode of operation is that the final products
of the hydrolysis are oligomers, produced by reversion of monosaccharides. Therefore, a
posthydrolysis stage (with dilute acids, at a moderate temperature) is needed to obtain the
corresponding sugars.
3.5.1.2 Enzymatic Hydrolysis

Compared to acid hydrolysis of cellulose, biotechnologic processes have the advantage of
using moderate temperatures and pressure conditions in noncorrosive media, which means
significant savings in energy and equipment. In addition, enzymatic hydrolysis is selective,
affecting only the polysaccharides without attacking the phenolic fraction, which results in
purer and more easily fermentable solutions. When opting for the biotechnologic pathway
(enzymatic hydrolysis), one must take into account the fact that, in a native lignocellulosic
material, lignin and cellulose form a system of interpenetrable polymers that are both
physically and chemically bound, so accessibility of the enzymes to cellulose is limited.
Therefore, in the development of processes for the conversion of lignocellulosic materials
by enzymatic hydrolysis, it is essential to alter their structure and composition by means of
physical, chemical or biological pretreatment (Barbado et al. 1991; Parajo et al. 1997;
Romano et al. 2005; Sveinsdottir et al. 2009).
Most of the time, the elimination of lignin is not a sufficient condition for the enzymatic
hydrolysis of the native wood to pass with acceptable reaction kinetics. It has been demon-
strated (Parajó et al. 1996) that after delignifying the eucalyptus wood following the
Acetosolv process, wood containing up to 90% cellulose is obtained and its enzymatic
hydrolysis achieves conversions of only 5% of the initial polysaccharides. To improve the
effectiveness of enzymatic hydrolysis, it is necessary to submit the pulp to a second treat-
ment called “conditioning,” which achieves a series of structural modifications such as the
reduction of the degree of polymerization, the reduction of crystallinity, and/or an increase
of the specific surface. This type of treatment can be applied to all types of lignocellulosic
materials; the different pretreatments proposed for the improvement of enzymatic sac-
charification can be classified into physical, chemical and physical–chemical treatments.
After the stages of pretreatment and conditioning of the cellulose fraction, there are
many species of bacteria, fungi and actinomycetes that produce enzymatic complexes
called cellulases (Szakäcs‐Dobozi et al. 1985; Li et al. 2011), which are very specific and
able to hydrolyze the β‐1,4‐glucosidic bonds of the cellulose chains in a reaction that shows
inhibition by the reaction products (Bisaria 1991; Béguin and Aubert 1994, 2000; Duff and
Murray 1996; Sreenath et al. 2001; Schimper et al. 2004; Medina et al. 2010).
3.5.2 Media Rich in Xylose

One of the applications of the hemicellulosic fraction is the use of its constituent sugars in
bioconversion processes (Amador et al. 2012). To obtain media rich in xylose, enzymes
(xylanases) can be used, which are frequently employed in the paper industry to reduce the
concentration in xylans of the delignified pulp (stage of chlorine‐free bleaching technolo-
gies and their derivatives). This is usually called TCF (Buchert et al. 1994; Woolridge 2014;
Singh et al. 2016). Bajpai (1999) has mentioned that factors such as temperature, pH, dos-
age of enzymes, pulp consistency, and reaction time affect the efficacy of xylanase treat-
ment. However, Sharma et al. (2014) concluded that the optimal pH for xylan treatment
varies between enzymes.
In order to be able to carry out the enzymatic hydrolysis of hemicellulosic liquors, it is
first necessary to submit the lignocellulosic material to a pretreatment of autohydrolysis or
chemical hydrolysis that allows us to solubilize the hemicellulosic fraction and make it
available for enzymatic attack. Acid prehydrolysis has the advantage that potentially valu-
able xylose‐rich media can be obtained directly for use in bioconversion processes in a
single stage. By the other method, autohydrolysis, since the oligomers produced during the
treatment cannot be assimilated by microorganisms, a subsequent stage (catalyzed by acids
or enzymes) is needed to obtain their corresponding monomeric sugars (Cuevas et al. 2010;
García et al. 2010a, 2010b). Depending on the starting materials, in addition to xylose (the
majority sugar), other sugars such as glucose, arabinose, and mannose appear, as well as
different reaction by‐products, which can act as inhibitors of microbial metabolism in the
fermentation process (Table 3.2). Inhibitors can decrease the rate of consumption of the
carbon source or even prevent fermentation (Parekh et al. 1987; Jönsson et al. 2013).
Occasionally, the inhibition may also be the result of synergistic effects (Saini et al. 2015).
3.6 Fermentation of Second-Generation Sugars
The constant increase in energy demand and the global concern about greenhouse gas
emissions and climate change have motivated the search for renewable and environmen-
tally friendly energy sources. Several biofuels can be produced from plant resources, such
as ethanol, butanol, and biodiesel. Among these, bioethanol has attracted attention due to
its potential as a fuel in motor vehicles (Robak and Balcerek 2018). Bioethanol can be used
as a pure fuel or mixed with gasoline in different proportions. Mixtures of 90% gasoline and
10% ethanol can be used in most internal combustion engines, while mixtures with higher
ethanol content can only be used in modified engines (Jeuland et al. 2004).
Bioethanol can be produced from different feedstocks. First‐generation ethanol is
obtained mainly from plant sugars or starches and utilizes food crops such as corn, wheat,
and sugarcane directly. In Brazil, most of the fuel ethanol is produced by fermentation of
Table 3.2 Raw materials, hydrolysis technologies, and composition of hemicellulosic hydrolyzates used as fermentative media.
Sugars (g/L) Inhibitors (g/L)
Raw material Hydrolysis Xylose Glucose Arabinose Acetic acid Furfural HMF References
Sugarcane H2SO4 21.6 3.0 — 3.65 0.5 — Aguilar et al. (2002)

bagasse HCl 9.2 2.6 1.1 1.1 0.21 — Tizazu and Moholkar (2018)
HNO3 22.6 3.77 3.31 3.59 1.54 — Bustos et al. (2003)
H3PO4 22.59 1.50 1.29 0.15 1.19 Laopaiboon et al. (2010)
18.6 2.87 2.04 0.9 1.32 — Rodrıguez‐Chong et al. (2004)
17.7 1.6 2.1 3.61 0.65 — Gámez et al. (2004)
Agave bagasse H2SO4 24.6 1.0 0.4 — 0.5 — Saucedo‐Luna et al. (2010)
Pinnapple husk H2SO4 26.90 2.61 — 7.71 0.29 — Amador et al. (2012)
Sorgum straw H2SO4 18.27 6.78 — 1.35 0.7 — Tellez‐Luis et al. (2002)
HCl 16.2 3.8 — 1.9 2.0 — Herrera et al. (2003)
Rice straw H2SO4 18.33 3.29 3.40 1.05 0.10 0.17 Mussatto (2002)
Wheat Straw H2SO4 12.8 1.7 2.6 2.7 0.15 — Nigam (2001)
Corn stover H2SO4 9.09 2.13 1.01 1.48 0.56 0.08 Cao et al. (2009)
Oil palm empty fruit bunch fiber H2SO4 29.4 2.34 — 1.25 0.87 — Rahman et al. (2006)
Vine trimming wastes H2SO4 17.4 11.1 4.3 4.0 0.7 0.1 Bustos et al. (2005)
Giant reed H2SO4 21.8 2.7 — 3.1 0.9 0.7 Shatalov and Pereira (2012)
Water hyacinth H2SO4 12.4 1.7 2.2 5.6 1.82 0.47 Nigam (2002)
Sunflower cake H2SO4 25.81 21.96 6.52 — — — Camargo et al. (2014)
Sunflower meal H2SO4 24.88 22.33 8.6 — — — Camargo et al. (2014)
HMF, 5‐hydroxymethylfurfural.
c03.indd 37 11/12/2019 6:03:03 PM

sugarcane, while in North America and Europe, most is produced from corn but also from
other grains (Niphadkar et al. 2018).
The use of first‐generation bioethanol can contribute to reducing the emission of green-
house gases. For example, in 2016 the use of corn‐derived ethanol in the United States
reduced greenhouse emissions from transportation by 43.5 million metric tonnes CO2
equivalent (Robak and Balcerek 2018). However, the sustainability and economy of first‐
generation bioethanol have been severely questioned. The main concerns about the pro-
duction of the first generation of biofuels are related to competition for land and water used
for food and fiber production (Fargione et al. 2008), as well as the high production and
processing costs that often require government subsidies in order for them to compete with
petroleum products (Sims et al. 2010).
Concerns about the sustainability of first‐generation biofuels have motivated the devel-
opment of the second generation. Second‐generation bioethanol uses renewable and inex-
pensive nonfood lignocellulosic biomass as feedstock. The production of ethanol from
forestry and agricultural residues does not directly compete with food/feed and has negli-
gible effects on their prices (Zhou et al. 2018).
The production of second‐generation bioethanol involves a series of steps to convert the
lignocellulosic biomass to ethanol. In the classic approach, lignocellulose is subjected to a
physical, chemical or biochemical pretreatment that makes the fibers more accessible to
hydrolysis; then, the pretreated biomass is hydrolyzed chemically or enzymatically to
release fermentable sugars; finally, the sugars are fermented by yeasts to ethanol (Niphadkar
et al. 2018). Different integrative strategies have been proposed for the production of
second‐generation ethanol, such as Simultaneous Saccharification and Fermentation (SSF)
and Consolidated Bioprocessing (CBP). SSF involves the enzymatic hydrolysis of cellulose
and alcoholic fermentation in a single step. This approach may reduce processing time and
energy consumption (de Araujo Guilherme et al. 2019). CBP refers to the combining of four
biological events required for the conversion of lignocellulose to ethanol (production of
saccharolytic enzymes, hydrolysis of polysaccharides present in pretreated biomass, fer-
mentation of hexose sugars, and fermentation of pentose sugars) in one reactor (Van Zyl
et al. 2007).
Ethanol production from sugar derived from starch and sucrose has been commercially
dominated by the yeast Saccharomyces cerevisiae. However, sugar derived from lignocellu-
losic biomass is a mixture of hexoses (primarily glucose) and pentoses (primarily xylose)
and most wild‐type strains of S. cerevisiae do not metabolize xylose (Gray et al. 2006).
3.6.1 Pentose Fermenting by Recombinant Microorganisms

Sustainable development and the recycling of resources have become increasingly vital in
recent years. The transformation of waste from different productive sectors has become an
important strategy for recycling and obtaining new by‐products of industrial interest (Yang
et al. 2018). The environment is a rich source of fermentative microorganisms for research.
However, wild‐type strains do not produce the high yields required for the industrial pro-
duction of second‐generation bioethanol. These include a high tolerance to high tempera-
tures, the ability to grow in lignocellulosic hydrolyzate, and the effective utilization of
xylose (Ali et al. 2017), since they cannot use both pentoses and hexoses. Therefore, they
are modified by genetic engineering linking genes from several organisms to create modi-
fied cells of Escherichia coli, S. cerevisiae, or Zymomonas mobilis (Martins et al. 2018).
Metabolic engineering allows the design of biochemical pathways that do not exist in the
natural world, as well as the redesign of biochemical pathways (Robak and Balcerek 2018).
Metabolic engineering applied to fermentative microorganisms improves their resistance
to fermentative conditions and enhances their resistance to inhibitors generated during
pretreatment, as well as their tolerance to ethanol and high concentrations of sugar, mak-
ing ethanol production more profitable (Kim et al. 2010).
Xylose is one of the main derivatives of lignocellulosic biomass hydrolyzate, essential
for the bioconversion of lignocellulose to fuels and other chemical products. Certain very
specific microorganisms can metabolize xylose into xylulose using enzymes such as oxi-
doreductase xylose reductase and xylitol dehydrogenase or isomerase such as xylose
isomerase, and finally through the Pentose Phosphate Pathway (PPP) (Moysés et al. 2016;
Robak and Balcerek 2018). The fermentation of xylose remains a challenge due to the
complexity of cellulosic hydrolyzate (Zhang and Geng 2012); in this sense, applying tech-
niques of metabolic engineering has added a pathway for the conversion of pentose or
other sugars into a yeast strain that produces natural ethanol, such as S. cerevisiae or the
bacterium Z. mobilis (Robak and Balcerek 2018). Z. mobilis genetically modified xylose
fermentation has been developed through the introduction of enzymes (such as transaldo-
lase and transketolase) in the pentose phosphate pathway and operons responsible for the
adaptation of xylose.
The second mode of recombination involves the genetic modification of microorganisms
that metabolize multiple sugars, to allow them to produce ethanol in the path of glycolysis
(Gamage et al. 2010). Attempts to use genetic engineering to allow the simultaneous use of
mixed sugars for the production of ethanol have focused mainly on the yeast S. cerevisiae
and the bacteria Clostridium cellulolyticum, Lactobacillus casei, Z. mobilis, E. coli, and
Klebsiella oxytoca (Robak and Balcerek 2018). Methods to obtain microorganisms capable
of simultaneously consuming glucose and xylose include mutagenesis and the introduc-
tion of a heterologous metabolic pathway for the utilization of xylose in conventional well‐
known strains such as S. cerevisiae.
Zhang and Geng (2012) modified the genome of modified S. cerevisiae strain ScF2
through the shuffling method and achieved an ScF2 strain that used both glucose and
xylose at a higher rate and produced more ethanol. Although the rate of glucose consump-
tion for ScF2 was slower compared to the control, de Figueiredo Vilela et al. (2015) demon-
strated that the heterologous expression of a bacterial xylose isomerase gene (xylA) from
Burkholderia cenocepacia allowed a strain of S. cerevisiae to ferment the xylose anaerobi-
cally, without accumulation of xylitol. However, the recombinant yeast fermented the
xylose slowly. In this study, an evolutionary engineering strategy was applied to improve
the fermentation of xylose by means of the yeast strain that expresses xylA, which involved
the sequential cultivation of batches in xylose. The resulting yeast strain co‐fermented glu-
cose and xylose rapidly and almost simultaneously, exhibiting improved ethanol yield and
productivity. It was also observed that when the cells were cultured in a medium contain-
ing higher glucose concentrations before being transferred to the fermentation medium,
higher rates of xylose consumption and ethanol production were obtained, demonstrating
that the use of xylose is not regulated by catabolic repression.
3.7 Conclusion
Agro‐industrial wastes are especially attractive sources of various compounds (sugars, pig-
ments, food fiber, proteins, polyphenols, lignin, etc.) and can be potentially useful when
they are transformed by appropriate reactions into high added‐value products. Obtaining
these compounds would thus revalorize a proportion of the residues and originate useful
compounds for the food, medical, and chemical fields. Nowadays, lignocellulosic biomass
is considered as the future raw material for biofuels production because their potential
characteristics such as natural abundance, renewal and recycling capacity and ease of
access throughout the year, all over the world, make residual biomass an eco‐attractive
alternative energy source.
R
eferences
Aguilar, R., Ramırez, J.A., Garrote, G., and Vázquez, M. (2002). Kinetic study of the acid
hydrolysis of sugar cane bagasse. Journal of Food Engineering 55 (4): 309–318.
Ali, S.S., Wu, J., Xie, R. et al. (2017). Screening and characterizing of xylanolytic and xylose‐
fermenting yeasts isolated from the wood‐feeding termite, Reticulitermes chinensis. PLoS
One 12 (7): e0181141.
Alonso, D.M., Bond, J.Q., and Dumesic, J.A. (2010). Catalytic conversion of biomass to
biofuels. Green Chemistry 12 (9): 1493–1513.
Alvira, P., Tomás‐Pejó, E., Ballesteros, M.J., and Negro, M.J. (2010). Pretreatment technologies
for an efficient bioethanol production process based on enzymatic hydrolysis: a review.
Bioresource Technology 101 (13): 4851–4861.
Amador, K.R., Carrillo, Ó.R., Aguilar, P.A., and Baudrit, J.V. (2012). Obtención de xilosa a
partir de desechos lignocelulósicos de la producción y proceso industrial de la piña
(Ananascomusus). Uniciencia 26 (1): 75–89.
AOAC (Association of Official Agricultural Chemists) (2005a). AOAC Official Method 962.09:
Fiber (crude) in animal feed and pet food ceramic fiber filter method. Official Methods of
Analysis of AOAC International, Gaithersburg.
AOAC (Association of Official Agricultural Chemists) (2005b). AOAC Official Method 2002.04:
Amylase‐treated neutral detergent fiber in feeds using refluxing in beakers or crucibles.
Official Methods of Analysis of AOAC International, Gaithersburg.
de Araujo Guilherme, A., Dantas, P.V.F., de Araújo Padilha, C.E. et al. (2019). Ethanol
production from sugarcane bagasse: use of different fermentation strategies to enhance an
environmental‐friendly process. Journal of Environmental Management 234: 44–51.
Arevalo‐Gallegos, A., Ahmad, Z., Asgher, M. et al. (2017). Lignocellulose: a sustainable
material to produce value‐added products with a zero waste approach‐a review.
Bajpai, P. (1999). Application of enzymes in the pulp and paper industry. Biotechnology
Progress 15 (2): 147–157.
Barbado, D.V., Liñares, J.C.P., Vázquez, G., and Yusty, M.A.L. (1991). Transformación de
materiales lignocelulósicos. Composición, fraccionamiento y aprovechamiento. Revista de
agroquímica y tecnología de alimentos 31 (2): 143–164.
Barrier, J.W., Moore, M.R., Farina, G.E., et al. (1986). Experimental production of ethanol from
agricultural cellulosic materials using low‐temperature acid hydrolysis. Third Southern
Biomass Energy Research Conference, Gainesville, FA, USA.
Béguin, P. and Aubert, J.P. (1994). The biological degradation of cellulose. FEMS Microbiology
Reviews 13 (1): 25–58.
Béguin, P. and Aubert, J. (2000). Cellulases. In: Encyclopedia of Microbiology, 2e, vol. 1 (eds. M.
Alexander, B. Bloom, D. Hopwood, et al.), 744–758. New York: Academic Press.
Bisaria, V.S. (1991). Bioconversion of waste materials to industrial products. In: Bioprocessing of
Agro‐Residues to Glucose and Chemicals (ed. A.M. Martin), 187–223. UK: Elsevier Science.
Brandt, A., Gräsvik, J., Hallett, J.P., and Welton, T. (2013). Deconstruction of lignocellulosic
biomass with ionic liquids. Green Chemistry 15 (3): 550–583.
Buchert, J., Tenkanen, M., Kantelinen, A., and Viikari, L. (1994). Application of xylanases in
the pulp and paper industry. Bioresource Technology 50 (1): 65–72.
Bustos, G., Ramírez, J.A., Garrote, G., and Vázquez, M. (2003). Modeling of the hydrolysis of
sugar cane bagasse with hydrochloric acid. Applied Biochemistry and Biotechnology 104 (1):
51–68.
Bustos, G., Moldes, A.B., Cruz, J.M., and Domínguez, J.M. (2005). Production of lactic acid
from vine‐trimming wastes and viticulture lees using a simultaneous saccharification
fermentation method. Journal of the Science of Food and Agriculture 85 (3): 466–472.
Bustos, G., de la Torre, N., Moldes, A. et al. (2007). Revalorization of hemicellulosic trimming
vine shoots hydrolyzates trough continuous production of lactic acid and biosurfactants by
L. pentosus. Journal of Food Engineering 78 (2): 405–412.
Camargo, D., Gomes, S.D., Felipe, M.D.G., and Sene, L. (2014). Response of by‐products of
sunflower seed processing to dilute‐acid hydrolysis aiming fermentable sugar production.
Journal of Food, Agriculture and Environment 12 (2): 239–246.
Cao, G., Ren, N., Wang, A. et al. (2009). Acid hydrolysis of corn stover for biohydrogen
production using Thermoanaerobacterium thermosaccharolyticum W16. International
Journal of Hydrogen Energy 34 (17): 7182–7188.
Cardona, C.A., Quintero, J.A., and Paz, I.C. (2010). Production of bioethanol from sugarcane
bagasse: status and perspectives. Bioresource Technology 101 (13): 4754–4766.
Carrillo‐Nieves, D., Alanís, M.J.R., de la Cruz Quiroz, R. et al. (2019). Current status and future
trends of bioethanol production from agro‐industrial wastes in Mexico. Renewable and
Sustainable Energy Reviews 102: 63–74.
Chandel, A.K., Kapoor, R.K., Singh, A., and Kuhad, R.C. (2007). Detoxification of sugarcane
bagasse hydrolysate improves ethanol production by Candida shehatae NCIM 3501.
Chandel, A.K., Singh, O.V., and Rao, L.V. (2010). Biotechnological applications of
hemicellulosic derived sugars: state‐of‐the‐art. In: Sustainable Biotechnology (eds. O. Singh
and S. Harvey), 63–81. Dordrecht: Springer.
Chandel, A.K., da Silva, S.S., and Singh, O.V. (2011). Detoxification of lignocellulosic
hydrolysates for improved bioethanol production. In: Biofuel Production – Recent
Developments and Prospects (ed. M.A.S. Bernardes), 225–246. UK: InTech.
Chandler, C., Villalobos, N., González, E. et al. (2012). Hidrólisis ácida diluida en dos etapas de
bagazo de caña de azúcar para la producción de azúcares fermentables. Multiciencias 12 (3):
245–253.
Chen, H., Liu, J., Chang, X. et al. (2017). A review on the pretreatment of lignocellulose for
high‐value chemicals. Fuel Processing Technology 160: 196–206.
Chesson, A. and Forsberg, C.W. (1997). Polysaccharide degradation by rumen microorganisms.
In: The Rumen Microbial Ecosystem (eds. P.N. Hobson and C.S. Stewart), 329–381.
Dordrecht: Springer.
Cuevas, M., Sánchez, S., Bravo, V. et al. (2010). Determination of optimal pre‐treatment
conditions for ethanol production from olive‐pruning debris by simultaneous
saccharification and fermentation. Fuel 89 (10): 2891–2896.
Dhingra, D., Michael, M., Rajput, H., and Patil, R.T. (2012). Dietary fibre in foods: a review.
Journal of Food Science and Technology 49 (3): 255–266.
Duff, S.J. and Murray, W.D. (1996). Bioconversion of forest products industry waste cellulosics
to fuel ethanol: a review. Bioresource Technology 55 (1): 1–33.
El‐Zawawy, W.K., Ibrahim, M.M., Abdel‐Fattah, Y.R. et al. (2011). Acid and enzyme hydrolysis
to convert pretreated lignocellulosic materials into glucose for ethanol production.
Carbohydrate Polymers 84 (3): 865–871.
Englyst, H., Wiggins, H.S., and Cummings, J.H. (1982). Determination of the non‐starch
polysaccharides in plant foods by gas‐liquid chromatography of constituent sugars as alditol
acetates. Analyst 107 (1272): 307–318.
Fan, L.T., Lee, Y.H., and Gharpuray, M.M. (1982). The nature of lignocellulosics and their
pretreatments for enzymatic hydrolysis. In: Microbial Reactions: Advances in Biochemical
Engineering (Conference Proceedings), 157–187. Berlin: Springer.
Fargione, J., Hill, J., Tilman, D. et al. (2008). Land clearing and the biofuel carbon debt. Science
319 (5867): 1235–1238.
de Figueiredo Vilela, L., de Araujo, V.P.G., de Sousa Paredes, R. et al. (2015). Enhanced xylose
fermentation and ethanol production by engineered Saccharomyces cerevisiae strain. AMB
Express 5 (1): 16.
Galbe, M. and Zacchi, G. (2002). A review of the production of ethanol from softwood. Applied
Microbiology and Biotechnology 59 (6): 618–628.
Galbe, M. and Zacchi, G. (2007). Pretreatment of lignocellulosic materials for efficient
bioethanol production. Advances in Biochemical Engineering/Biotechnology 108: 41–65.
Gamage, J., Lam, H., and Zhang, Z. (2010). Bioethanol production from lignocellulosic
biomass, a review. Journal of Biobased Materials and Bioenergy 4 (1): 3–11.
Gámez, S., Ramírez, J.A., Garrote, G., and Vázquez, M. (2004). Manufacture of fermentable
sugar solutions from sugar cane bagasse hydrolyzed with phosphoric acid at atmospheric
pressure. Journal of Agricultural and Food Chemistry 52 (13): 4172–4177.
García, J., Cuevas, M., Bravo, V., and Sánchez, S. (2010a). Ethanol production from olive
prunings by autohy‐drolysis and fermentation with Candida tropicalis. Renewable Energy 35:
1602–1608.
García, J.F., Sánchez, S., Bravo, V., and Cuevas, M. (2010b). Autohydrolysis and acid post‐
hydrolysis of olive pruning debris. Afinidad 67 (548): 279–282.
Geboers, J.A., van de Vyver, S., Ooms, R. et al. (2011). Chemocatalytic conversion of cellulose:
opportunities, advances and pitfalls. Catalysis Science & Technology 1 (5): 714–726.
Gírio, F.M., Fonseca, C., Carvalheiro, F. et al. (2010). Hemicelluloses for fuel ethanol: a review.
Gray, K.A., Zhao, L., and Emptage, M. (2006). Bioethanol. Current Opinion in Chemical Biology
10 (2): 141–146.
Hamelinck, C.N., van Hooijdonk, G., and Faaij, A.P. (2005). Ethanol from lignocellulosic
biomass: techno‐economic performance in short‐, middle‐and long‐term. Biomass and
Bioenergy 28 (4): 384–410.
Harris, J., Baker, A., and Zerbe, J. (1984). Two‐stage, dilute sulfuric acid hydrolysis of
hardwood for ethanol production. Energy from Biomass and Wastes 8: 1151–1170.
Helm, R.F. (1987). The reversion and dehydration reactions of glucose during the dilute
sulphuric acid hydrolysis of cellulose. www.osti.gov/biblio/5380955.
Herrera, A., Téllez‐Luis, S.J., Ramırez, J.A., and Vázquez, M. (2003). Production of xylose from
sorghum straw using hydrochloric acid. Journal of Cereal Science 37 (3): 267–274.
Huang, H.J., Ramaswamy, S., Tschirner, U.W., and Ramarao, B.V. (2008). A review of
separation technologies in current and future biorefineries. Separation and Purification
Technology 62 (1): 1–21.
Jeuland, N., Montagne, X., and Gautrot, X. (2004). Potentiality of ethanol as a fuel for
dedicated engine. Oil & Gas Science and Technology 59 (6): 559–570.
Jönsson, L.J., Alriksson, B., and Nilvebrant, N.O. (2013). Bioconversion of lignocellulose:
inhibitors and detoxification. Biotechnology for Biofuels 6 (1): 16.
Jouzani, G.S. and Taherzadeh, M.J. (2015). Advances in consolidated bioprocessing systems for
bioethanol and butanol production from biomass: a comprehensive review. Biofuel Research
Journal 2 (1): 152–195.
Karimi, K. and Taherzadeh, M.J. (2016). A critical review of analytical methods in
pretreatment of lignocelluloses: composition, imaging, and crystallinity. Bioresource
Technology 200: 1008–1018.
Kim, J.H., Block, D.E., and Mills, D.A. (2010). Simultaneous consumption of pentose and
hexose sugars: an optimal microbial phenotype for efficient fermentation of lignocellulosic
biomass. Applied Microbiology and Biotechnology 88 (5): 1077–1085.
Kumar, P., Barrett, D.M., Delwiche, M.J., and Stroeve, P. (2009). Methods for pretreatment of
lignocellulosic biomass for efficient hydrolysis and biofuel production. Industrial &
Engineering Chemistry Research 48 (8): 3713–3729.
Laopaiboon, P., Thani, A., Leelavatcharamas, V., and Laopaiboon, L. (2010). Acid hydrolysis of
sugarcane bagasse for lactic acid production. Bioresource Technology 101 (3): 1036–1043.
Li, H.Q. and Xu, J. (2013). A new correction method for determination on carbohydrates in
lignocellulosic biomass. Bioresource Technology 138: 373–376.
Li, D.C., Li, A.N., and Papageorgiou, A.C. (2011). Cellulases from thermophilic fungi: recent
insights and biotechnological potential. Enzyme Research 2011: 308730.
Madadi, M. and Abbas, A. (2017). Lignin degradation by fungal pretreatment: a review. Journal
of Plant Pathology & Microbiology 8 (2) https://doi.org/10.4172/2157‐7471.1000398.
Martins, G.M., Bocchini‐Martins, D.A., Bezzerra‐Bussoli, C. et al. (2018). The isolation of
pentose‐assimilating yeasts and their xylose fermentation potential. Brazilian Journal of
Microbiology 49 (1): 162–168.
Medina, D.A.P., Nuñez, M.F.A., and Ordoñes, M.S. (2010). Obtención de enzimas celulasas por
fermentación sólida de hongos para ser utilizadas en el proceso de obtención de bioalcohol
de residuos del cultivo de banano. Revista Tecnológica‐ESPOL 23 (1): 81–88.
Mertens, D.R. (2002). Gravimetric determination of amylase‐treated neutral detergent fiber in

feeds with refluxing in beakers or crucibles: collaborative study. Journal of AOAC
International 85 (6): 1217–1240.
Moldes, A., Cruz, J., Domínguez, J., and Parajó, J. (2002). Production of a cellulosic substrate
susceptible to enzimatic hydrolysis from prehidrolized barley husks. Agricultural and Food
Science 11 (1): 51–58.
Moysés, D., Reis, V., Almeida, J. et al. (2016). Xylose fermentation by Saccharomyces cerevisiae:
challenges and prospects. International Journal of Molecular Sciences 17 (3): 207.
Mussatto, S.I. (2002). Influencia do tratamento do hidrolisado hemicelulosico de palha de arroz
na producao de xilitol por Candida guilliermondii. Masters thesis. Faculdade de Engenharia
Química de Lorena, São Paulo, Brazil.
Nigam, J.N. (2001). Ethanol production from wheat straw hemicellulose hydrolysate by Pichia
stipitis. Journal of Biotechnology 87 (1): 17–27.
Nigam, J.N. (2002). Bioconversion of water‐hyacinth (Eichhornia crassipes) hemicellulose acid
hydrolysate to motor fuel ethanol by xylose‐fermenting yeast. Journal of Biotechnology 97
(2): 107–116.
Niphadkar, S., Bagade, P., and Ahmed, S. (2018). Bioethanol production: insight into past,
present and future perspectives. Biofuels 9 (2): 229–238.
Parajó, J.C., Domínguez, H., and Domínguez, J.M. (1996). Charcoal adsorption of wood
hydrolysates for improving their fermentability: influence of the operational conditions.
Parajo, J.C., Alonso, J.L., and Moldes, A.B. (1997). Production of lactic acid from lignocellulose
in a single stage of hydrolysis and fermentation. Food Biotechnology 11 (1): 45–58.
Paramanandham, J. and Ronald Ross, P. (2015). Lignin and cellulose content in coir waste on
subject to sequential washing. Journal of Chemistry and Chemical Research 1 (1): 10–13.
Parekh, S., Parekh, R., and Wayman, M. (1987). Fermentation of wood‐derived acid
hydrolysates in a batch bioreactor and in a continuous dynamic immobilized cell bioreactor
by Pichia stipitis R. Process Biochemistry 22: 85–91.
Pilath, H.M., Nimlos, M.R., Mittal, A. et al. (2010). Glucose reversion reaction kinetics. Journal
of Agricultural and Food Chemistry 58 (10): 6131–6140.
Quiroz, A.P.B., Coca, A.L.B., and Baquero, P.P. (2016). Sostenibilidad del aprovechamiento del
bagazo de caña de azùcar en el Valle del Cauca, Colombia. Ingeniería Solidaria 12 (20):
133–149.
Rahman, S.H.A., Choudhury, J.P., and Ahmad, A.L. (2006). Production of xylose from oil palm
empty fruit bunch fiber using sulfuric acid. Biochemical Engineering Journal 30 (1): 97–103.
Reddy, N. and Yang, Y. (2005). Biofibers from agricultural byproducts for industrial
applications. Trends in Biotechnology 23 (1): 22–27.
Robak, K. and Balcerek, M. (2018). Review of second‐generation bioethanol production from
residual biomass. Food Technology and Biotechnology 56 (2): 174–187.
Rodrıguez‐Chong, A., Ramı́rez, J.A., Garrote, G., and Vázquez, M. (2004). Hydrolysis of sugar
cane bagasse using nitric acid: a kinetic assessment. Journal of Food Engineering 61 (2):
143–152.
Romano, S., González, E., and Laborde, M. (2005). Combustibles Alternativos. Buenos Aires,
Argentina: Red CYTED: Nuevas Tecnologías para la obtención de Biocombustibles.
Ediciones Cooperativas.
SAGARPA (Secretaria de Agricultura y Desarrollo Rural) (2016). Atlas Agroalimentario. Mexico

City: SAGARPA.
Saini, J.K., Saini, R., and Tewari, L. (2015). Lignocellulosic agriculture wastes as biomass
feedstocks for second‐generation bioethanol production: concepts and recent developments.
Biotechnology 5 (4): 337–353.
Saucedo‐Luna, J., Castro‐Montoya, A.J., Rico, J.L., and Campos‐García, J. (2010). Optimización
de hidrólisis ácida de bagaso de Agave tequilana Weber. Revista Mexicana de Ingeniería
Química 9 (1): 91–97.
Scheller, H.V. and Ulvskov, P. (2010). Hemicelluloses. Annual Review of Plant Biology 61: 263–289.
Schimper, C., Keckeis, R., Ibanescu, C. et al. (2004). Influence of steam and dry heat
pretreatment on fibre properties and cellulase degradation of cellulosic fibres. Biocatalysis
and Biotransformation 22 (5–6): 383–389.
Sharma, A., Thakur, V.V., Shrivastava, A. et al. (2014). Xylanase and laccase based enzymatic
kraft pulp bleaching reduces adsorbable organic halogen (AOX) in bleach effluents: a pilot
scale study. Bioresource Technology 169: 96–102.
Shatalov, A.A. and Pereira, H. (2012). Xylose production from giant reed (Arundo donax L.):
modeling and optimization of dilute acid hydrolysis. Carbohydrate Polymers 87 (1): 210–217.
Sims, R.E., Mabee, W., Saddler, J.N., and Taylor, M. (2010). An overview of second generation
biofuel technologies. Bioresource Technology 101 (6): 1570–1580.
Sindhu, R., Binod, P., and Pandey, A. (2016). Biological pretreatment of lignocellulosic
biomass – an overview. Bioresource Technology 199: 76–82.
Singh, G., Capalash, N., Kaur, K. et al. (2016). Enzymes: applications in pulp and paper
industry. In: Agro‐Industrial Wastes as Feedstock for Enzyme Production (eds. G.S. Dhillon
and S. Kaur), 157–172. St Louis, MO: Elsevier Academic Press.
Sluiter, J.B., Ruiz, R.O., Scarlata, C.J. et al. (2010). Compositional analysis of lignocellulosic
feedstocks. 1. Review and description of methods. Journal of Agricultural and Food
Chemistry 58 (16): 9043–9053.
Soltani, N., Bahrami, A., Pech‐Canul, M.I., and González, L.A. (2015). Review on the
physicochemical treatments of rice husk for production of advanced materials. Chemical
Engineering Journal 264: 899–935.
Soni, S.K., Sharma, A., and Soni, R. (2018). Cellulases: role in lignocellulosic biomass
utilization. Methods in Molecular Biology 1796: 3–23.
Southgate, D.A.T. (1969). Determination of carbohydrates in foods II.‐unavailable
carbohydrates. Journal of the Science of Food and Agriculture 20 (6): 331–335.
Sreenath, H.K., Moldes, A.B., Koegel, R.G., and Straub, R.J. (2001). Lactic acid production from
agriculture residues. Biotechnology Letters 23 (3): 179–184.
Sveinsdottir, M., Rafn, S., Baldursson, B., and Orlygsson, J. (2009). Ethanol production from
monosugars and lignocellulosic biomass by thermophilic bacteria isolated from Icelandic
hot springs. Icelandic Agricultural Sciences 22: 45–58.
Szakäcs‐Dobozi, M., Szakäcs, G., Meyer, D., and Klappach, G. (1985). Enhancement of
enzymatic degradation of cellulose by application of mixed enzyme systems of different
fungal origin. Acta Biotechnologica 5 (1): 27–33.
Tandon, G. (2015). Bioproducts from residual lignocellulosic biomass. In: Advances in
Biotechnology (eds. N.N. Nawani, M. Khetmalas, P.N. Razdan and A. Pandey), 52–75. New
Delhi: I.K. International Publishing House Pvt. Ltd.
Tellez‐Luis, S.J., Ramırez, J.A., and Vázquez, M. (2002). Mathematical modelling of

hemicellulosic sugar production from sorghum straw. Journal of Food Engineering 52 (3):
285–291.
Tian, J., Wang, J., Zhao, S. et al. (2010). Hydrolysis of cellulose by the heteropoly acid H 3 PW
12 O 40. Cellulose 17 (3): 587–594.
Tizazu, B.Z. and Moholkar, V.S. (2018). Kinetic and thermodynamic analysis of dilute acid
hydrolysis of sugarcane bagasse. Bioresource Technology 250: 197–203.
Van Zyl, W.H., Lynd, L.R., den Haan, R., and McBride, J.E. (2007). Consolidated bioprocessing
for bioethanol production using Saccharomyces cerevisiae. Advances in Biochemical
Engineering/Biotechnology 108: 205–235.
Ven Soest, P.J. (1963). Use of detergents in the analysis of fibrous feeds II. A rapid method for
the determination of fiber and lignin. Journal of the Association of Official Agricultural
Chemists 46 (5): 829–835.
Ven Soest, P.J. (1967). Use of detergents in the analysis of fibrous feeds. Determination of plant
cell wall constituents. Journal of the Association of Official Analytical Chemists 50: 50–55.
Watkins, D., Nuruddin, M., Hosur, M. et al. (2015). Extraction and characterization of lignin
from different biomass resources. Journal of Materials Research and Technology 4 (1): 26–32.
Wijayapala, H.R.T. (2014). Catalytic Conversion of Biomass to Bio‐Fuels. Mississippi State:
Mississippi State University.
Wilkie, K. (1983). Hemicelluloses. ChemTech 5: 306–319.
Woolridge, E. (2014). Mixed enzyme systems for delignification of lignocellulosic biomass.
Catalysts 4 (1): 1–35.
Wright, J.D., Power, A.J., and Bergeron, P.W. (1985). Concentrated halogen acid hydrolysis
processes for alcohol fuel production (No. SERI/TP‐232‐2690; CONF‐850513‐2). Solar
Energy Research Institute, Golden, CO, USA.
Yang, M., Yun, J., Zhang, H. et al. (2018). Genetically engineered strains: application and
advances for 1, 3‐propanediol production from glycerol. Food Technology and Biotechnology
56 (1): 3–15.
Zhang, W. and Geng, A. (2012). Improved ethanol production by a xylose‐fermenting
recombinant yeast strain constructed through a modified genome shuffling method.
Biotechnology for Biofuels 5 (1): 46.
Zhou, Z., Lei, F., Li, P., and Jiang, J. (2018). Lignocellulosic biomass to biofuels and
biochemicals: a comprehensive review with a focus on ethanol organosolv pretreatment
technology. Biotechnology and Bioengineering 115 (11): 2683–2702.
47
Production of Biohydrogen from Lignocellulosic

Feedstocks
Sheetal Radhakrishnan1, Shiv Prasad2, Sandeep Kumar 2, and
Dhanya Subramanian3
1
Regional Research Station, ICAR-Central Arid Zone Research Institute, Bikaner, Rajasthan, India
2
Centre for Environment Science and Climate Resilient Agriculture, Indian Agricultural Research Institute, New Delhi, India
3
Centre for Environmental Science and Technology, Central University of Punjab, Bathinda, Punjab, India
4.1 Introduction
Fossil‐based fuels have been the main energy source since the Industrial Revolution.
However, the current situation of climate change with increasing concentrations of green-
house gases (GHG), mainly CO2, demands an alternate cleaner fuel, particularly for vehi-
cles, which contribute a major share to GHG levels (Balat and Kırtay 2010; Prasad et al.
2014). Here we can take the example of a developing country like India, where the trans-
portation sector alone consumes more than 75% of its diesel and petrol and contributes 60%
of the GHG emissions from related activities (Sheetal et al. 2017). If we are able to provide
a clean, renewable, domestically produced fuel in place of fossil fuels, it would be a major
breakthrough toward the low carbon economy. Moreover, with limited reserves worldwide,
crude oil is a nonrenewable fuel resource that is rapidly becoming limited and more expen-
sive. These factors, combined with environmental and political concerns, have led to stead-
ily growing interest in the eco‐friendly biofuels.
Biofuels may be liquid or gaseous fuels obtained mainly from biomass which can easily
replace conventional fossil fuels in vehicles (Sarma et al. 2013; Prasad et al. 2017, 2019),
offering a chance to switch to renewable, sulfur and aromatics free, lower emission fuels
(Prasad et al. 2012; Das et al. 2017). Recent advances in their production technologies are
also helping to make biofuels cost‐competitive with fossil fuels and playing a major part in
the future of transport.
A fuel currently gaining momentum is hydrogen, one of the cleanest renewable fuels
which does not emit oxides of carbon or any other harmful compounds and also releases
much more energy (2.75 times higher) than fossil fuels (Balat and Kırtay 2010). It is a flex-
ible energy carrier that may be produced from any primary energy source that is locally
abundant, and transformed into other energy forms for use in transport, power generation,

and industry, supporting the low carbon economy (IEA 2018). With an energy density vary-
ing from 120 to 142 MJ per kilogram of hydrogen, one unit (kg) can replace the energy
obtained from 3.55 L of diesel (Sarma et al. 2013). It can also be utilized for electricity gen-
eration via fuel cells. However, being a gas under normal conditions creates a problem in
storage and transport, which can be reduced by storage in solid or liquid compounds
through chemical or physiochemical means (Shakya et al. 2005).
To achieve a low‐carbon economy and energy source, hydrogen production needs to be
shifted from its current feedstock comprising the fossil fuels (as discussed in later sections)
to the more eco‐friendly and green biomass (Demirbas 2009; Das et al. 2017). Only when
renewable sources like biomass become a feedstock will hydrogen energy become a sus-
tainable and carbon‐free energy carrier, which can meet future energy needs. Similar to
other biofuels, hydrogen can be sustainably produced from lignocellulosic biomass which,
in addition to being more climate friendly, also makes hydrogen production renewable, as
the lignocellulosic biomass is composed primarily of agricultural wastes and residues
(Prasad et al. 2012; Bajpai 2016).
This chapter discusses the potential of lignocellulosic biomass for hydrogen production,
different conversion routes, their advantages and limitations along with examples of stud-
ies conducted using these methods and also how production may be made more economi-
cal. However, these production technologies are still under development, and only small
amounts of data about plants and their operation are currently available (Singh and
Rathore 2017). Therefore, it may be difficult for energy policy decision makers to identify
the advantages and disadvantages of production technologies based on biomass
feedstock.
4.2 Global Hydrogen Production and Use: Current Status
With global production of more than 50 million tons annually, hydrogen has become an
essential feedstock for its users (US DoE 2013). Currently, almost all the hydrogen pro-
duced in the world comes from fossil fuels. Natural gas accounts for about 50% of the base
material for hydrogen production, followed by heavy oils, naphtha, and coal. However,
hydrogen production from these sources poses the risk of higher GHG (mainly CO2) emis-
sions during the production process, which partially offsets the benefits from using hydro-
gen as fuel. In addition to the production stage, the extraction processes, such as fracking
to extract oil and gas from subterranean sources, create pollution and also release huge
amounts of methane, which is a more potent GHG. The ideal feedstock would thus be
renewable biomass as well as solar and/or wind energy. Biomass accounts for only 1% of
hydrogen production presently, indicating the huge potential that this resource has for fur-
ther utilization (Balat and Kırtay 2010). Current annual worldwide hydrogen use ranges
from 400 to 500 billion Nm3 (Demirbas 2009), increasing at the rate of almost 23 mt annu-
ally. Worldwide hydrogen production is given in Figure 4.1.
Present hydrogen consumption comprises around 3% of total energy use, and an annual
growth rate of 5–10% is projected (Mohan et al. 2007). As of 2015, hydrogen demand was 8
EJ (IRENA 2018). The highest hydrogen demand comes from the industrial chemical sector
Production of Biohydrogen from Lignocellulosic Feedstocks 49
40
Worldwide Total Production [1]*
Worldwide Merchant Production

30
(Million metric tons/year)
20
10
0
2005 2006 2007 2008 2009 2010
Figure 4.1 Worldwide hydrogen production. Source: CryoGas International; February 2006; February
2007; February 2008; February 2009; April 2010, February 2011. Note: [1]* Excludes hydrogen
production from syngas, by-product gases, and on-site plants not owned and operated by the
end-user. “Merchant” hydrogen production is defined here to mean any hydrogen produced by one
company for consumption by another company.
Table 4.1 Properties of hydrogen versus other conventional fuels (NHEB 2007).
Fuel/property Hydrogen Natural gas Petrol Liquid petroleum gas
Lower heating value (MJ/kg) 120.7 49.54 41.87–44.19 46.05

Higher heating value (MJ/kg) 141.89 54.89 43.73–47.45 50.24
Density at standard conditions (kg/m3) 0.08 0.6 720–780 510
Phase at standard conditions Gas Gas Liquid Liquid
Autoignition temperature in air (°C) 566–582 540 257 454–510
Ignition limit in air (vol %) 4.1–74 5.3–15 1.4–7.6 2.2–9.5
Diffusion coefficient in air (cm2/s) 0.61 0.16 0.05 0.11
for ammonia production (55%), fuel refining (25%), and methanol production (10%). Other
users include the glass, iron and steel, electronics, food, and chemicals industries, account-
ing for about 10% of hydrogen use. In 2018, the hydrogen generation market was expected
to be valued at $135.5 billion and grow further to $199 billion by 2023 (Markets and Markets
Report 2018), showing an annual growth rate of 8% during this period.
The advantages of hydrogen fuel in transportation include that it can be used directly as
a transportation fuel, unlike solar or wind energy. With high octane number, good burning
speeds, and its pollution‐free status, hydrogen can be a great alternative transportation fuel
(Table 4.1). It has much wider limits of flammability in air (4–75% by volume) than meth-
ane (5.3–15% by volume) and gasoline (1–7.6% by volume) (Das et al. 2017).
4.3 Biomass Importance and Contribution to Global

Energy Supply
Biomass is the organic material derived from plants or animals, such as crop residues,
waste wood, crop processing wastes, municipal waste, algae, etc. that is not fossilized.
Humans have been using biomass since the beginning of history for heat energy produc-
tion through direct combustion, and even now it accounts for 90% of bioenergy use (WEC
2016). Forest residue or fuelwood is the oldest form of energy used by humans. Biomass is
now gaining popularity as a renewable energy source, through conversion into other energy
forms like biofuels or biohydrogen, which can easily replace petrol and diesel fuels in vehi-
cles. Biofuels are considered to have net zero CO2 emission because it is believed that only
the CO2 absorbed during its growth is emitted during its use. Due to scarcity of fossil fuels
and the increasing environmental concerns, the use of abundant renewable sources for
energy/fuel production will become inevitable. Potentially the largest and most sustainable
renewable resource (Jekayinfa and Scholz 2009), in 2012 bioenergy accounted for 14% of
global energy use, with approximately 2.6 billion population dependent on biomass for
traditional uses (World Energy Resources 2016).
The largest renewable energy source, biomass now accounts for 10% of global energy
supply. Biomass‐derived energy is generally obtained from sugars or glucose units that
make up the biomass structure (Prasad et al. 2007). Biomass resources for the production
of energy can be categorized as first‐generation biomass (crops and grains), second‐
generation biomass (lignocellulosic wastes), and third‐generation biomass (algae). For food
and energy security concerns, nonfood biomass, also called lignocellulosic biomass, is
mainly taken into consideration for energy production (Olsson and Hahn‐Hagerdal 1996;
Prasad et al. 2007; Alves et al. 2013).
Lignocellulosic biomass may be categorized into the following types.
●● Agriculture residues – includes crop residues such as straw from crops like rice or wheat,
stover of maize or sorghum, etc.
●● Residues from forest trees – harvesting and other operations like pruning lead to genera-
tion of residues such as branches, foliage, and roots which can be utilized for energy
production.
●● Wastes from food processing – kitchen wastes, wastes (solid or liquid streams) from food
processing industries, municipal solid wastes, and market wastes are included under this
section.
The major difficulty in the use of lignocellulosic biomass compared to other biomass is
its recalcitrant structure. Understanding of the chemical conformation and structure of
lignocellulosic biomass is important to develop processes for further production of fuels
and chemicals. Lignocellulosic biomass is made of three polymers – cellulose (36–61%),
hemicellulose (13–39%), and lignin (6–29%) – with the composition of these three com-
ponents varying with the type of biomass, plant species, age, and other factors (Olsson
and Hahn‐Hagerdal 1996; Prasad et al. 2007). These three components are integrated
into a complex heteromatrix (Bajpai 2016), requiring a mechanical/physical/chemical
step (pretreatment) to separate them, so that they become accessible to chemicals,
microbes or enzymes facilitating breakdown into simpler sugars/by‐products which can
be easily converted to the end energy sources (Sheetal et al. 2013). Depending on the
conversion method, the end energy carrier may be gaseous/liquid/solid fuels, heat or
electricity.
Fuel production, be it biofuels or biohydrogen, from lignocellulosic biomass provides
several benefits to the producing nation. Besides helping to meet CO2 targets, biologically
produced fuels help to meet oil demands domestically as they can be produced from region-
ally available materials. Currently, a major part of the generated lignocellulosic biomass is
not utilized but has great potential for use in energy generation activities.
4.4 Hydrogen Production: Feedstock and Production

Technologies
With an annual worldwide availability of about 220 billion tons or about 4500 EJ, on an
energy basis, of primary production, lignocellulosic biomass accounts for the largest uni-
versal energy source (Balat and Kırtay 2010). Considerable focus has been placed on biohy-
drogen production in recent years. The starchy and sugary substrates that comprise the
first‐generation biomass provide greater hydrogen yields compared to second‐ and third‐
generation biomass. However, it is less favored as it competes with nutrition security. In
this chapter, focus has been on the second‐generation biomass or lignocellulosic wastes.
Lignocellulosic biomass, such as residues, forestry wastes, organic wastes (municipal or
industrial), food processing wastes or effluents, provides an environmentally friendly,
renewable, economic and sustainable feedstock for biohydrogen production (Prasad et al.
2007; Jekayinfa and Scholz 2009; Balat and Kırtay 2010). Several materials have been stud-
ied for their hydrogen production, including peanut shells, crop straw, municipal solid
wastes, manure, biogas, etc. (Demirbas 2001; Ntaikou et al. 2010; Alves et al. 2013), through
various production routes in several studies described later. Energy crops (e.g., Miscanthus,
poplar), forestry wastes, and residues may also serve as feedstocks for hydrogen produc-
tion. However, unlike hydrogen from fossil fuels which has an established method, further
refinement in the lignocellulosic biomass‐to‐hydrogen technology is required to help make
it more economical.
Based on the dry weight and hydrogen content of biomass, the hydrogen yields obtained
vary from 16% to 18% and are quite low (Demirbas 2001; Balat and Kırtay 2010). Studies
have looked at different feedstocks and different production routes for hydrogen produc-
tion from biomass, which currently accounts for only 1% of hydrogen production globally.
There are two main processes for the production of biohydrogen: thermochemical and bio-
logical. Thermochemical conversion processes include gasification, direct liquefaction,
pyrolysis and steam reforming of bio‐oils and biogas while biological conversion may be
done generally via fermentation and application of microbial electrolysis. Figure 4.2 shows
the different conversion routes of biomass to hydrogen.
4.4.1 Thermochemical Conversion Technologies

The advantage of thermochemical conversion technologies is their higher conversion effi-
ciency (>50%) and lower production costs compared to other methods (Balat and Kırtay
2010). Among these methods, Meng et al. (2006) observed the superior performance by
pyrolysis and gasification.
Thermochemical processes Biological processes
Fermentation
Direct Biomass Biomass Biogas,
liquefaction pyrolysis gasification bioethanol
Photo- Dark- Biocatalyzed Hybrid

Steam reforming & Water-gas shift reactions fermentation fermentation electrolysis system
H2
Figure 4.2 Hydrogen production methods from lignocellulosic biomass.
4.4.1.1 Direct Liquefaction

This process uses catalytic hydrogenation to convert lignocellulosic biomass anaerobically
at higher temperatures of 525–600 K and pressure of 5–20 MPa, into a mixture of liquid
biofuels, which can be used as fuel or as raw materials for further processing (Stevens 1987;
Meier and Rupp 1991). With a catalyst (e.g., Na2CO3), carbon monoxide and water vapor
convert biomass into a gas and oil mixture with high methane content. However, this
method gives low hydrogen yields, and also the operating conditions are difficult to achieve
and maintain.
4.4.1.2 Biomass Pyrolysis

Pyrolysis is a thermal process of destructive distillation of organic matter. Under anaerobic
conditions, and at high temperatures (650–800 K) and pressure (0.1–0.5 MPa), biomass con-
verts into solid (charcoal/char, other inert materials), liquid (oils, tar) and gaseous (mixture
of H2, CO, CH4, CO2) compounds, in a general ratio of 35:35:30, subject to the organic
composition of the biomass (Jalan and Srivastava 1999). Pyrolysis of lignocellulosic bio-
mass can be performed by either fast or slow pyrolysis. Flash or fast pyrolysis yields higher
quantities of hydrogen (Eq. 4.1) as long as higher temperatures and an adequate volatile
phase residence period are provided for the reaction.
Biomass heat H 2 CH 4 CO other products (4.1)
Hydrocarbon gases and vapors produced may be steam reformed (Eq. 4.2), followed by
water–gas shift reaction (Eq. 4.3) for 5–7% higher biohydrogen yields.
CH 4 H 2O CO 3H 2 (4.2)
CO H 2O CO2 H 2 (4.3)
Table 4.2 Hydrogen yields obtained by pyrolysis in literature.
Feedstock Conditions Hydrogen yield Reference
Olive husk 13% ZnCl2 as catalyst at about 1025 K 70.3% Demirbas

Cotton temperature 59.9% (2001)
cocoon shell 60.3%
Tea waste
Wheat straw 10% NiAl2O3 as catalyst in a two‐stage 16.38 mmol/g Akubo et al.
Palm kernel fixed bed reactor system at 550 °C 25.35 mmol/g (2018)
shells pyrolysis temperature and 750 °C 18.22 mmol/g
Rice husk catalytic bed temperature
22.11 mmol/g
Coconut shell 22.96 mmol/g
Sugarcane 20.74 mmol/g
bagasse
Cotton stalk
Rice straw Cr2O3 as catalyst 49.5% of gas vol. Chen et al.
Sawdust 51.4% of gas vol. (2003)
The bio‐oils produced along with the gases can also be treated for production of hydro-
gen. The bio‐oil is composed of two fractions based on water solubility: a monomer‐rich
water‐soluble fraction and a lignin‐derived oligomer‐rich water‐insoluble fraction, both of
which can be used in steam reforming reactions for hydrogen production.
Selection of appropriate reactor type (ablative, fluidized bed, circulating fluidized bed,
entrained flow), catalysts and heat transfer modes helps to regulate temperatures, heating
rates, and residence time which influence product yields (Bridgwater 1999; Balat and
Kırtay 2010; Garcia‐Nunez et al. 2017; Sobamowo and Ojolo 2018). Among different reac-
tors for biomass pyrolysis, studies show that the fluidized bed reactor seems to be the most
suitable for hydrogen production. The maximum hydrogen yields could touch 90% in
experiments when a Ni‐based catalyst was used, which may be improved by further steam
reforming and water–gas shift reactions. Table 4.2 lists some studies done by pyrolysis.
4.4.1.3 Lignocellulosic Biomass Gasification

Gasification is a two‐stage thermochemical reaction consisting of oxidation and reduction
processes under controlled volumes of oxygen and/or steam. Gasification of biomass can
be achieved at high temperatures (>1000 K), wherein it undergoes partial oxidation giving
a mixture of charcoal and gas. The charcoal is then reduced to a mix of hydrogen, carbon
oxides (CO, CO2), and methane (called synthesis gas or syngas or product gas). This conver-
sion process can be represented as:
Biomass steam heat H 2 CO CO2 CH 4 hydrocarbons char (4.4)
As discussed in the previous section, the product gases can be further reacted through
steam reforming and water–gas shift reactions for higher yields. As the products are
predominantly vapors, the gasification process is more promising than pyrolysis for the
production of biohydrogen.
Gasification reactors can be divided into several types (autothermal gasifiers, allothermal
gasifiers, fluidized‐bed reactors, entrained flow reactors), depending on the method of heat
transfer and the gasification medium (Hannula 2009). Supercritical water gasification is
another process which promises to solve key issues in biomass gasification and is ideal for
wet biomass containing as much as 99% water, eliminating the need to dry materials prior
to processing. The normal gasification process requires drying of biomass to a moisture
content below 35% (Demirbas 2002).
One of the major limitations to the gasification process is the formation of tar and ash
during the process. The undesirable tar fraction results in the formation of tar aerosols
and polymerization to a more composite structure, which may block heat exchangers or
pipes and reduce hydrogen production during steam reforming (Mishra et al. 2015;
Sikarwar et al. 2016). Studies have helped identify ways in which tar formation may be
reduced such as maintenance of optimum operating parameters like temperature, gasify-
ing agent and residence time inside the gasifier, plus proper design of gasifier. Temperatures
above 1273 K are reported to thermally crack tar (Sikarwar et al. 2016). Use of additives
such as char, dolomite, olivine, etc. reduces tar formation inside the gasifier. Dolomite is
the best eliminator of tar. The addition of catalysts (such as Ni‐based catalysts, nonmetal-
lic oxides, alkaline metal oxides) also reduces tar formation as well as increasing gas
yields, quality, and conversion efficiency (Corella et al. 1999). Tar reduction can also be
possible by a slight modification in methodology by two‐stage gasification and secondary
air injection in the gasifier. Ash deposition is also a problem that can be resolved by frac-
tionation and leaching. Fractionation may decrease ash quality, while leaching improves
ash quality by removing inorganic fraction from the biomass. Table 4.3 looks at some
studies done by pyrolysis.
Table 4.3 Hydrogen yields obtained by gasification in literature.
Feedstock Reaction conditions H2 yield Reference
Pinewood Conical spouted bed reactor, and further 117 g/kg Arregi et al.
sawdust steam reforming of the pyrolysis vapors in a biomass (2016)
fluidized bed reactor on a Ni‐based catalyst
Pine Continuous‐feed supercritical water 41.7% Faires (2003)
gasification at 800 K and 5% biomass
concentration
Palm kernel Fluidized‐bed gasifier 5.52% dry wt. Ghani et al.
Coconut shell 5.04% dry wt. (2009)
Sawdust Downdraft reactor at 870 °C 35.39% vol. Pengmei
et al. (2007)
Sawdust Fluidized‐bed gasifier 850 °C 57.4% vol. Turn et al.
(1998)
Rice straw Air gasification 103 MNm3 Cardenas
Woody biomass 409 MNm3 et al. (2007)
Olive oil residue 24 MNm3
4.4.1.4 Catalytic Steam Reforming of Biogas and Bioethanol

Alternate sources like biogas, produced by anaerobic decomposition of lignocellulosic bio-
mass have high potential as a versatile feedstock for biohydrogen production, besides help-
ing to reduce GHGs by utilizing the evolved methane (Alves et al. 2013). Biogas comprises
a methane‐rich gas mixture, which can be steam reformed at high temperatures (500–
950 °C) in the presence of the catalyst to yield hydrogen gas (Doucek et al. 2007), via the
following reactions:
CH 4 H 2O 3H 2 CO CO2 (4.5)
CH 4 2H 2O 4H 2 CO2 (4.6)
The CO can be further reacted to yield H2 via water–gas shift reaction.

Bioethanol is also produced from lignocellulosic biomass. In a similar manner as
described above, steam reforming can be used to convert bioethanol to hydrogen (Eq. 4.7).
C 2 H 5OH 3H 2O 6H 2 2CO2 (4.7)
4.4.1.5 Hydrogen Separation

Hydrogen gas production from thermochemical processing of biomass is normally coupled
with production of other gaseous mixtures. Consequently, hydrogen gas needs to be sepa-
rated and purified for its final use. Numerous methods have been successfully developed
for this process, including drying/chilling, CO2 absorption, pressure swing adsorption, and
membrane separation (Sircar and Golden 2000; Adhikari and Fernando 2006; Uehara
2013). It can be anticipated that the coming years will see a surge in large‐scale commercial
production of hydrogen, with the availability of newly developed gasifiers and other
technologies.
4.5 Biological Hydrogen Production Technologies
Biohydrogen evolution as a byproduct of microbial metabolism has gained momentum in

recent years for the production of sustainable energy from renewable resources (Balat and
Kırtay 2010). These processes are more eco‐friendly and consume less energy compared to
thermochemical processes. Biomass availability and cost, as well as the content of carbohy-
drates and fermentability, are major factors that influence the suitability of biomass
(Kapdan and Kargi 2006; Argun et al. 2016).
Biohydrogen production from lignocellulosic biomass involves two steps: pretreatment
followed by hydrogen production. Pretreatment is recognized as an essential requirement
which breaks the recalcitrant crystalline lignocellulosic biomass structure, loosens the fib-
ers, breaks the lignin seals, and creates monomers which are accessible to enzyme attack
(by enzyme or microbes) in order to obtain higher hydrogen yields (Argun et al. 2016). This
may be achieved by physical, chemical or biological processes, following a size reduction
process. Particle size has been observed as a factor affecting hydrogen generation, with
studies suggesting that a reduction in particle size enhances hydrogen formation (Yuan
et al. 2011; Argun et al. 2016). However, one disadvantage of pretreatment, mainly the
chemical treatment methods, is the formation of inhibitory by‐products such as furfural,

hydroxymethylfurfural (HMF), and phenolics, which have been observed by several
researchers to inhibit microbial activity subsequently (Ntaikou et al. 2010) and might
require a detoxification step to reduce the concentration of toxic compounds. Another
challenge in producing hydrogen from lignocellulosic biomass is the high cost of pretreat-
ment. Dark and photofermentation are the major biohydrogen‐producing processes using
microorganisms.
4.5.1 Photofermentation
Photofermentation requires optimization and maintenance of strict environmental condi-
tions (light source type, light intensity, lighting regime) and media composition (Argun
and Kargi 2010). In addition, essential elements which need to be added to the fermenta-
tion media include Fe and Mo for better hydrogen yield (Kars and Ceylan 2013). Hydrogen
evolution by photosynthetic bacteria is mediated by nitrogenase activity. Purple nonsulfur
bacteria are the main hydrogen‐producing photofermentative microorganisms: Rhodobacter
sphaeroides, Rhodobacter capsulatus, Rhodospirillum rubrum, etc. (Argun et al. 2016).
Organic acids such as acetate, butyrate, and lactate are the main substrates for photofer-
mentative hydrogen production (Doucek et al. 2007; Keskin et al. 2011; Kars and Ceylan
2013). A wide variety of effluents rich in organic acids can be used in this process. Azwar
et al. (2014) noted that although theoretically high yields can be obtained from the process,
low light conversion efficiency (3–10%) and production volumes may be obstacles in pho-
tofermentation. Hydrogen generation rates of the order of 145–160 mmol/h/L have been
reported in the literature (Levin et al. 2004). Budiman and Wu (2018) have reported that
certain chemicals like iron, molybdenum, EDTA, vitamins, buffer solution, etc. have sig-
nificant beneficial effects on biohydrogen production rates. Table 4.4 shows the hydrogen
yields obtained by photofermentation in the literature.
4.5.2 Dark Fermentation

Fermentation is one of the best routes for sustainable biohydrogen production. Dark fer-
mentation is the most investigated approach among these. Nandi and Sengupta (1998) state
that research on hydrogen production using anaerobic bacteria started in the 1980s. Simple
operations and higher production rates make it superior to photosynthetic processes
(Chong et al. 2009). Glucose is the basic compound of lignocellulosic biomass that gets
converted to hydrogen and theoretically, conversion of 1 mol of glucose (C6H12O6) yields
12 mol of hydrogen. However, the final yield depends on the fermentation pathway and
end‐products (Ntaikou et al. 2010; Sveinsdottir et al. 2011). More than 80% of end‐products
are made of acetate and butyrate (Balat and Kırtay 2010). Other end‐products of fermenta-
tion include ethanol and lactic acid which are nonhydrogen forming and propionic acid
which consumes hydrogen (Argun et al. 2016). For example, the theoretical yield of hydro-
gen if acetic acid and butyric acid are the final products follows Eqs (4.8) and (4.9),
respectively:
C6 H12O6 4H 2O 2CH3COO 2HCO3 4H 2 4H (4.8)

Table 4.4 Hydrogen yields obtained by photofermentation in literature.
Feedstock Culture H2 yield Reference
Corncob Rhodospirillum rubrum, Rhodobacter 229 mmol Zhang et al.

Sorghum stover capsulatus, Rhodopseudomonas H2/L (2014)
Corn stover palustris 149.67
Rice straw 145.67
Soybean stalk 140.45
Cotton stalk 131.12
118.46
Wheat straw R. capsulatus‐PK 372 mL H2/L Mirza et al.
(2013)
Cassava wastewater Mixed bacterial consortium 36.1 mmol Lazaro et al.
H2/L culture (2015)
Combined palm oil Rhodobacter sphaeroides NCIMB 8253 8.72 mL H2/ Budiman and
plus pulp and paper mL medium Wu (2016)
mill effluent
Sugar beet molasses R. capsulatus JP91 10.5 mol H2/ Keskin and
mol sucrose Hallenbeck
(2012)
Apple waste Mixed bacterial consortium 11.85 mol Lu et al. (2016)
H2/g TS
Corn stalk pith Mixed bacterial consortium 2.61 mol H2/ Jiang et al.
mol glucose (2016)
C6 H12O6 2H 2O CH3CH 2CH 2COO 2HCO3 2H 2 3H (4.9)
However, nearly 2.0–2.5 mol of hydrogen yield from 1 mol of glucose is achieved (Kapdan
and Kargi 2006), which is less than theoretical yield estimations. The low yield of hydrogen
might be due to the production of a mix of acetate and butyrate reaching to almost 2.5 mol
of hydrogen/mol of glucose yield, production of nonhydrogen‐forming or hydrogen‐
consuming end‐products, or the utilization of feedstock substrate as an energy source for
microbial growth rather than expected organic acid formation (Argun et al. 2016).
The fermentation process can be done through a wide array of microorganisms (e.g.,
Clostridium spp., Enterobacter, Thermoanaerobacterium spp., Thermotoga spp.), which
vary in their substrate, pH and temperature requirements, and subsequently the metabolic
pathway followed and hydrogen yields obtained (Ntaikou et al. 2010). Studies done on fer-
mentation processes claim a higher yield of hydrogen using extremophilic bacteria reported
to produce up to 4 mol of hydrogen and 2 mol of acetate in pure cultures due to improved
reaction kinetics at high temperatures, resistance to higher hydrogen partial pressure in
media, and less sensitivity to contaminants, while mesophilic and moderate thermophilic
bacteria produce reduced by‐products such as ethanol, lactic or butyric acids under high
hydrogen concentrations in media (Sveinsdottir et al. 2011).
Dark fermentation uses anaerobic bacteria to convert organic matter into hydrogen,
along with other by‐products like carbon dioxide, organic acids, and solvents, which
a ccumulate in the medium as it cannot be used by these anaerobes, thus lowering

medium pH and hampering attainment of theoretically possible maximum hydrogen
yields (Brar and Sarma 2013). Also, the by‐products of the initial pretreatment given to
biomass such as furfural and HMF are observed to inhibit microbial growth (Sveinsdottir
et al. 2011). Several studies have been done to analyze the hydrogen production poten-
tial by fermentation. Table 4.5 lists studies on hydrogen yields obtained from various
lignocellulosic biomass after various pretreatment conditions using the dark fermenta-
tion process.
A disadvantage of the dark fermentation process is that the organic load in the substrate
remains more or less in the medium itself. Also, a remarkable quantity of hydrogen also
remains trapped in these compounds, resulting in lower hydrogen yields (Ntaikou et al.
2010). Methods to utilize more of the remaining organic matter in the medium, either
through a second stage for hydrogen extraction or generation of high‐value products (e.g.,
polyhydroxyalkaonates) from effluents, may improve the energy output and economics
positively.
4.5.3 Two-Stage Hydrogen Production

A hybrid system involving both dark and photofermentation is said to be a suitable remedy
for the drawbacks faced by the individual processes. Here, the end‐products of the first
dark fermentation phase (Eq. 4.10) are utilized in the second photofermentative phase (Eq.
4.11), leading to higher production potential.
C 6 H12O6 2H 2O 4H 2 2CO2 2CH3COOH (4.10)
CH3COOH 2H 2O light energy 4H 2 2CO2 (4.11)
Anaerobic bacteria produce organic acids, energy, and electrons through biomass degra-
dation. Photosynthetic bacteria use these organic acids in the presence of light energy to
produce hydrogen. However, the sensitivity of photofermentative organisms to higher
organic acids and ammonia levels in media and lower photosynthetic efficiency requiring
larger surface area and cost are factors to be taken into consideration during the operation
of this dual system (Ntaikou et al. 2010). Hydrogen yield obtained by the hybrid system is
shown in Table 4.6.
4.5.4 Biocatalyzed Electrolysis

The acetate‐rich effluent from dark fermentation can be used for further hydrogen produc-
tion using this system. Microbial electrolysis cell (Figure 4.3) is another promising
approach, in which a microbial reactor acting as the anode chamber generates protons,
electrons (absorbed by anode), and other by‐products (Eq. 4.12). Application of minute
electric currents (around 0.2 V) reduces the generated protons to hydrogen (Eq. 4.13).
Anode : CH3COOH 2H 2O 2CO2 8H 8e (4.12)
Cathode : 8H 8e 4H 2 (4.13)
Table 4.5 Hydrogen yields obtained by dark fermentation in literature.
H2 yield (mol H2 mol/

Feedstock Culture glucose equiv.) Reference
Corn stover Thermoanaerobacterium thermosaccharolyticum 2.24 Cao et al. (2009)

Mixed 1.53 Liu and Cheng (2010)
Napier grass Mixed 1.2 Lo et al. (2009)
Wheat straw Caldicellulosiruptor saccharolyticus 3.8 Ivanova et al. (2009)
Mixed 1.0–2.54 Kongjan and Angelidaki (2010)
Kongjan et al. (2010)
Clostridium sp. IODB03 2.52 mol/mol sugar Patel et al. (2015)
Bacillus coagulans NCIM 2323 and Enterobacter 0.23–1.40 mol/mol Kumar et al. (2014)
aerogens NCIM5139 glucose
Barley hulls Clostridium thermocellum 1.24 Magnusson et al. (2008)
Grass Clostridium AK14 0.8–0.9 Almarsdottir et al. (2010)
Food waste Mixed 0.6–2.4 Shin et al. (2004); Shin and Youn (2005)
Vegetable waste Mixed 1.7 Lee et al. (2010)
Miscanthus Thermotoga elfii 1.1 de Vrije et al. (2002)
T. neapolitana 3.2 de Vrije et al. (2009)
Caldicellulosiruptor saccharolyticus 3.4 de Vrije et al. (2009)
Sweet sorghum Rumicococcus albus 3.15 Ntaikou et al. (2008)
stalk
Bagasse Mixed 13.39 Chairattanamanokorn et al. (2009)
Maize leaves C. saccharolyticus 3.6 Ivanova et al. (2009)
Rice straw Heat‐treated sludge 0.44 mol/mol sugar Liu et al. (2013)
Soybean straw Clostridium buytricum 47.65 mL/g substrate Han et al. (2012)
c04.indd 59 11/12/2019 6:01:59 PM

Table 4.6 Hydrogen yields obtained by the hybrid system in literature.
Feedstock H2 yield Reference
Beet molasses 13.7 mol H2/mol of sucrose Özgür et al. (2010)

Sugarcane bagasse 1753 mL H2/L Rai et al. (2014)
Arthrospira platensis biomass 337.0 mL H2/g DW Cheng et al. (2012)
Sugarcane bagasse 26–35.7% vol. Hema and Pushpa (2012)
Corncob 713.6 ml H2/g COD Yang et al. (2010)
Water hyacinth 751.5 ml H2/g TVS Cheng et al. (2013)
COD, chemical oxygen demand; DW, dry weight; TVS, total volatile solids.
e– Power e–
CO2 Supply
H2
H+
Bacteria
Anode Cathode
PEM
Figure 4.3 Biocatalyzed electrolysis.
Using this process, end‐products like acetate from fermentative processes, when used as
the substrate, give eight hydrogen molecules in place of four obtained via fermentation
(Geelhoed et al. 2010; Brar and Sarma 2013). Studies by Liu et al. (2005) and Rozendal et al.
(2006) noted yields up to 73% and 53% with an external supply of 250 mV and 500 mV,
respectively.
4.6 Production Costs
Steam methane reforming (SMR) is currently the most popular and least expensive method
for generating hydrogen from natural gas. Chemically, the content of hydrogen (H) in bio-
mass is comparatively less (6–6.5%), compared to nearly 25% in natural gas. As the material
has a low content for generating hydrogen, it cannot compete on a cost basis with the well‐
developed commercial technology for SMR of natural gas. Table 4.7 presents a comparison
of hydrogen production cost via various methods.
Table 4.7 Comparison of hydrogen production parameters via different methods.
Conversion
Process efficiency (%) Production cost References
Natural gas reforming 75 0.45 US$/Nm3 H2 Reith et al. (2003)

Steam reforming of biomethane ~20 0.45 US$/Nm3 H2 Reith et al. (2003)
Photobiological hydrogen ~10 ~10 US$/MBTU Benemann (1997)
Two‐stage bioprocess from 15 0.35 US$/ Nm3 H2 Reith et al. (2003)
biomass
Fermentative hydrogen ~22 ~40 US$/MBTU Das and Veziroglu
(2001)
Hydrogen from coal/biomass NA 4 US$/MBTU Bockris (1981)
Biomass pyrolysis NA 8.86–15.52 US$/ Padro and Putsche
GJ (1999)
Wind energy‐based electrolysis NA 20.2 US$/GJ Mahishi et al. (2014)
PV‐based electrolysis NA 41.8 US$/GJ Mahishi et al. (2014)
GJ, gigajoule; MBTU, million British thermal units; NA, not applicable; PV, photovoltaic.
An integrated process, in which a portion of the biomass is used to produce more valua-
ble substances or chemicals and the residual fractions are utilized to generate hydrogen,
may be an economically viable option (Balat and Kırtay 2010).
4.7 Conclusion
Biohydrogen production from sources like lignocellulosic feedstocks is an exciting area of

renewable energy development. Technologic advancement offers the viable production of
useable hydrogen from renewable bioresources to satisfy the world’s energy needs.
Although hydrogen production has reached great heights, progress in its production, espe-
cially from lignocellulosic biomass, has not kept pace with other developments in this field
like fuel cells or durability. A major limitation is the low rates and yields of hydrogen from
biomass. The hydrogen production process constraints or barriers are well understood, and
intense investigations are under way worldwide to surmount these barriers and produce
biohydrogen which is competitive with conventionally produced fuels.
References
Adhikari, S. and Fernando, S. (2006). Hydrogen Membrane Separation Techniques. Ind. Eng.
Chem. Res. 45 (3): 875–881.
Akubo, K., Nahil, M.A., and Williams, P.T. (2018). Pyrolysis‐catalytic steam reforming of
agricultural biomass wastes and biomass components for production of hydrogen/syngas.
J. Energy Inst. https://doi.org/10.1016/j.joei.2018.10.013.
Almarsdottir, A.R., Taraceviz, A., Gunnarsson, I., and Orlygsson, J. (2010). Hydrogen
production from sugars and complex biomass by Clostridium species AK14, isolated from
Icelandic hot spring. Icel. Agric. Sci. 23: 61–71.
Alves, H.J., Junior, C.B., Niklevicz, R.R. et al. (2013). Overview of hydrogen production
technologies from biogas and the applications in fuel cells. Int. J. Hydrog. Energy 38 (13):
5215–5225.
Argun, H., Gokfiliz, P., and Karapinar, I. (2016). Biohydrogen production potential of
different biomass sources. In: Biohydrogen Production: Sustainability of Current
Technology and Future Perspective (eds. A. Singh and D. Rathore). Dordrecht:
Springer Publications.
Argun, H. and Kargi, F. (2010). Effects of light source, intensity and lighting regime on
bio‐hydrogen production from ground wheat starch by combined dark and photo‐
fermentation. Int. J. Hydrog. Energy 35: 1604–1612.
Arregi, A., Lopez, G., Amutio, M. et al. (2016). Hydrogen production from biomass by
continuous fast pyrolysis and in‐line steam reforming. RSC Adv. 6 (31): 25975–25985.
Azwar, M.Y., Hussain, M.A., and Abdul‐Wahab, A.K. (2014). Development of biohydrogen
production by photobiological, fermentation and electrochemical processes: a review.
Renew. Sust. Energ. Rev. 31: 58–173.
Bajpai, P. (2016). Pretreatment of Lignocellulosic Biomass for Biofuel Production. Dordrecht:
Springer.
Balat, H. and Kırtay, E. (2010). Hydrogen from biomass present scenario and future prospects.
Int. J. Hydrog. Energy 35: 7416–7426.
Benemann, J.R. (1997). Feasibility analysis of photobiological hydrogen production. Int. J.
Hydrog. Energy 22: 979–987.
Bockris, J.O.M. (1981). The economics of hydrogen as a fuel. Int. J. Hydrog. Energy 6: 223–241.
Brar, S.K. and Sarma, S.J. (2013). Bio‐hydrogen production: a promising strategy for organic
waste management. Hydrol. Current Res. S5: e002.
Bridgwater, A.V. (1999). Principles and practice of biomass fast pyrolysis processes for liquids.
J. Anal. Appl. Pyrolysis 51: 3–22.
Budiman, P.M. and Wu, T.Y. (2016). Ultrasonication pre‐treatment of combined effluents from
palm oil, pulp and paper mills for improving photo‐fermentative biohydrogen production.
Energy Convers. Manag. 119: 142–150.
Budiman, P.M. and Wu, T.Y. (2018). Role of chemicals addition in affecting biohydrogen
production through photofermentation. Energy Convers. Manag. 165: 509–527.
Cao, G.L., Ren, N.Q., Wang, A.J. et al. (2009). Effect of lignocellulose‐derived inhibitors on
growth and hydrogen production by Thermoanaerobacterium thermosaccharolyticum W16.
Cardenas, R., Alfonso, D., Penalvo, E. et al. (2007). Potential for hydrogen production from
biomass residues in the Valencian community. Conference proceedings. http://conference.
ing.unipi.it/ichs2007/fileadmin/user_upload/CD/PAPERS/13SEPT/2.1.98.pdf.
Chairattanamanokorn, P., Penthamkeerati, P., Reungsang, A. et al. (2009). Production of
biohydrogen from hydrolyzed bagasse with thermally preheated sludge. Int. J. Hydrog.
Energy 34: 7612–7617.
Chen, G., Andries, J., and Spliethoff, H. (2003). Catalytic pyrolysis of biomass for hydrogen‐
rich gas fuel production. Energy Convers. Manag. 44: 2289–2296.
Cheng, J., Xia, A., Liu, Y. et al. (2012). Combination of dark‐ and photo‐fermentation to
improve hydrogen production from Arthrospira platensis wet biomass with ammonium
removal by zeolite. Int. J. Hydrog. Energy 37 (18): 13330–13337.
Cheng, J., Xia, A., Su, H. et al. (2013). Promotion of H2 production by microwave‐assisted
treatment of water hyacinth with dilute H2SO4 through combined dark fermentation and
photofermentation. Energy Convers. Manag. 73: 329–334.
Chong, M.L., Sabaratnam, V., Shirai, Y., and Hassan, M.A. (2009). Biohydrogen production from
biomass and industrial wastes by dark fermentation. Int. J. Hydrog. Energy 34: 3277–3287.
Corella, J., Aznar, M.P., Gil, J., and Caballero, M.A. (1999). Biomass gasification in fluidized
bed: where to locate the dolomite to improve gasification. Energy Fuel 13: 1122–1127.
CryoGas International. https://www.gasworld.com.
Das, D., Khanna, N., and Dasgupta, C.N. (2017). Biohydrogen Production: Fundamentals and
Technology Advances. Boca Raton, FL: CRC Press.
Das, D. and Veziroglu, T.N. (2001). Hydrogen production by biological processes: a survey of
literature. Int. J. Hydrog. Energy 26: 13–28.
Demirbas, A. (2001). Yields of hydrogen‐rich gaseous products via pyrolysis from selected
biomass samples. Fuel 80 (13): 1885–1891.
Demirbas, A. (2002). Hydrogen production from biomass by the gasification process. Energy
Sources 24: 59.
Demirbas, A. (2009). Biohydrogen for Future Engine Fuel Demands. London: Springer.
Doucek, A., Prokes, O., and Tenkrat, D. (2007). New trends in hydrogen production from
biomass. Energy z biomass VII. – Odborny seminar: 39–44.
Faires, K.B. (2003). Gasification of in‐forest biomass residues. Thesis. University of
Washington.
Garcia‐Nunez, J.A., Pelaez‐Samaniego, M.R., Garcia‐Perez, M.E. et al. (2017). Historical
developments of pyrolysis reactors: a review. Energy Fuel 31 (6): 5751–5775.
Geelhoed, J.S., Hamelers, H.V.M., and Stams, A.J.M. (2010). Electricity‐mediated biological
hydrogen production. Curr. Opin. Microbiol. 13: 307–315.
Ghani, W.A.W.A.K., Moghadam, R.A., Mohd Salleh, M.A., and Alias, A.B. (2009). Air
gasification of agricultural waste in a fluidized bed gasifier: hydrogen production
performance. Energies 2: 258–268.
Han, H., Wei, L., Liu, B. et al. (2012). Optimization of biohydrogen production from soybean
straw using anaerobic mixed bacteria. Int. J. Hydrog. Energy 37: 13200–13208.
Hannula, I. (2009). Hydrogen production via thermal gasification of biomass in near‐to‐
medium term. VTT Working Papers 131. VTT Technical Research Centre of Finland.
Hema, R. and Pushpa, A. (2012). Production of clean fuel from waste biomass using combined
dark and Photofermentation. IOSR J. Comput. Eng. 1 (4): 39–47.
IEA (2018). www.iea.org/tcep/energyintegration/hydrogen.
IRENA (2018). Hydrogen from renewable power: technology outlook for the energy transition.
International Renewable Energy Agency, Abu Dhabi. www.irena.org.
Ivanova, G., Rakhely, G., and Kovács, K.L. (2009). Thermophilic biohydrogen production from
energy plants by Caldicellulosiruptor saccharolyticus and comparison with related studies.
Jalan, R.K. and Srivastava, V.K. (1999). Studies on pyrolysis of a single biomass cylindrical
pellet‐kinetic and heat transfer effects. Energy Convers. Manag. 40: 467.
Jekayinfa, S.O. and Scholz, V. (2009). Potential availability of energetically usable crop residues
in Nigeria. Energy Sources Part A 31: 687–697.
Jiang, D., Ge, X.M., Zhang, T. et al. (2016). Photo‐fermentative hydrogen production from
enzymatic hydrolysate of corn stalk pith with a photosynthetic concortium. Int. J. Hydrog.
Energy 41: 16778–16785.
Kapdan, I.K. and Kargi, F. (2006). Bio‐hydrogen production from waste materials. Enzym.
Microb. Technol. 38: 569–582.
Kars, G. and Ceylan, A. (2013). Biohydrogen and 5‐aminolevulinic acid production from waste
barley by Rhodobacter sphaeroides O.U.001 in a biorefinery concept. Int. J. Hydrog. Energy
38: 5573–5579.
Keskin, T., Abo‐Hashesh, M., and Hallenbeck, P.C. (2011). Photofermentative hydrogen
production from wastes. Bioresour. Technol. 102: 8557–8568.
Keskin, T. and Hallenbeck, P.C. (2012). Hydrogen production from sugar industry wastes using
single‐stage photofermentation. Bioresour. Technol. 112: 131–136.
Kongjan, P. and Angelidaki, I. (2010). Extreme thermophilic biohydrogen production from
wheat straw hydrolysate using mixed culture fermentation: effect of reactor configuration.
Bioresour. Technol. 101: 7789–7796.
Kongjan, P., O‐Thong, S., Kotay, M. et al. (2010). Biohydrogen production from wheat straw
hydrolysate by dark fermentation using extreme thermophilic mixed culture. Biotechnol.
Bioeng. 105: 899–908.
Kumar, S., Prasad, S., and Gupta, N. (2014). Hydrogen production potential of wheat straw by
enzymatic saccharification and fermentation. In: Strategic Technologies Complex
Environmental Issues: A Sustainable Approach (ed. G.C. Mishra). New Delhi: Excellent
Publishing House.
Lazaro, C.Z., Bosio, M., dos Santos, F.J. et al. (2015). The biological hydrogen production
potential of agroindustrial residues. Waste Biomass Valoriz. 6: 273–280.
Lee, Z., Li, S., Kuo, P. et al. (2010). Thermophilic bio‐energy process study on hydrogen
fermentation with vegetable kitchen waste. Int. J. Hydrog. Energy 35: 13458–13466.
Levin, D.B., Pitt, L., and Love, M. (2004). Biohydrogen production: prospects and limitations to
practical application. Int. J. Hydrog. Energy 29: 173–185.
Liu, C. and Cheng, X. (2010). Improved hydrogen production via thermophilic fermentation of
corn stover by microwave‐assisted acid pretreatment. Int. J. Hydrog. Energy 35 (17): 8945–8952.
Liu, C.M., Chu, C.Y., Lee, W.Y. et al. (2013). Biohydrogen production evaluation from rice
straw hydrolysate by concentrated acid pre‐treatment in both batch and continuous systems.
Liu, H., Grot, S., and Logan, B. (2005). Electrochemically assisted microbial production of
hydrogen from acetate. Environ. Sci. Technol. 39: 4317–4320.
Lo, Y.C., Huang, C.Y., Fu, T.N. et al. (2009). Fermentative hydrogen production from
hydrolyzed cellulosic feedstock prepared with a thermophilic anaerobic bacterial isolate. Int.
J. Hydrog. Energy 34 (15): 6189–6200.
Lu, C., Zhang, Z., Ge, X. et al. (2016). Bio‐hydrogen production from apple waste by
photosynthetic bacteria HAU‐M1. Int. J. Hydrog. Energy 41: 13399–13407.
Magnusson, L., Islam, R., Sparling, R. et al. (2008). Direct hydrogen production from cellulosic
waste materials with a single‐step dark fermentation process. Int. J. Hydrog. Energy 33:
5398–5403.
Mahishi, M., Goswami, D.Y., Ibrahim, G., and Elnashaie, S.S.E.H. (2014). Hydrogen production
from biomass and fossil fuels. In: Handbook of Hydrogen Energy (eds. S.A. Sherif, D.Y.
Goswami, E.K. Stefanakos and A. Steinf), 113–137. Boca Raton, FL: CRC Press.
Markets and Markets Report (2018). Hydrogen Generation Market by Generation, Application
(Petroleum Refinery, Ammonia Production, Methanol Production, Transportation, Power
Generation), Technology (Steam Reforming, Water Electrolysis, & Others), Storage, and
Region – Global Forecast to 2023. www.researchandmarkets.com/reports/4669166/
hydrogen‐generation‐market‐by‐generation.
Meier, D. and Rupp, M. (1991). Direct catalytic liquefaction technology of biomass: status and
review. In: Biomass Pyrolysis Liquids Upgrading and Utilization (eds. A.V. Bridgwater and G.
Grassi). Dordrecht: Springer.
Meng, N., Leung, D.C., Leung, M.K.C., and Sumathy, K. (2006). An overview of hydrogen
production from biomass. Fuel Process. Technol. 87: 461–472.
Mirza, S.S., Iqbal, Q.J., Zhao, Q., and Chen, S. (2013). Photo‐biohydrogen production potential
of Rhodobacter capsulatus‐PK from wheat straw. Biotechnol. Biofuels 6 (1): 144–156.
Mishra, A.K., Singh, R.N., and Mishra, P.P. (2015). Effect of biomass gasification on
environment. Mesop. Environ. J. 1 (4): 39–49.
Mohan, S.V., Bhaskar, Y.V., and Sarma, P.N. (2007). Biohydrogen production from chemical
wastewater treatment in biofilm configured reactor operated in periodic discontinuous
batch mode by selectively enriched anaerobic mixed consortia. Water Res. 41: 2652–2664.
Nandi, R. and Sengupta, S. (1998). Microbial production of hydrogen – an overview. Crit. Rev.
Microbiol. 24: 61–84.
NHEB (2007). National Hydrogen Energy Road Map – Path Way for Transition to Hydrogen
Energy for India. National Hydrogen Energy Board, Ministry of New and Renewable
Energy, Government of India.
Ntaikou, I., Antonopoulou, G., and Lyberatos, G. (2010). Biohydrogen production from
biomass and wastes via dark fermentation: a review. Waste Biomass Valoriz. 1: 21–39.
Ntaikou, I., Gavala, H.N., Kornaros, M., and Lyberatos, G. (2008). Hydrogen production from
sugars and sweet sorghum biomass using Ruminococcus albus. Int. J. Hydrog. Energy 33:
1153–1163.
Olsson, L. and Hahn‐Hagerdal, B. (1996). Fermentation of lignocellulosic hydrolysates for
ethanol production. Enzym. Microb. Technol. 18: 312–331.
Özgür, E., Mars, A.E., Peksel, B. et al. (2010). Biohydrogen production from beet molasses by
sequential dark and photofermentation. Int. J. Hydrog. Energy 35: 511–517.
Padro, C.E.G. and Putsche, V. (1999). Survey of the economics of hydrogen technologies.
Technical Report, NREL/TP_570_27079. National Renewable Energy Laboratory, Golden,
CO, USA.
Patel, A.K., Debroy, A., Sharma, S. et al. (2015). Biohydrogen production from a novel
alkalophilic isolate Clostridium sp. IODB‐O3. Bioresour. Technol. 175: 291–297.
Pengmei, L., Yuan, Z., Ma, L. et al. (2007). Hydrogen‐ rich gas production from biomass air and
oxygen/steam gasification in a downdraft gasifier. Renew. Energy 32: 2173–2185.
Prasad, S., Dhanya, M.S., Gupta, N., and Kumar, A. (2012). Biofuels from biomass: a
sustainable alternative to energy and environment. Biochem. Cell. Arch. 12 (2): 255–260.
Prasad, S., Kumar, A., and Muralikrishna, K.S. (2014). Biofuels production: a sustainable
solution to combat climate change. Indian J. Agric. Sci. 84 (12): 1443–1452.
Prasad, S., Singh, A., and Joshi, H.C. (2007). Ethanol as an alternative fuel from agril,
industrial and urban residues. Resour. Conserv. Recycl. 1854: 1–39.
Prasad, S., Singh, A., and Rathore, D. (2017). Recent advances in biogas production. Chem.
Eng. Process. Technol. 3 (2): 1038.
Prasad, S., Venkatramanan, V., Kumar, S., and Sheetal, K.R. (2019). Biofuels: a clean
technology for environment management. In: Sustainable Green Technologies for
Environmental Management (eds. S. Shah et al.), 219–240. Singapore: Springer Nature
Singapore Pte Ltd.
Rai, P.K., Singh, S.P., Asthana, R.K. et al. (2014). Biohydrogen production from sugarcane
bagasse by integrating dark‐ and photo‐fermentation. Bioresour. Technol. 152: 140–146.
Reith, J.H., Wijffels, R.H., and Barten, H. (2003). Biomethane and Biohydrogen – Status and
Perspective of Biological Methane and Hydrogen Production, 118–125. The Hague: Dutch
Biological Hydrogen Foundation.
Rozendal, R.A., Hamelers, H.V.M., Euverink, G.J.W. et al. (2006). Principle and perspectives of
hydrogen production through biocatalyzed electrolysis. Int. J. Hydrog. Energy 31 (12): 1632–1640.
Sarma, S.J., Brar, S.K., Le Bihan, Y., and Buelna, G. (2013). Bio‐hydrogen production by
biodiesel‐derived crude glycerol bioconversion: a techno‐economic evaluation. Bioprocess
Biosyst. Eng. 36: 1–10.
Shakya, B.D., Aye, L., and Musgrave, P. (2005). Technical feasibility and financial analysis of
hybrid wind‐photovoltaic system with hydrogen storage for cooma. Int. J. Hydrog. Energy 30:
9–20.
Sheetal, K.R., Prasad, S., and Chandel, A.K. (2017). Perspectives on climate change, water and
land use, carbon accumulation, and effects on ecosystem after biofuel implementation in
India. In: Sustainable Biofuels Development in India (eds. A.K. Chandel and R.K.
Sukumaran), 517–540. Dordrecht: Springer International.
Sheetal, K.R., Prasad, S., Gupta, N., and Lata (2013). Evaluation of different pretreatment
processes for sugar recovery from rice straw for ethanol production. Biochem. Cell. Arch. 13
(1): 89–91.
Shin, H.S. and Youn, J.H. (2005). Conversion of food waste into hydrogen by thermophilic
acidogenesis. Biodegradation 16: 33–44.
Shin, H.S., Youn, J.H., and Kim, S.H. (2004). Hydrogen production from food waste in
anaerobic mesophilic and thermophilic acidogenesis. Int. J. Hydrog. Energy 29: 1355–1363.
Sikarwar, V.S., Zhao, M., Clough, P. et al. (2016). An overview of advances in biomass
gasification. Energy Environ. Sci. 9: 2939–2977.
Singh, A. and Rathore, D. (2017). Biohydrogen: next generation fuel. In: Biohydrogen
Production: Sustainability of Current Technology and Future Perspective (eds. A. Singh and D.
Rathore), 1–10. India: Springer.
Sircar, S. and Golden, T.C. (2000). Purification of hydrogen by pressure swing adsorption. Sep.
Sci. Technol. 35: 667–687.
Sobamowo, G.M. and Ojolo, S.J. (2018). Techno‐economic analysis of biomass energy
utilization through gasification technology for sustainable energy production and economic
development in Nigeria. J. Energy www.hindawi.com/journals/jen/2018/4860252/.
Stevens, D.J. (1987). An overview of biomass thermochemical liquefaction research sponsored
by the US DOE. In production, analysis, and upgrading of pyrolysis oils from biomass, ACS
div. Fuel Chem. 32 (2): 223–263.
Sveinsdottir, M., Sigurbjornsdottir, M.A., and Orlygsson, J. (2011). Ethanol and hydrogen
production with thermophilic bacteria from sugars and complex biomass. In: Progress in
Biomass and Bioenergy Production (ed. S. Shaukat). London: InTech Open.
Turn, S., Kinishita, C., Zhang, Z. et al. (1998). An experimental investigation of hydrogen
production from biomass gasification. Int. J. Hydrog. Energy 23: 641–648.
Uehara, I. (2013). Separation and Purification of Hydrogen. Energy Carriers and Conversion
Systems – Vol. I. Encyclopedia of Life Support Systems (EOLSS). www.eolss.net/sample‐
chapters/C08/E3‐13‐05‐01.pdf.
US DoE (2013). Report of the Hydrogen Production Expert Panel: A Subcommittee of the
Hydrogen & Fuel Cell Technical Advisory Committee. United States Department of Energy,
Washington DC. www.hydrogen.energy.gov/pdfs/hpep_report_2013.pdf.
de Vrije, T., Bakker, R.R., Budde, M.A.W. et al. (2009). Efficient hydrogen production from the
lignocellulosic energy crop Miscanthus by the extreme thermophilic bacteria
Caldicellulosiruptor saccharolyticus and Thermotoga neapolitana. Biotechnol. Biofuels 2: 12.
de Vrije, T., de Haas, G.G., Tan, G.B. et al. (2002). Pretreatment of Miscanthus for hydrogen
production by Thermotoga elfii. Int. J. Hydrog. Energy 27: 1381–1390.
WEC (2016). Biomass. www.worldenergy.org/data/resources/resource/biomass.
World Energy Resources (2016). Bioenergy. www.worldenergy.org/wp‐content/
uploads/2017/03/WEResources_Bioenergy _2016.pdf.
Yang, H., Guo, L., and Liu, F. (2010). Enhanced bio‐hydrogen production from corncob by a
two‐step process: dark‐ and photo‐fermentation. Bioresour. Technol. 101: 2049–2052.
Yuan, X., Shi, X., Zhang, P. et al. (2011). Anaerobic biohydro‐ gen production from wheat stalk
by mixed microflora: kinetic model and particle size influence. Bioresour. Technol. 102:
9007–9012.
Zhang, Z., Yue, J., Zhou, X. et al. (2014). Photo‐fermentative bio‐ hydrogen production from
agricultural residue enzymatic hydrolyzate and the enzyme reuse. Bioresources 9:
2299–2310.
69
Recent Advances in the Production of Biodiesel Using

Rahul Bhagat1, Harris Panakkal1, Indarchand Gupta1, and Avinash P. Ingle2
1
Department of Biotechnology, Government Institute of Science, Aurangabad, Maharashtra, India
2
Department of Biotechnology, Engineering School of Lorena, University of São Paulo, Lorena, São Paulo, Brazil
5.1 Introduction
The world is presently confronted with two important crises related to energy: continuous
depletion in our fossil fuel resources and environmental degradation due to emission of
greenhouse gases. Therefore, there is an immediate necessity to research alternative,
renewable, safer and sustainable fuel resources to produce clean energy which should be
nontoxic, efficient, economically viable, and environmentally friendly. The production of
clean energy has attracted a great deal of attention worldwide because it can be used for the
transportation of people and commodities and to perform mechanical work.
It is expected that renewable energy for transportation is likely to grow by 19% until 2023
whereas approximately 15% rise in biofuel production is expected in the same time. Among
biofuels, ethanol production still makes up two‐thirds of total production and the rest is
contributed by biodiesel and hydrotreated vegetable oil (HVO). As far as world biofuel pro-
duction is concerned, Asian countries including India, China, and the ASEAN countries
are expected to contribute huge growth in the same period (IEA 2018).
In this context, liquid biofuels such as bioethanol and biodiesel obtained from renewable
sources are considered as a potential option to address the energy issue and they also reduce
greenhouse gas emissions which cause global warming (Mohammed et al. 2018). Fuels
derived from biological material or biomass are called biofuels and can be used to replace
petrol, diesel, and other fossil‐based fuels. Biofuel derived from natural products has sev-
eral advantages and is considered an attractive, sustainable alternative to fossil fuels
(Ahorsu et al. 2018; IEA 2018). Organic substances used to create biofuels are easily and
naturally broken down and are less toxic than fossil fuels. Moreover, the manufacturing
process for biofuel is much safer than the fossil fuel production process. In addition to the
above‐mentioned concerns about fossil fuel, other key concerns include no proper supply
chain with complete dependency on oil‐exporting countries and volatility in energy prices.
These factors act as a driving force for production as well as consumption of biofuels.

Among the liquid biofuels, in this chapter special emphasis has been given to biodiesel.
Biodiesel is a renewable energy source that can be used for partial or even total replace-
ment of diesel (Ramalingam et al. 2018). Chemically, it consists of monoalkyl esters of
long‐chain fatty acids, obtained through transesterification of biological matter (Lapuerta
et al. 2008). It is considered eco‐friendly as it emits fewer gases than fossil fuels. It reduces
carbon dioxide emission by 78% in comparison with conventional fuel (Nasreen et al.
2018). It is biodegradable and is easily degraded by biological agents within a short period
of time. Biodiesel can be produced from different conventional and nonconventional
sources. Blends of biodiesel can be used in engines without modification or with only slight
modification to achieve fossil fuel‐like performance.
Various microbial sources such as bacteria, fungi, yeasts, and algae have the ability to
accumulate oils (Li et al. 2008; Meng et al. 2009; Musa et al. 2018). Among these, algae are
the most promising source for biodiesel production (Chisti 2010; Liandong et al. 2017). The
biodiesel produced from microbial sources has certain advantages over conventional fuels
in terms of environmental benefits and they are also economically competitive (Sharma
and Singh 2017). On the other hand, there are a few demerits of microbial biodiesel pro-
duction such as higher production cost and advanced methodologies required for large‐
scale biomass production (Arenas et al. 2016).
Currently, biodiesel is produced from several raw materials like oil from rapeseed, can-
ola, soybean, palm, etc. Animal wastes like poultry oil, animal fat, and fish oil are also used
as feedstocks (Adewale et al. 2015; Kudre et al. 2017; Kirubakaran and Selvan 2018). Other
plant‐based biodiesel sources include almond, andiroba, barley, coconut, Jatropha curcus,
kanranja, cumaru, etc. (Pinto et al. 2005; Nasreen et al. 2018). Major biodiesel‐producing
countries utilize different oils as raw material depending on availability. Rapeseed oil and
soybean oil are preferably used as feedstocks for biodiesel production in European coun-
tries and the United States, respectively, whereas coconut oil and palm oil are used as pri-
mary sources in countries like Malaysia and Indonesia. India and Southeast Asian countries
mainly use jatropha tree, karanja and mahua as important sources of biodiesel production
(Atabani et al. 2013; Mohammed et al. 2018).
Challenges associated with biodiesel production technologies include the high cost of
raw materials like oilseeds or animal fats and their lower availability. The cost of biodiesel
mainly depends on raw material costs which account for 60–75% of the total cost of bio-
diesel fuel processing (Katabathini et al. 2007). Industrial‐scale production of biodiesel
largely depends on the availability of vegetable oils, such as soybean and canola. Abundant
production of these raw materials requires excessive land use for oilseed farming (da Costa
Cardoso et al. 2018). All these limitations drive the search for a low‐cost raw material
source with large‐scale supply so as to bridge the gap between the realistic needs of bio-
diesel and current production. A great deal of attention has focused on the use of lignocel-
lulosic biomass for biodiesel production. The biodiesel obtained from lignocellulosic
biomass can replace transportation fuels such as petroleum, diesel, and gasoline and pro-
vide viable options for improving energy security and reducing greenhouse emissions
(Wyman 1999; Rubin 2008). Biodiesel production technologies based on lignocellulosic
substrate are economical as well as eco‐friendly and also address key environmental issues
(de Bhowmicka et al. 2018).
Recent Advances in the Production of Biodiesel Using Lignocellulosic Biomass 71
The aim of this chapter is to discuss possibilities for the use of various lignocellulosic
biomasses as feedstocks for the production of biodiesel. In addition, other important
aspects are covered including different types and composition of lignocellulosic biomass,
approaches to the pretreatment of lignocellulosic biomass, common fermentation methods
used for biodiesel production, etc.
5.2 Lignocellulosic Biomass as Novel Feedstock

for Biodiesel
A biological material derived from living organisms such as animals and plants is termed
biomass. Biomass, in general, contains various carbon‐based molecules composed of car-
bon, hydrogen, oxygen, and nitrogen with minor quantities of heavy metals and other ele-
ments. Lignocellulosic biomass is a natural and renewable substrate commonly used for
large‐scale production of biofuel, including biodiesel (Limayem and Sticke 2012). It
includes woods, agricultural residues, and industrial waste. It also contributes to environ-
mental sustainability and does not compete with food and animal feed production
(Demirbas 2003).
More specifically, lignocellulosic biomass is mainly composed of cellulose (30–50%),
hemicellulose (15–35%), lignin (10–20%), and some extractives (5–10%) (Sanchez 2009). It
is the most abundant natural material and represents potential volume for low‐cost biofuel
(Zhang et al. 2016; Sharma et al. 2019). It is produced as waste material from various
sources including agriculture, forest, domestic, and industrial waste (Cardona and Sanchez
2007) and poses a severe disposal problem. The depolymerization of lignocellulose is quite
difficult but microbial enzymes can act sequentially to achieve complete saccharification of
lignocellulosic biomass. Due to its immense biotechnological potential, lignocellulose bio-
mass processing is gaining huge attention as a cheap and renewable natural resource for
the production of value‐added products such as biodiesel.
As mentioned earlier, cellulose constitutes the greater part of lignocellulosic material. It
has been claimed that in the case of terrestrial ecosystems, cellulose is synthesized through
half of the fixed carbon per year. It is produced mainly by plants, but some algae, some
microorganisms, and a few animals also synthesize cellulose. Cellulose, generic formula
(C6H10O5)n, is one of the main components of the plant cell wall; it is composed of glu-
copyranosyl monomeric units linked by 1‐4‐β glycosidic bonds with a high degree of polym-
erization: 102–105. It is crystalline in nature and very fibrous with high rigidity due to
hydrogen links between cellulose molecules (Kucharska et al. 2018). A disaccharide named
cellobiose is the repeating unit of cellulose. Three‐dimensionally, cellulose consists of sheets
of glucopyranose rings arranged in a plane with sheets stacked on one another, forming a
highly organized structure. This structure is intermixed with unorganized amorphous
regions which possess suboptimal hydrogen bonding, which makes them accessible to water
molecules and hydrolyzing enzymes. Long chains of cellulose polymers with hydrogen and
van der Waals bond linkages are packed into microfibrils and covered by hemicelluloses and
lignin (Kumar et al. 2009). Lignocellulosic material with higher amounts of cellulose is
considered as a promising substrate for biofuel production (Raud et al. 2014).
Hemicellulose is the second most abundant polymer in the world, consisting of short
linear and branched polymers. It is a short, highly branched polymer with β‐1,4‐glucosidic
bonds as the main chains and β‐1,3; β‐1,6‐glucosidic bonds as side chains with a lower
degree of polymerization (200–300 units) compared to cellulose (10 000 units) (Brandt et al.
2013). It is an amorphous, complex heteropolymer comprising polyxylose, galactoglu-
comannan, and glucomannan polymers. Xylan and mannan are the main hemicellulosic
components of hardwood and softwood respectively. Acid hydrolysis of hemicelluloses
results in many products such as xylose, mannose, glucose, galactose, arabinose, and trace
amounts of rhamnose, glucuronic acid, methylglucuronic acid, and galacturonic acid
(Dworzanski et al. 2006; Kucharska et al. 2018). Hemicellulose is hydrolyzed quickly owing
to its amorphous and branched nature compared to cellulose.
Lignin is the main structural component of lignocellulose. It is one of the constituents of
the plant cell wall and plays an important role in protection of the plant from microbial
invasion (Hallenbeck 2012). Lignin is a heterogeneous, aromatic polymer whose main aro-
matic components are trans‐coniferyl, trans‐sinapyl, and trans‐p‐coumaryl alcohols
(Brandt et al. 2013; Saini et al. 2015). Basic subunits of lignin are p‐hydroxyphenylpropane,
syringylpropane, and guaiacylpropane, and the majority of phenylpropane units are linked
by ether bonds (mainly b‐O‐4) and the rest with carbon–carbon bonds (Kucharska et al.
2018). Lignin bonds the cellulose and hemicellulose fibers through different linkages
(Dworzanski et al. 2006). It is supposed to be the toughest part of the lignocellulose due to
its structural complexity. Cellulose and hemicellulose can be degraded by microorganisms
whereas lignin is resistant to microbial degradation (Birhanli and Yeşilada 2013) and chem-
ical degradation as well. Extractives represent a smaller fraction of wood component, up to
5% m/m. They include lipophilic and hydrophilic compounds such as terpenoids and ster-
oids, fats and waxes, phenolic constituents, and inorganic components (Kuhad et al. 2010;
Karimi and Taherzadeh 2016).
The large‐scale production of biofuels from lignocellulosic biomass is anticipated to ben-
efit society and the environment in various ways. Biomass feedstock for energy can be
derived from plant material like food crops, wood, grassy and woody plants, agriculture
and forestry residues, high oil‐containing algae, household waste (municipal), and indus-
trial waste. Fuel generated from biomass includes liquid fuels such as bioethanol and bio-
diesel. Anaerobic fermentation of lignocellulosic biomass results in production of
biohydrogen and biogas (Kucharska et al. 2018). The components of lignocellulosic bio-
mass serve as the potential energy source.
5.3 Methods for Pretreatment of Lignocellulosic Biomass
As discussed earlier, lignocellulosic biomass is composed of three important polymers: cel-

lulose, hemicellulose, and lignin. Out of these, carbohydrate polymers (i.e., cellulose and
hemicellulose) account for about 75% and are the important source of fermentable sugars
which are further used for the production of liquid fuels. However, the release of such sug-
ars from biomass is a major challenge for lignocellulose biorefining technologies due to the
compact and rigid structure of the biomass (Jørgensen et al. 2007). The complex structure
of these polymers and their spatial interlinks make the biomass resistant to processing.
This characteristic creates a physical barrier to protect the carbohydrate polymers from
degradation by microorganisms or enzymes (Zhao et al. 2012). Therefore, pretreatment of
lignocellulosic biomass helps to degrade its complex structure and release the fermentable
sugars which can be further converted to biodiesel through different fermentation
approaches (Maurya et al. 2015).
In recent decades several pretreatment approaches have been routinely used for different
types of biomass. The main objective of each pretreatment approach is to break the physical
barriers of the biomass and make the carbohydrate polymers available for enzymatic digest-
ibility by altering the chemical composition and physical structure of the biomass feedstock.
Therefore, knowledge of the chemical composition and physical structure of biomass and
how it affects enzymatic hydrolysis plays a key role in the improvement of existing pretreat-
ment technologies and also helps to develop novel pretreatment processes.
Different conventional pretreatment approaches like physical, chemical, physicochemi-
cal, biological, and biochemical methods are commonly used for a variety of lignocellulosic
feedstocks. Among these, important physical pretreatments include mechanical extrusion
(Lamsal et al. 2010; Zheng and Rehmann 2014), milling and grinding (Kumar et al. 2009;
Kim et al. 2018), microwave (Hassan et al. 2018), ultrasonication (Bussemaker and Zhang
2013), pyrolysis (Oudenhoven et al. 2016), and application of pulsed electric field (Kumar
and Sharma 2017). Chemical pretreatment approaches mainly include use of various
chemical agents like dilute acids (Liu et al. 2018), alkali (Kim et al. 2018), ozone (Fang et al.
2018), etc. In addition, other chemical methods are used like organosolv (Ichwan and Son
2011), ionic liquids (Behera et al. 2014), deep eutectic solvents (Zhang et al. 2012), and
natural deep eutectic solvents (Dai et al. 2013). Physicochemical pretreatment methods like
steam explosion (Pielhop et al. 2016), liquid hot water (Yu et al. 2010), ammonia fiber
explosion (Teymouri et al. 2004), ammonia recycle percolation (Kim and Lee 2005), and
supercritical fluid pretreatment (Gu et al. 2013), etc. are used.
Apart from these, biological pretreatments are considered as the most effective methods
due to their eco‐friendly and economically viable nature. They generally involve the use of
different white‐ and brown‐rot fungi and a variety of bacteria (Akhtar et al. 2016). The
enzymes produced by these microorganisms are responsible for the degradation of carbo-
hydrate polymers present in lignocellulosic biomass. For some lignocellulosic biomass, a
combination of two or more different approaches may be most effective. Bioorganosolv and
bioozonolysis are combinational approaches used for the pretreatment of some biomass
(Ingle et al. 2019). Table 5.1 summarizes the different pretreatment methods.
5.4 Fermentation Approaches Used in Biodiesel Production
Different fermentation processes are commonly used for the biotechnological production
of single cell oil from lignocellulosic feedstocks; this mainly involves submerged/liquid
fermentation (Zhu et al. 2008; Huang et al. 2009) and solid‐state fermentation (Peng and
Chen 2008; Fakas et al. 2009). In the submerged fermentation process, the substrate used
for fermentation is always in liquid state which contains the nutrients needed for growth.
Therefore, when used for biomass, it requires prior sugar extraction from the biomass
feedstock to the bulk liquid which makes the process expensive and time‐consuming.
Table 5.1 Important methods used for the pretreatment of lignocellulosic biomass (LB).
Pretreatment
methods Brief description of process
Physical pretreatments
Mechanical Heating of biomass at high temperature (>300 °C) under shear mixing
extrusion
Milling Chipping and grinding of biomass
Microwave Degradation of biomass through microwave irradiation
Ultrasound Degradation of biomass with the application of ultrasound/ultrasonic
waves
Pyrolysis Biomass is subjected to high‐temperature treatment (500–800 °C) in the
absence of oxidizing agent
Pulsed electric field Biomass is subjected to a sudden burst of high voltage (5.0–20.0 kV/cm)
for short durations
Chemical pretreatments
Acids Use of dilute acids like sulfuric acid, oxalic acid, and maleic acid for
biomass pretreatment
Alkali Use of different alkalis such as sodium hydroxide, potassium hydroxide,
calcium hydroxide, and ammonium hydroxide, for biomass pretreatment
Ozonolysis Degradation of biomass using ozone treatment
Organosolv Application of aqueous organic solvents such as ethanol, methanol,
ethylene glycol, acetone, etc.
Ionic liquids Use of various derivatives of imidazolium salts
Deep eutectic Biomass exposure with solvents made up of two or three components
solvents capable of self‐association through hydrogen bond interactions
Natural deep Application of choline, urea, sugars, amino acids, some organic acids,
eutectic solvents etc.
Physicochemical pretreatments
Steam explosion LB is subjected to high pressure and temperature for a short residence
time followed by rapid depressurization enabling fiber explosion
Liquid hot water LB is treated with water at high temperatures (160–240 °C) and high
pressure
Ammonia fiber LB is subjected to liquid hydrous ammonia under high pressures and
explosion moderate temperatures (60–100 °C) followed by rapid depressurization
Ammonia recycle Ammonium hydroxide (5–15% wt), at high temperature (140–210 °C)
percolation compared to ammonia fiber expansion (AFEX) where the moderate
temperature is about 60–100 °C
Supercritical fluid Use of supercritical fluid materials in liquid or gaseous form
pretreatment
Biological pretreatments
White‐rot fungi Punctularia spp., Irpex lacteus, Ceriporiopsis subvermispora, Phlebia
brevispora, P. floridensis, P. radiata, Echinodontium taxodii, Gonoderma
spp., Oxysporus spp., Trametes versicolor, Pleurotus sajor‐caju and
Trichoderma reesei
Table 5.1 (Continued)
Pretreatment
methods Brief description of process
Brown‐rot fungi Serpula lacrymans, Coniophora puteana, Meruli poria incrassata,

Laetoporus sulphureus and Gleophyllum trabeum
Bacteria Caldicellulosiruptor bescii, Clostridium thermocellum, Clostridium
clariflavum and C. cellulolyticum, Cupriavidus basilensis
Biochemical pretreatments
Bioorganosolv, Combination of biological and chemical methods
bioozonolysis
Source: Adapted from Ingle et al. (2019) with permission from John Wiley & Sons Ltd.
Table 5.2 The different lignocellulosic feedstocks and their fermentation process to accumulate
lipid by oleaginous microorganism.
Lignocellulosic feedstocks Fermentation process involved
Wheat straw and wheat bran Solid‐state

Pitch pine Submerged/liquid
Pear pomace Solid‐state
Rice straw Submerged/liquid
Wheat straw Submerged/liquid
Sweet sorghum Semi‐solid state
Tomato waste Submerged/liquid
Corn stalk Submerged/liquid
Tree (Populus euramevicana) leaves Submerged/liquid
Rice straw Submerged/liquid
Source: Adapted and modified from Yousuf (2012) with permission from Elsevier.
However, solid‐state fermentation is considered most suitable for biomass like bran,
bagasse, and paper pulp. In this process, the desired microorganisms are generally grown
on moist, solid materials in the absence of free‐flowing water, so that limited lipid accumu-
lation can be obtained (Yousuf 2012).
Compared with submerged fermentation, solid‐state fermentation has many advantages
such as simple processing, a bioreactor with small capacity is sufficient, simple down-
stream processing, less energy required, low wastewater output and, most importantly,
economic viability (Pérez‐Guerra et al. 2003; Yousuf 2012). Economou et al. (2010) demon-
strated that a small modification of solid‐state fermentation, making it into a semi‐solid
fermentation system by increasing the water content, encouraged easy growth of the
desired fungus and also enhanced the production of single cell oil. According to Pandey
et al. (2000), solid‐state fermentation can use various natural resources like agricultural
waste, wood residues, energy crops, and by‐products of the food industry. Table 5.2 lists the
lignocellulosic biomass and suitable fermentation processes used to accumulate lipid by

oleaginous microorganisms.
5.5 Production of Biodiesel from Lignocellulosic Biomass
Biodiesel is generated primarily based on transesterification of plant oils. It is one of the

liquid biofuels that includes pure plant oil and bioethanol. In global production of biofuels,
ethanol, primarily obtained from starchy crops, maize, and sugar, is at the top followed by
biodiesel production. Biodiesel can be used in engines in pure form or blended with con-
ventional diesel fuels. Biodiesel is used in generator engines as well as vehicle engines.
Numerous processes exist to convert different feedstocks into biofuels.
Biofuels produced through familiar and well‐established processes such as cold pressing,
chemical synthesis, hydrolysis and fermentation, and transesterification are referred to as
first‐generation biofuels. Second‐generation biofuels are yet to reach commercialization on
a large scale as their conversion technologies are still in the nascent stage. Hydro treat-
ment, advanced hydrolysis and fermentation, and gasification and synthesis are some of
the advanced processes that are being explored for second‐generation biofuel production.
A broad range of feedstocks can be used in the production of next‐generation biofuels,
including lignocellulosic sources (Kimble et al. 2008). Lignocellulosic feedstock materials
include crop straw, bagasse, forest waste, switchgrass, Miscanthus, Indian grass, and short‐
rotation forests. These second‐generation biofuels can address concerns associated with
first‐generation biofuels and potentially bring about significant cost reductions in the
longer term.
Kim et al. (2015) have explored nitric acid‐pretreated co‐production of bioethanol and
biodiesel from corn stover with promising outcomes. Saccharomyces cerevisiae produced an
ethanol concentration of 22.4 g/L, equivalent to 69.1% theoretical ethanol yield related to
initial cellulose weight, while Cryptococcus curvatus in hydrolyzate medium produced
1.04 g/L lipid yield, which was higher with reference to a defined medium having pure
xylose (Kim et al. 2015).
Li et al. (2015) reported biogas and biodiesel production from corncob in a multistep
bioprocess. Initially, biogas production from corncob by anaerobic fermentation was fol-
lowed by biogas residue conversion by black soldier fly larvae; finally biodiesel was pro-
duced from larvae grease. 86.70 L biogas was obtained through anaerobic fermentation
from 400 g corncob which translates to a biogas yield of 220.71 mL/g, and 3.17 g of biodiesel
was produced from the larvae grease (Li et al. 2015).
In order to meet our ever increasing energy needs, converting microbial lipids to bio-
diesel fuel is one of the many approaches in shifting toward a future less dependent on
limited fossil fuel reserves. Conversion of vegetable oil to biodiesel is costly and is an eco-
nomically nonviable proposition. In search of a way to circumvent the cost barrier, research-
ers have turned their attention to microbial lipids as a competitive alternative source for
biodiesel production by growing the microbes on readily available and cheap lignocellu-
losic waste materials (Galafassi et al. 2012). Inhibitors derived from biomass pretreatment
and biofuel fermentation limit the efficiency of microbial strains (Wang et al. 2018). As a
promising microbial lipid producer, the oleaginous yeast Rhodotorula graminis is capable
of lipid production from a broad range of carbon sources with a complementary ability to
tolerate inhibitors normally generated in the course of preprocessing of lignocellulosic
materials, especially in the hydrolysis step. With the yeast R. graminis, using undetoxified
corn stover hydrolysate as substrate, Galafassi et al. (2012) have shown a lipid productivity
of 0.21 gL/h and a total lipid content of 34% w/w. Their corresponding results with crude
glycerol as the carbon source were 0.15 gL/h and 54% w/w, respectively. Work with
R. graminis has thus revealed a suitable candidate for fermentation processes involving
renewable resources.
In another interesting study investigating biodiesel production from cheap, nonedible
and abundant lignocellulosic biomass, a novel oleaginous yeast Rhodosporidium kratoch-
vilovae HIMPA1 was identified, having the ability to grow and accumulate high amounts of
triacylglycerides as large intracellular lipid droplets of 4.35 μm size, total lipids
(4.86 ± 0.54 g/L) with lipid content of 53.18% (w/w) from aqueous extract of Cassia fistula
L. fruit pulp as the sole nutritional source (Patel et al. 2015). The lipid profile revealed pal-
mitic acid (C16:0) 43.06%, stearic acid (C18:0) 28.74%, and oleic acid (C18:1) 17.34% as
major fatty acids. High saturated fatty acids content (72.58%) can be blended with high
polyunsaturated fatty acids (PUFA) feedstocks to make an industrially viable renewable
energy product.
Slininger (2016) reported four robust oleaginous yeast strains able to produce high lipid
concentrations from both acid‐ and base‐pretreated biomass. The screening involved a two‐
tier arrangement involving a primary screening medium of undetoxified enzyme hydrolyz-
ates of ammonia fiber expansion (AFEX)‐pretreated corn stover and a secondary screening
medium applied to strains passing the primary screen obtained from acid‐pretreated switch-
grass. Three of these reported isolates are novel bioconverters of lignocellulose to lipids.
Among the various current approaches employed to efficiently convert lignocellulose to
biofuel, the potential of mass spectrometry‐based metabolomic analysis of biofuel‐
producing microbes is being explored. Identifying a robust next‐generation enzyme is key
for a cost‐effective biofuel production industry. Metabolic engineering focused on inhibitor
tolerance and evolutionary engineering have been able to obtain microbes with the desired
tolerance (Wang et al. 2018). Information acquired from metabolomic analysis has been
utilized to improve the stress tolerance of biofuel producers, produce unique bioproducts,
characterize the metabolism of emerging biofuel producers, and enable efficient utilization
of lignocellulosic biomass and productivity of engineered pathways (Martien and Amador‐
Noguez 2017).
One of the major classes of inhibitory compounds derived from lignocellulosic biomass
pretreatment has been identified as aldehydes that interfere with microbial growth and
subsequent fermentation for advanced biofuel production (Wang et al. 2017a). Wang et al.
(2017a) demonstrated and identified five different uncharacterized putative aryl‐alcohol
dehydrogenase genes (AADs) from Scheffersomyces stipitis (Pichia) as a new source of
resistance against biomass fermentation inhibitor, 2‐furaldehyde (furfural) by gene expres-
sion, gene cloning, and direct enzyme assay analysis using partially purified proteins.
Analysis revealed that genes SsAAD2, SsAAD3, and SsAAD4 are inducible and displayed
significantly enhanced levels of expression in response to the challenge of furfural. Their
encoding proteins also showed higher levels of specific activity toward furfural and were
suggested as core functional enzymes contributing to aldehyde resistance in S. stipitis.
When trying to optimize lignocellulosic feedstock bioconversion to biofuel, one of the

important aspects is to look at the availability of xylose which is the second most abundant
monosaccharide. Thus many studies have developed efficient xylose‐utilizing microorgan-
isms to generate biofuel. S. cerevisiae has been thoroughly engineered to assimilate xylose
due to its robustness under industrial fermentation conditions (Kwak and Jin 2017). Cetyl
alcohol and palmityl alcohol have been used as an emulsifier and a lubricant in various
industrial fields and are considered as a potential advanced biofuel.
An approach that can completely avoid generation of toxic inhibitors that interfere with
efficient microbial bioconversion process was demonstrated by Agematu et al. (2017). Their
work involved a rumen‐mimicking bioprocess on lignocellulosic biomass which was dry‐
pulverized and continuously cultivated with ruminal bacteria in a controlled pH using
ammonium as a nitrogen source. The study was carried out by continuously cultivating rumi-
nal bacteria for over 60 days that digested microcrystalline cellulosic materials including rice
straw and Japanese cedar to produce volatile fatty acids (VFAs) (Agematu et al. 2017).
Avoiding the toxic inhibitors that interfere with efficient microbial bioconversion process
can be an alternative approach. One such approach is the use of insects as a promising
resource for biodiesel production (Wang et al. 2017b). Wang et al. (2017b) have successfully
demonstrated the potential of certain kinds of insects, many of which are voracious eaters
of organic wastes, as a promising green alternative to meet the growing demands of the
global population for food, feed, and energy. These insects have been highlighted as source
of protein that may help address environmental, economic, and health issues. The study
demonstrated that successive co‐conversion of corn stover by insects possessing different
feeding habits could be an attractive option for tackling multiple environmental issues
simultaneously by efficient utilization of lignocellulosic resources, and represents a poten-
tially valuable solution to farm waste management, the rise of global energy needs, and
demand for animal feed. The study by Wang et al. (2017b) included use of two different
insects in succession: initially yellow mealworm (Tenebrio molitor L.) followed by black sol-
dier fly (Hermetia illucens L.) for corn stover degradation; the residues produced during the
first stage were consumed by the insect in the second stage. The use of insects in succession
resulted in a waste dry mass reduction rate of 51.32%, yielding 1.95 g crude grease from lar-
val biomass translating to 1.76 g biodiesel and by‐products such as 6.55 g protein and 111.59 g
biofertilizer. The two insect‐based biorefinery yielded a total insect biomass of 8.50 g. The
efficiency of conversion of crude grease free fatty acids into biodiesel touched 90%.
All the latest approaches mentioned above indicate that the major hindrance in efficient
utilization of lignocellulosic waste material for biodiesel production is the toxic metabolites
produced during preprocessing. Efficient and effective utilization of metabolic genes iden-
tified through metabolomics capable of providing tolerance to stress and toxins will even-
tually lead us to success in harnessing the full potential of lignocellulosic feedstock
materials for biodiesel production.
5.6 Challenges in the Production of Biodiesel
The implementation of any new technology greatly depends upon its inclusiveness of the
“whole system.” The technology can be considered successful if it has a positive impact on
the stakeholders and society. Therefore, there is always a need to pinpoint and meet the
challenges which can make a technology successful. Biodiesel production has also socio-
economic challenges which need to be considered before it can achieve worldwide
implementation.
5.6.1 Social Challenges

Development of any technology should consider its impact on social issues that may be
linked with it. Interestingly, this challenge associated with biofuel production is still not
given enough emphasis. The social acceptance of the technology hugely affects its sustain-
ability and success.
For cultivation of a biofuel crop, a huge amount of land is required. Hence, if land previ-
ously used for food crops is turned over to biofuel crops, this will affect the food supply.
Moreover, it will also increase the food price, which mostly affects poor people, especially
those from developing countries (OECD/FAO 2007). Thus inflation in food price will make
biofuel and food more costly (Searchinger 2011). The land may be made available by defor-
estation or acquiring agricultural land, which will affect the livelihood of the huge popula-
tion across the globe which depends upon forest and agricultural land. Hence, sustainable
development of this technology without affecting these populations will be a great
challenge.
5.6.2 Environmental Challenges

It has been claimed that biofuels are more environmentally friendly than fossil fuels, pro-
ducing reduced amounts of toxic gases (den Haan et al. 2013). However, as for other fuels,
the production and consumption of biofuel will add to the environmental burden in the
form of toxic gases generated, especially after consumption. This will lead to environmen-
tal pollution via emission of greenhouse gases. Biofuel technology will also have an impact
on essential natural resources such as food, water, and land. Therefore, there is a need to
draw attention to climate change which might occur due to biofuel consumption. There is
also a requirement to set criteria for the assessment of the effect of lignocellulosic biofuel
on rural populations.
After setting up a biorefinery, it is necessary to perform a life cycle analysis (LCA) of
the steps involved in biofuel production. Air pollution is the major factor to be consid-
ered as this is increased during the harvesting, grinding, and chemical pretreatment pro-
cesses. The LCA should be taken seriously by biorefineries for assessment of the
environmental impact of this technology. Such analysis will help biorefineries to modu-
late the production process of biofuel. However, the LCA will add to the production costs
of biofuel. Hence, it will be a great challenge for the industry to produce biofuel at an
affordable price.
Another important environmental impact is the effect of biofuel production on soil ero-
sion. The plants used for biofuel can help in preventing soil erosion due to afforestation and
also preventing water run‐off and loss of sediment (Neumann et al. 2010). In this context,
it has been reported that planting jatropha and other biofuel crops helps in improving the
quality of soil and can even create a soil environment good enough for other food crops
(Achten et al. 2008; Chum et al. 2011).
5.7 Future Prospects
Many big industries are demonstrating interest in biofuel. However, before investing their
capital, there is a need to focus on some aspects related to its production. The concerned
research and development team should preferably try to run a pilot‐scale project of sustain-
able biofuel production. The challenges of the separation and purification processes need
to be addressed so that they can generate highly pure biofuel without or with only a
minimum quantity of other unwanted residues. However, significant focus is required on
deciding the type of pretreatment method, lignocellulosic biomass, and catalyst for effi-
cient biofuel production. Additionally, we need more in‐depth analysis of the effects of this
technology at the societal and environmental levels, so that biofuel can become a great
alternative to fossil fuel at large scale.
5.8 Conclusion
As the natural resources of fuel are diminishing, there is a pressing need to look for a truly
viable alternative, a technology which can produce fuel sustainably and at an affordable
price. In this context, biodiesel technology has the potential to meet the global energy
demand. It has been found that biofuel, especially made from lignocellulosic biomass, has
the edge over other options. It can be obtained via various routes, purified and used like
conventional diesel.
This technology looks promising but negative factors associated with its use need to be
taken into consideration. The first and foremost is the social and environmental challenges,
including the exploitation of productive land for biomass substrate and high production of
greenhouse gases. Hence, before adopting this technology for worldwide application, there
is a need to consider all aspects of boons and banes associated with it. Therefore, there is
huge responsibility on the shoulders of concerned scientists to develop it so that it can be a
greatly needed resource for humankind.
References
Achten, W., Verchot, L., Franken, Y. et al. (2008). Jatropha bio‐diesel production and use.
Biomass and Bioenergy 32: 1063–1084.
Adewale, P., Dumont, M.J., and Ngadi, M. (2015). Recent trends of biodiesel production from
animal fat wastes and associated production techniques. Renewable and Sustainable Energy
Reviews 45: 574–588.
Agematu, H., Takahashi, T., and Hamano, Y. (2017). Continuous volatile fatty acid production
from lignocellulosic biomass by a novel rumen‐mimetic bioprocess. Journal of Bioscience
and Bioengineering 124 (5): 528–533.
Ahorsu, R., Medina, F., and Constantí, M. (2018). Significance and challenges of biomass as a
suitable feedstock for bioenergy and biochemical production: a review. Energies 11: 3366.
Akhtar, N., Gupta, K., Goyal, D., and Goyal, A. (2016). Recent advances in pretreatment
technologies for efficient hydrolysis of lignocellulosic biomass. Environmental Progress &
Sustainable Energy 35 (2): 489–511.
Arenas, E.G., Rodriguez Palacio, M.C., Juantorena, A.U. et al. (2016). Microalgae as a potential
source for biodiesel production: techniques, methods and other challenges. International
Journal of Energy Research 41 (6): 761–789.
Atabani, A.E., Silitonga, A.S., Ong, H.C. et al. (2013). Non‐edible vegetable oils: a critical
evaluation of oil extraction, fatty acid compositions,biodiesel production, characteristics,
engine performance and emissions production. Renewable and Sustainable Energy Reviews
18: 211–245.
Behera, S., Arora, R., Nandhagopal, N., and Kumar, S. (2014). Importance of chemical
pretreatment for bioconversion of lignocellulosic biomass. Renewable and Sustainable
Energy Reviews 36: 91–106.
Birhanli, E. and Yeşilada, Ö. (2013). The utilization of lignocellulosic wastes for laccase
production under semisolid‐state and submerged fermentation conditions. Turkish Journal
of Biology 37: 450–456.
Brandt, A., Gräsvik, J., Halletta, J.P., and Welton, T. (2013). Deconstruction of lignocellulosic
biomass with ionic liquids. Green Chemistry 15: 550–583.
Bussemaker, M.J. and Zhang, D. (2013). Effect of ultrasound on lignocellulosic biomass as a
pretreatment for biorefinery and bio‐fuel applications. Industrial and Engineering Chemistry
52: 3563–3580.
Cardona, C.A. and Sanchez, O.J. (2007). Fuel ethanol production: process design trends and
integration opportunities. Bioresource Technology 98: 2415–2457.
Chisti, Y. (2010). Fuels from microalgae. Biofuels 1: 233–235.
Chum, H., Faaij, A., Moreira, J. et al. (2011). Bioenergy. In: IPCC Special Report on Renewable
Energy Sources and Climate Change Mitigation (eds. O. Edenhofer, R. Pichs‐Madruga, Y.
Sokona, et al.), 209–332. Cambridge: Cambridge University Press.
da Costa Cardoso, L., de Almeida, F.N.C., Keiff, G. et al. (2018). Synthesis and optimization of
ethyl esters from fish oil waste for biodiesel production. Renewable Energy 133: 743–748.
Dai, Y., van Spronsen, J., Witkamp, G.J. et al. (2013). Natural deep eutectic solvents as new
potential media for green technology. Analytica Chimica Acta 766: 61–68.
De Bhowmicka, G., Sarmaha, A.K., and Sen, R. (2018). Lignocellulosic biorefinery as a model
for sustainable development of biofuels and value added products. Bioresource Technology
247: 1144–1154.
Demirbas, A. (2003). Biodiesel fuels from vegetable oils via catalytic and non‐catalytic
supercritical alcohol transesterifications and other methods: a survey. Energy Conversion
and Management 44: 2093–2109.
Dworzanski, J.P., Buchanan, R.M., Chapman, J.N., and Meuzelaar, H.L.C. (2006).
Characterization of lignocellulosic materials and model compounds by combined tg/(Gc)/Ft
Ir/Ms, Symp. Pyrolysis. Natural and Synthetic Macromolecules 36: 725–732.
Economou, C.N., Makri, A., Aggelis, G. et al. (2010). Semi‐solid state fermentation of sweet
sorghum for the biotechnological production of single cell oil. Bioresource Technology 101:
1385–1388.
Fakas, S., Makri, A., Mavromati, M. et al. (2009). Fatty acid composition in lipid fractions
lengthwise the mycelium of Mortierella isabellina and lipid production by solid state
fermentation. Bioresource Technology 100: 6118–6120.
Fang, S., Wang, W., Tong, S. et al. (2018). Evaluation of the effects of isolated lignin on
cellulose enzymatic hydrolysis of corn stover pretreatment by NaOH combined with ozone.
Molecules 23 (6): 1495.
Galafassi, S., Cucchetti, D., Pizza, F. et al. (2012). Lipid production for second generation
biodiesel by the oleaginous yeast Rhodotorula graminis. Bioresource Technology 111:
398–403.
Gu, T., Held, M.A., and Faik, A. (2013). Supercritical CO2 and ionic liquids for the
pretreatment of lignocellulosic biomass in bioethanol production. Environmental Science
and Technology 34 (13–16): 1735–1749.
den Haan, H., Kroukamp, H., Mert, M. et al. (2013). Engineering Saccharomyces cerevisiae for
next generation ethanol production. Journal of Chemical Technology & Biotechnology 88 (6):
983–991.
Hallenbeck, P.C. (2012). Microbial Technologies in Advanced Biofuels Production. Dordrecht:
Springer.
Hassan, S.S., Williams, G.A., and Jaiswal, A.K. (2018). Emerging technologies for pretreatment
of lignocellulosic biomass. Bioresource Technology 262: 310–318.
Huang, C., Zong, M., Wu, H., and Liu, Q. (2009). Microbial oil production from rice straw
hydrolysate by Trichosporon fermentans. Bioresource Technology 100: 4535–4538.
Ichwan, M. and Son, T.W. (2011). Study on organosolv pulping methods of oil palm biomass.
Proceedings of the 2nd International Seminar on Chemistry, Jatinangor, Indonesia, pp.
364–370.
Ingle, A.P., Chandel, A.K., Antunes, F.A.F. et al. (2019). New trends in application of
nanotechnology for the pretreatment of lignocellulosic biomass. Biofuels, Bioproducts &
Biorefining 13 (3): 776–788.
International Energy Agency (IEA) (2018). Renewables Information 2018: Overview. Paris,
France: IEA.
Jørgensen, H., Kristensen, J.B., and Felby, C. (2007). Enzymatic conversion of lignocellulose
into fermentable sugars: challenges and opportunities. Biofuels, Bioproducts & Biorefining 1:
119–134.
Karimi, K. and Taherzadeh, M.J. (2016). A critical review of analytical methods in
pretreatment of lignocelluloses: composition, imaging, and crystallinity. Bioresource
Technology 200: 1008–1018.
Katabathini, N., Lee, A., and Wilson, K. (2007). Catalysts in production of biodiesel: a review.
Journal of Biobased Materials and Bioenergy 1: 19–30.
Kim, I., Seo, Y.H., Kim, G.‐Y., and Han, J.‐I. (2015). Co‐production of bioethanol and biodiesel
from corn stover pretreated with nitric acid. Fuel 143: 285–289.
Kim, S. (2018). Enhancing bioethanol productivity using alkali‐pretreated empty palm fruit
bunch fiber hydrolysate. BioMed Research International 2018: 5272935.
Kim, S.J., Um, B.H., Im, D.J. et al. (2018). Combined ball milling and ethanol organosolv
pretreatment to improve the enzymatic digestibility of three types of herbaceous biomass.
Energies 11: 2457.
Kim, T.H. and Lee, Y.Y. (2005). Pretreatment and fractionation of corn stover by ammonia
recycle percolation process. Bioresource Technology 96 (18): 2007–2013.
Kimble, M., Pasdeloup, M.V., and Spencer, C. (2008). Sustainable Bioenergy Development in
UEMOA Member Countries, 45–56. http://large.stanford.edu/courses/2011/ph240/demori2/
docs/2008_10_unf_bioenergy_full_report1.pdf.
Kirubakaran, M. and Selvan, V.A.M. (2018). A comprehensive review of low cost biodiesel
production from waste chicken fat. Renewable and Sustainable Energy Reviews 2: 390–401.
Kucharska, K., Hołowacz, I., Konopacka‐Łyskawa, D. et al. (2018). Key issues in modeling and
optimization of lignocellulosic biomass fermentative conversion to gaseous biofuels.
Renewable Energy 129: 384–408.
Kudre, T.G., Bhaskar, N., and Sakhare, P.Z. (2017). Optimization and characterization of
biodiesel production from rohu (Labeo rohita) processing waste. Renewable Energy 113:
1408–1418.
Kuhad, R.C., Gupta, R., Khasa, Y.P., and Singh, A. (2010). Bioethanol production from Lantana
camara (red sage): pretreatment, saccharification and fermentation. Bioresource Technology
101: 8348–8354.
Kumar, A.K. and Sharma, S. (2017). Recent updates on different methods of pretreatment of
lignocellulosic feedstocks: a review. Bioresource Bioprocess 4: 7.
Kumar, P., Barrett, D.M., Delwiche, M.J., and Stroeve, P. (2009). Methods for pretreatment of
lignocellulosic biomass for efficient hydrolysis and biofuel production. Industrial Chemical
Engineering Research 48: 3713–3729.
Kwak, S. and Jin, Y.S. (2017). Production of fuels and chemicals from xylose by engineered
Saccharomyces cerevisiae: a review and perspective. Microbial Cell Factories 16: 82.
Lamsal, B., Yoo, J., Brijwani, K., and Alavi, S. (2010). Extrusion as a thermo‐mechanical pre
treatment for lignocellulosic ethanol. Biomass and Bioenergy 34 (12): 1703–1710.
Lapuerta, M., Armas, O., and Fernandez, J.R. (2008). Effect of biodiesel fuels on diesel engine
emissions. Progress in Energy and Combustion Science 34: 198–224.
Li, Q., Du, W., and Liu, D. (2008). Perspectives of microbial oils for biodiesel production.
Applied Microbiology and Biotechnology 80 (5): 749–756.
Li, W., Li, Q., Zheng, L.Y. et al. (2015). Potential biodiesel and biogas production from corncob
by anaerobic fermentation and black soldier fly. Bioresource Technology 194: 276–282.
Liandong, Z., Nugroho, Y.K., Shakeel, S.R. et al. (2017). Using microalgae to produce liquid
transportation biodiesel: what is next? Renewable and Sustainable Energy Reviews 78:
391–400.
Limayem, A. and Sticke, S.C. (2012). Lignocellulosic biomass for bioethanol production:
current perspectives, potential issues and future prospects. Progress in Energy and
Combustion Science 38 (4): 449–467.
Liu, W., Chen, W., Hou, Q. et al. (2018). Effects of combined pretreatment of dilute acid pre‐
extraction and chemical‐assisted mechanical refining on enzymatic hydrolysis of
lignocellulosic biomass. RSC Advances 8: 10207–10214.
Martien, J.I. and Amador‐Noguez, D. (2017). Recent applications of metabolomics to advance
microbial biofuel production. Current Opinion in Biotechnology 43: 118–126.
Maurya, D.P., Singla, A., and Negi, S. (2015). An overview of key pretreat‐ment processes for
biological conversion of lignocellulosic biomass to bioethanol. 3 Biotech 5 (5): 597–609.
Meng, X., Yang, J., Xu, X. et al. (2009). Biodiesel production from oleaginous microorganisms.
Mohammed, N.I., Kabbashi, N.A., Alade, A.O., and Sulaiman, S. (2018). Advancement in the
utilization of biomass‐derived heterogeneous catalysts in biodiesel production. Green and
Sustainable Chemistry 8: 74–91.
Musa, S.D., Tang, Z., Ibrahim, A.O., and Mukthar, H. (2018). China’s energy status: a critical
look at fossils and renewable options. Renewable and Sustainable Energy Reviews 81 (2):
2281–2290.
Nasreen, S., Nafees, M., Qureshi, L. et al. (2018). Review of catalytic transesterification
methods for biodiesel production. IntechOpen, UK. doi: 10.5772/intechopen.75534.
Neumann, K., Verburg, P.H., Stehfest, E., and Muller, C. (2010). The yield gap of global grain
production: a spatial analysis. Agricultural Systems 103: 316–326.
OECD/FAO (2007). OECD‐FAO Agricultural Outlook 2007–2016. Paris: OECD Publications.
Oudenhoven, S.R.G., van der Ham, A.G.J., van den Berg, H. et al. (2016). Using pyrolytic acid
leaching as a pretreatment step in a biomass fast pyrolysis plant: process design and
economic evaluation. Biomass and Bioenergy 95: 388–404.
Pandey, A., Soccol, C.R., Nigam, P., and Soccol, V.T. (2000). Biotechnological potential of agro
industrial residues. I: sugarcane bagasse. Bioresource Technology 74 (1): 69–80.
Patel, A., Sindhu, D.K., Arora, N. et al. (2015). Biodiesel production from non‐edible
lignocellulosic biomass of Cassia fistula L. fruit pulp using oleaginous yeast Rhodosporidium
kratochvilovae HIMPA1. Bioresource Technology 197: 91–98.
Peng, X. and Chen, H. (2008). Single cell oil production in solid‐state fermentation by
Microsphaeropsis sp. from steam‐exploded wheat straw mixedwith wheat bran. Bioresource
Pérez‐Guerra, N., Torrado‐Agrasar, A., López‐Macias, C., and Pastrana, L. (2003). Main
characteristics and applications of solid substrate fermentation. Electronic Journal of
Agricultural and Food Chemistry 2 (3): 343–350.
Pielhop, T., Amgarten, J., von Rohr, P.R., and Studer, M.H. (2016). Steam explosion
pretreatment of softwood: the effect of the explosive decompression on enzymatic
digestibility. Biotechnology for Biofuels 9: 152.
Pinto, A.C., Guarieiro, L.L.N., Rezende, M.J.C. et al. (2005). Biodiesel: an overview. Journal of
the Brazilian Chemical Society 16: 1313–1330.
Ramalingam, S., Rajendran, S., Ganeshan, P., and Govindaswamy, M. (2018). Effect of
operating parameters and antioxidant additives with biodiesels to improve the performance
and reducing the emissions in a compression ignition engine‐A review. Renewable and
Raud, M., Kesperi, R., Oja, T. et al. (2014). Utilization of urban waste in bioethanol production:
potential and technical solutions. Agronomy Research 12 (2): 397–406.
Rubin, E.M. (2008). Genomics of cellulosic biofuels. Nature 454: 841–845.
3 Biotech. 5 (4): 337–353.
Sanchez, C. (2009). Lignocellulosic residues:biodegradation and bioconversion by fungi.
Biotechnology Advances 27: 185–194.
Searchinger, T. (2011). How biofuels contribute to the food crisis. Washington Post, Washington
DC.
Sharma, H.K., Xu, C., and Qin, W. (2019). Biological pretreatment of lignocellulosic biomass
for biofuels and bioproducts: an overview. Waste and Biomass Valorization 10: 235–251.
Sharma, Y.C. and Singh, V. (2017). Microalgal biodiesel: a possible solution for India’s energy
security. Renewable and Sustainable Energy Reviews 67: 72–78.
Slininger, P.J. (2016). Comparative lipid production by oleaginous yeasts in hydrolyzates of
lignocellulosic biomass and process strategy for high titers. Biotechnology and Bioengineering
113 (1): 2305–2305.
Teymouri, F., Laureano‐Pérez, L., Alizadeh, H., and Dale, B.E. (2004). Ammonia fiber
explosion treatment of corn stover. Applied Biochemistry and Biotechnology: Part A 115
(1–3): 951–963.
Wang, H., Rehman, K.U., Liu, X. et al. (2017a). Insect biorefinery: a green approach for
conversion of crop residues into biodiesel and protein. Biotechnology for Biofuels 10: 304.
Wang, S., Sun, X., and Yuan, Q. (2018). Strategies for enhancing microbial tolerance to
inhibitors for biofuel production: a review. Bioresource Technology 258: 302–309.
Wang, X., Lewis Liu, Z., Zhang, X., and Ma, M. (2017b). A new source of resistance to 2‐
furaldehyde from Scheffersomyces (Pichia) stipitis for sustainable lignocellulose‐to‐biofuel
conversion. Applied Biochemistry and Biotechnology 101 (12): 4981–4993.
Wyman, C.E. (1999). Biomass ethanol: technical progress, opportunities and commercial
challenges. Annual Review of Environment and Resources 24: 189–226.
Yousuf, A. (2012). Biodiesel from lignocellulosic biomass: prospects and challenges. Waste
Management 32: 2061–2067.
Yu, G., Yano, S., Inoue, H. et al. (2010). Pretreatment of rice straw by a hot‐compressed water
pro‐cess for enzymatic hydrolysis. Applied Biochemistry and Biotechnology 160 (2): 539–551.
Zhang, K., Pai, Z., and Wang, D. (2016). Organic solvent pretreatment of lignocellulosic
biomass for biofuels and biochemicals: a review. Bioresource Technology 199: 21–33.
Zhao, X., Zhang, L., and Liu, D. (2012). Biomass recalcitrance. Part I: the chemical
compositions and physical structures affecting the enzymatic hydrolysis of lignocellulose.
Biofuels, Bioproducts & Biorefining 6 (4): 465–482.
Zheng, J. and Rehmann, L. (2014). Extrusion pretreatment of lignocel‐lulosic biomass: a
review. International Journal of Molecular Sciences 15 (10): 18967–18984.
Zhu, L.Y., Zong, M.H., and Wu, H. (2008). Efficient lipid production with Trichosporon
fermentans and its use for biodiesel preparation. Bioresource Technology 99: 7881–7885.
87
Bioelectricity Production from Lignocellulosic Biomass

Samar Das1, Shayaram Basumatary1, Pankaj Kalita1, Vinayak Kulkarni1,
Pranab Goswami1, Akhil Garg2, and Xiongbin Peng2
1
Centre for Energy, Indian Institute of Technology Guwahati, Guwahati, Assam, India
2
Department of Mechatronics Engineering, Shantou University, Shantou, Guangdong, China
6.1 Introduction
The world is witnessing a tremendous industrial development which encourages urbaniza-

tion by creating prodigious economic growth. This industrialization and urbanization is
changing human life and enhancing living standards, which has boosted energy demand
across the world. Higher energy consumption has been experienced especially by develop-
ing countries (Ali et al. 2012; Feng and Lin 2017) and one of the major instruments that
plays a key role in economic progress and human development is access to a reliable and
adequate electricity supply (Kaur and Luthra 2018).
Electricity demand is continuously increasing due to rapid rise in population and indus-
trial development. The growth rate of global electricity generation was estimated to be 2.8%
for 2017, which is higher than the average growth rate of the last decade (2010–2016), and
the average primary energy consumption growth was observed to be 2.2% in 2017, which is
1.2% higher than the previous year and the fastest since 2013 (British Petroleum 2018). It is
also projected that from 2000 to 2030, global primary energy demand will increase by 1.7%
per year but more than 90% of energy demand will be fulfilled by fossil fuel (Bilgen et al.
2004). World energy consumption structure showed the complete dominance of fossil fuel
as coal remains the world’s most important source of power, with a 38.1% share in 2017
which is about 973.4 terawatt‐hours of electricity, followed by natural gas with a global
share of 23.2% (British Petroleum 2018). This excessive use of fossil fuel is creating huge
environmental challenges, including emission of greenhouse gases (GHGs) and degrada-
tion of air quality by releasing pollutants such as SOx, NOx, and particulate matter (Wang
et al. 2017). According to the Intergovernmental Panel on Climate Change (IPCC), con-
tinuous emissions from fossil fuels will result in global temperature rise of between 1.4 and
5.8 °C over the period from 1990 to 2100 (Mahmoud et al. 2009). Environmental effects
including global warming, urban smog, acid rain, etc. have become major threats to modern

society and necessitate a shift toward new options which are less harmful to the environ-
ment and also sustainable (Saidur et al. 2011). In this context, electricity generation from
renewable energy sources will play a crucial role in meeting a variety of primary and sec-
ondary energy needs along with improvement of energy diversity and security, local pollut-
ant and GHG emissions reduction, regional and rural development by providing
employment opportunities, fostering social structure and adding value at the local and
regional level.
Biomass comprises combustible organic materials mainly derived from plant and animal
origin, present in land and aquatic environments (Panwar et al. 2012). It includes a vast
range of feedstocks such as wood and wood waste, farming crops, their residues and by‐
products, municipal solid and animal wastes, by‐products and wastes of process industries,
aquatic plants and algae (Balat et al. 2009a). These are carbon‐neutral, low‐emission, low‐
cost and the most abundantly available renewable sources, with the potential to produce
heat, electricity, fuel, chemicals, and other products (Wang et al. 2017). Biomass is consid-
ered to be carbon neutral because the amount of carbon it releases during its lifetime is
equivalent to the amount it absorbs. Hence, as a renewable source, conversion of biomass
to fuel also supports environmental protection and energy security (Xu et al. 2009;
Kobayashi and Fan 2011). On the other hand, compared to fossil fuel, biomass has lower
heating value on a similar weight basis (e.g., the heating value of biomass, agro‐residues,
and wood materials is in the range of 15–19 GJ/t, 15–17 GJ/t, and 18–19 GJ/t respectively, in
comparison with 20–30 GJ/t for coal). The bulk density or energy density of biomass is
10–40% that of fossil fuels (Zhang et al. 2010).
Another useful way of comparing biomass and fossil fuel is in terms of their H/C and
O/C ratios which can be represented using the van Krevelen diagram, shown in Figure 6.1.
3.3
3.0 Ethanol
2.7
2.4 Gasoline
Atomic H/C ratio (×10–1)
2.1 Kerosene
Treated MSW Juliflora
Diesel
1.8
Ta
1.5 rg et Bio-oil s
asse
Biom Refuse
1.2
te derived
Coal Ligni Sewage sludge fuel
0.9
0.6 Wood
Anthracite
0.3 Bio-char
0.0
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0
Atomic O/C ratio
Figure 6.1 Van Krevelen diagram for various conventional and nonconventional fuels. Source:
Adapted from Reddy and Vinu (2018) with permission from Springer Nature.
Bioelectricity Production from Lignocellulosic Biomass 89
It is evident from the figure that biomass has higher oxygen content compared to coal and
petroleum. Also, the lower the ratios of O/C and H/C, the higher the energy content of the
material (McKendry 2002a).
Application of various conversion routes can enhance the quality of biomass in the form
of bio‐derived fuel and can be utilized in different ways. Direct conversion biomass results
in high‐value end‐products (biochar, biodiesel, biogas, etc.) and with different conversion
technologies, it is possible to obtain fuels in three different forms (solid, liquid, gas) which
is an added advantage over the use of other renewable sources (Hamelinck and Faaij
2006). These biomass‐derived fuels depend on the biomass conversion technologies and
the technologies rely on the type of feedstocks used (Demirbas 2001). There are other fac-
tors which determine the choice of biomass conversion pathway, such as the availability
of biomass, end‐use of the derived fuel, economic and environmental standards etc.
(Saidur et al. 2011).
There are two main routes used to transform biomass into an operational form of
energy: thermochemical conversion and biochemical conversion. In the first, four differ-
ent techniques – combustion, gasification, pyrolysis, and liquefaction – can be used to
harness biomass energy. The second route includes fermentation and anaerobic digestion
which involves microbial activity to release energy from biomass (Zhang et al. 2013). Each
conversion technology has specific criteria which need to be fulfilled by the biomass feed-
stock in order to select the proper conversion route. Identification of feedstock character-
istics is crucial at this point because it not only helps in matching biomass with technology
but also determines to what extent it can be used. Fundamental characterization of feed-
stock also identifies the risks associated with each technology and can be classified into
four groups or classes, “most desirable” as class 1 to “least desirable” as class 4, as pre-
sented in Table 6.1.
Three major products can be obtained from conversion of lignocellulosic biomass: heat/
power generation, transportation fuel, and chemicals. This chapter mainly concentrates on
the thermochemical conversion technologies for generating clean power from lignocellu-
losic biomass. First, the different types of lignocellulosic biomass, its fundamental
composition, and various thermochemical properties are explained briefly. Then the pre-
treatment methods related to these conversion processes are discussed. Different technolo-
gies of thermochemical conversion have also been covered.
6.2 Lignocellulosic Biomass
Lignocellulosic biomass is a term which mainly refers to perennial herbaceous plant spe-
cies and woody crops, covering all forms of agriculture, forestry, agro‐industrial residues,
and energy crops (e.g., cardoon, miscanthus, switchgrass). These feedstocks are abundantly
available in many regions across the globe and do not compete with food sources (Cotana
et al. 2015; Patel et al. 2016). As these biomasses are available from natural resources, feed-
stock cost is less and it is considered to be one of the most sustainable sources of energy for
production of fuels and chemicals around the world. It has been estimated that about
2.2 × 1011 tons of dry lignocellulosic biomass is available annually, out of which only
1.2 × 1010 dry tons are from sustainable sources, and it is predicted that about 38% of the
Table 6.1 Fundamental biomass properties relevant for thermal conversion and anaerobic

digestion.
Classification
Relevant conversion
Property Unit 1 2 3 4 technology
Thermal conversion
Chlorine content wt% d.m. <0.02 0.02–0.1 0.1–0.4 >0.4 Thermal conversion
Ash DT °C >1200 1000– 800– <800 Thermal Conversion
1200 1000
Ash content wt% d.m. <1 1–3 3–10 >10 Thermal and
biochemical conversion
Nitrogen content wt% d.m. <0.3 0.3–1 1–2.5 >2.5 Thermal conversion
Biological conversion
Carbohydrates wt% d.m. >65 50–65 30–50 <30 Biochemical conversion
Lignin content wt% d.m. <10 10–25 25–35 >35 Biochemical conversion
and anaerobic digestion
Anaerobic digestion
Biogas yield m3/ton >300 150–300 50–150 <50 Anaerobic digestion
a.r.
Does the digested Yes No Anaerobic digestion
material have an application?
wt% d.m., weight percentage dry matter (dry basis); a.r., as received (wet basis).
Source: Adapted from Elbersen et al. (2017) with permission from Elsevier.
world’s direct fuel and about 17% of its electricity can be provided using lignocellulosic
biomass (Feng and Lin 2017; Zhao et al. 2017).
6.2.1 Types of Lignocellulosic Biomass

Lignocellulosic biomass can be divided into three categories: hardwood, softwood, and
grasses. There are considerable differences in terms of their chemical and structural
composition which affect their ability to deconstruct or disorganize (Zhu and Pan 2010;
Brandt et al. 2013). Hardwood is mainly characterized by the presence of large water‐
conducting pores or vessels which can be distinguished by their shape, perforated plate
size, and cell wall structure. This type of wood is usually found in broad‐leaved temper-
ate and tropical forests. There is a significant difference between hardwood and softwood
in terms of structural complexity and chemical composition. Hardwoods are generally
much harder than softwoods and slower growing in nature. The most dominant feature
that separates softwood from hardwood is the absence of pores. These woods are nor-
mally found in the northern hemisphere. Softwoods are finely packed with lignin and
hemicellulose that makes them resistant in nature and requires strong pretreatment con-
ditions (Brandt et al. 2013).
Table 6.2 Different types of lignocellulosic biomass.
Feedstock type Resources
Wood Softwood bark, ground softwood, pine, sawdust woodchips, hardwood, mixed
hardwoods
Forest residues Bark waste, peat moss, treetops, limbs
Agricultural Bagasse, cashew nut shell, corn bran
residues
Industrial Lignin from newsprint, paper waste, creosote‐treated wood waste, birch wood
residues waste, wood industry residues, black pulping liquor
Industrial Organosolv lignin, lignin from steam explosion of birch, lignosulfonate
lignins
Source: Adapted from Effendi et al. (2008) with permission from Elsevier.
Grasses are completely different from woods with respect to pore structure. Both annual
and perennial grasses are considered as low‐maintenance feedstocks highly utilized for
production of biofuel. Xylose is the major hemicellulose material present in grass which is
easily degradable. Perennial grasses are more productive but resistant in nature in compari-
son to annual grasses like rice straw, wheat straw, corn stalks, sugarcane bagasse, etc. The
low lignin content present in grasses makes them preferable feedstocks for biorefineries
(Bhowmick et al. 2017; Hassan et al. 2018). Different types of lignocellulosic biomass are
shown in Table 6.2.
6.2.2 Composition of Lignocellulosic Biomass

Lignocellulosic biomass is primarily composed of three major components: cellulose (35–
50%), hemicellulose (20–35%), and lignin (10–25%), along with other elements like lipids,
proteins, etc. (Sawatdeenarunat et al. 2015). The cellulose and hemicellulose, which are
known as holocellulose polymers, contribute roughly two‐thirds of the total dry weight of
biomass (Zabed et al. 2017) and are firmly linked to the lignin through chemical bonds
(hydrogen and covalent bonds), which creates a highly robust structure resistant to
depolymerization.
Cellulose is a linear chain constituting of β‐1,4 linked d‐glucose, having a large number
of hydroxyl groups which create extensive van der Waals and hydrogen bonding networks
within the cellulose molecules and hence form compact crystalline structures of cellulose
(Sawatdeenarunat et al. 2015; Zabed et al. 2017). Hemicellulose is a complex and heteroge-
neous group of branched polysaccharides. Its structural elements consist of different mon-
omers such as glucose, xylose, galactose, glucuronic acid, mannose, and arabinose (Dhyani
and Bhaskar 2018). Lignin is an aromatic phenylpropane‐based polymer (Paul and Dutta
2018). Phenolic compounds like sinapyl alcohol, p‐coumaryl, and coniferyl are the mono-
mers of lignin.
Among the lignocellulosic biomass, woods have a higher proportion of lignin. Softwoods
contain higher amount of lignin, in the range of 30–60%, compared to hardwoods with
30–55%, whereas grasses and agricultural wastes have comparatively lower amounts of
lignin, at 10–30% and 3–15% respectively (Zabed et al. 2017; Paul and Dutta 2018). Lignin
is mainly responsible for the impermeability and rigid structure of the cell walls of plants
and also acts as a biochemical and physical barrier that impedes the conversion processes
of biomass to useful bioenergy (Sawatdeenarunat et al. 2015; Dhyani and Bhaskar 2018).
The composition of lignocellulosic biomass is presented in Table 6.3.
6.2.3 Physiochemical Properties of Lignocellulosic Biomass

Lignocellulosic biomass can be converted into biofuels, bioenergy, and biochemicals by
various processes such as logistics, pretreatment and conversion. Biomass materials are so
diverse in their physical, chemical, and biological properties that there is not a single prop-
erty that is used for a particular process (Capareda 2014). Therefore, prior to design and
implementation of any biochemical or thermochemical processing technique, proper
Table 6.3 Composition of lignocellulosic biomass by dry basis.
Biomass name Cellolose (%) Hemicellulose (%) Lignin (%) Carbon/nitrogen ratio
Rice straw 35–44 27–34 12–13 47–67

Cotton straw 42 12 15
Cotton stalk 31 11 28 41
Wheat straw 38–42 20–27 20–22 50–60
Soybean straw 38 16 16 50
Barley straw 38–48 21–25 11–26 71
Oat straw 33 23 21 95
Rape seed straw 37 25 17 28
Corn stover 40 25–31 14–17 50–63
Corn cobs 45 25 15 123
Sugarcane bagasse 40–45 20–24 25–30 118–150
Poplar 45 21 24 103
Pine 25–44 26–32 28–48 140
Red oak 39 33 23 250
Spruce 28–43 20–30 28–35 170
Cucumber plant 17 17 3 11
Softwood stem 40 30 30 511
Pea vines 40 10 9 120
Green beans 17 16 8 11
Tomato pomace 39 5 11 –
Switchgrass 36–45 28–30 12–26 90
Tomato plant 39 29 12 35
Pepper plant 18 12 8 13
Source: Adapted from Paul and Dutta (2018) with permission from Elsevier.
understanding of the physicochemical properties of biomass is essential as they signifi-

cantly influence the performance of the various conversion processes along with material
handling, storage, transport, pretreatment methods etc. The importance of various physic-
ochemical properties in the area of engineering applications is presented in Table 6.4.
6.2.3.1 Chemical Properties

The identification and characterization of chemical properties of lignocellulosic biomass
are essential for the conversion of biomass feedstock to energy. The elemental composition
of biomass can be investigated by proximate and ultimate analysis which determine the
properties, quality, potential applications, and limitations necessary for designing a system
for energy conversion.
6.2.3.1.1 Proximate Analysis

Proximate analysis is used to investigate the amount of volatile matter (VM), moisture con-
tent (MC), fixed carbon (FC), and ash in the biomass sample. MC represents the amount of
water content in a biomass and is expressed in terms of percentage of weight of the material.
It is an important property of feedstock for biochemical or thermochemical conversion into
biofuels or bio‐oil. It has a strong impact on harvesting and preparation, storage, transport,
processing, and the resultant products (Cai et al. 2017). The VM of biomass indicates the
amount of vapor and gases, except for water, released when biomass is heated at high tem-
perature in the absence of oxygen. Biomass with higher VM increases the production of bio‐
oil. FC is the nonvolatile fraction of biomass which remains after biomass is heated. High FC
content is required for production of biochar (Cai et al. 2017; Dhyani and Bhaskar 2018). Ash
Table 6.4 Various applications for physicochemical properties.
Physicochemical
properties Areas of application
Density Handling, storage, and transportation facilities

Flowability Handling, feeding, and storage facilities
Grindability Grinding facilities
Particle size Feeding, grinding, and storage facilities
Moisture sorption Drying and storage facilities
Moisture content (MC) Handling, storage, drying, feeding facilities and conversion processes
Ash content Estimation of the potential risk of slagging and fouling issues during
biomass combustion or gasification
Volatile matter (VM) Conversion efficiency
content
Elemental composition Conversion efficiency
Energy content Conversion efficiency
Thermal properties Thermochemical conversion efficiency
Chemical composition Conversion efficiency
Source: Adapted from Cai et al. (2017) with permission from Elsevier.
Table 6.5 Proximate analysis of various lignocellulosic biomass feedstocks.
Biomass MC VM Ash Higher heating

feedstock (%) (%) FC (%) (%) value (MJ/kg) References
Softwood 8.8 70.0 28.1 1.7 19.6 Demirbas (1997)

Hardwood 7.8 72.3 25.0 2.7 19.0 Demirbas (1997)
Rice husk 8.47 61.81 16.95 21.24 14.69 Channiwala and Parikh
(2002)
Rice straw 8.10 65.70 13.91 20.38 14.85 Channiwala and Parikh
(2002)
Barley straw 30 46 18 6 16.1 McKendry (2002a)
Sugarcane 8.5 84 1.64 4.5–9 18.17 Al Arni et al. (2010)
bagasse
Bamboo 9.15 73.92 15.30 1.7–5 17.82 Dhyani and Bhaskar (2018)
Wheat straw 8.5 63.0 23.5 13.5 18.7 Demirbas (1997)
Tea waste 6.5 85.0 13.6 1.4 16.8 Demirbas (1997)
Banana leaves 7.8 78.2 15.6 6.2 17.1 Sellin et al. (2016)
Switchgrass 9.3 76.3 10.7 3.6 19.8 Demirbas (2004)
Corn stover 10.6 78.7 17.6 3.7 17.6 Demirbas (1997)
FC, fixed carbon; MC, moisture content; VM, volatile matter.
is the inorganic residue that remains after combustion which may contain SiO2 and CaO and
trace amounts of Al, Mg, K, and P oxides. The ash content varies depending on the type of
biomass. It may be very low (0.5%) for some biomass on a dry basis or up to 20% for cereal and
agriculture waste. The ash content of biomass helps to predict the erosion and abrasion of the
different plant components where a biomass system is used (Nunes et al. 2016). The proxi-
mate analysis of some lignocellulosic biomass feedstocks is presented in Table 6.5.
6.2.3.1.2 Ultimate Analysis

Ultimate analysis is an important aspect of studying the properties of biomass fuels. It is used
to determine the contents of carbon (C), nitrogen (N), hydrogen (H), oxygen (O), and sulfur
(S) in biomass feedstocks so that a better comparison among the feedstocks can be provided.
Ultimate analysis of various lignocellulosic biomass feedstocks is shown in Table 6.6.
Ultimate analysis helps to estimate the heating value and pollution potential of biomass
fuel (SuÁrez et al. 2002; Saidur et al. 2011). Estimation of higher heating value (HHV) from
the elemental composition of fuel is one of the basic aspects in performance modeling and
calculations on thermal systems (Channiwala and Parikh 2002). The heating value of a
biomass fuel can be determined by establishing mathematical empirical formula correla-
tions or by performing experiments using an adiabatic bomb calorimeter. Many research-
ers have correlated the HHV formula based on the data from proximate and ultimate
analyses of biomass fuels. Yin (2011) has also presented different empiric correlations
based on proximate and ultimate analysis of biomass samples developed by various
researchers as shown in Table 6.7.
Table 6.6 Ultimate analysis of various lignocellulosic biomass feedstocks.
Biomass feedstock C (%) H (%) O (%) N (%) S (%) References
Softwood 52.1 6.1 41.0 0.2 1.7 Demirbas (1997)

Hardwood 48.6 6.2 41.1 0.4 2.7 Demirbas (1997)
Sugarcane bagasse 45.04 5.78 47.33 1.75 – Al Arni et al. (2010)
Bamboo 45.88 5.36 37.10 0.32 0.26 Dhyani and Bhaskar (2018)
Wheat straw 45.50 5.10 34.10 1.8 – Demirbas 1997)
Tea waste 48.6 5.5 39.5 0.5 – Demirbas (1997)
Barley straw 45.7 6.1 38.3 0.4 0.1 McKendry (2002a)
Rice straw 41.4 5.0 39.9 0.7 0.1 McKendry (2002a)
Rice husk 38.9 5.1 32.0 0.6 – Raveendran et al. (1995)
Banana leaves 43.5 6.2 42.3 0.86 0.95 Sellin et al. (2016)
Switchgrass 46.7 5.9 37.4 0.8 0.19 Demirbas (2004)
Coconut shell 46.9 6.1 45.7 0.3 0.2 Capareda (2014)
Corn stover 40.0 5.9 46.2 1.3 0.2 Capareda (2014)
Corn stalk 43.83 5.75 48.94 0.97 0.13 Song et al. (2004)
C, carbon; H, hydrogen; N, nitrogen; O, oxygen; S, sulfur.
Table 6.7 Correlated higher heating value (HHV) equation based on proximate and ultimate
analysis of different biomass feedstocks.
Correlated HHV equation based on Correlated HHV equation based on ultimate

Serial no. proximate analysis, MJ/kg analysis, MJ/kg
1 HHV = 19.914 − 0.2324 Ash HHV = 0.4373 C − 1.6701
2 HHV = − 3.0368 + 0.2218 HHV = − 0.763 + 0.301 C + 0.525 H + 0.064 O
VM + 0.2601 FC
3 HHV = 0.3536 FC + 0.1559 HHV = 0.3259 C + 3.4597
VM − 0.0078 Ash
4 HHV = 354.3 FC + 170.8 VM HHV = 3.55 C2 − 232 C − 2230 H + 51.2 C * H
+ 131 N + 20600
5 HVV = 35430 − 183.5 VM − 354.3 Ash HHV = 0.3491 C + 1.178 H + 0.1005 S −
0.10340 O − 0.015 N − 0.0211 * Ash
6 HHV = − 10.8141 + 0.3133 (VM + FC) HHV = 0.2949 C + 0.8250 H
7 HHV = 0.1905 VM + 0.2521 FC
Source: Adapted from Yin (2011) with permission from Elsevier.

C, carbon; FC, fixed carbon; H, hydrogen; N, nitrogen; O, oxygen; VM, volatile matter.
6.2.3.2 Physical Properties

Particle size, density, grindability, flowability, moisture sorption, and thermal properties
are the main physical properties of lignocellulosic biomass.
6.2.3.2.1 Particle Size

The shape and size of particles are an important factor as they affect mixing and fluidiza-
tion, mass transfer, heating, and drying during the combustion process and the flow char-
acteristics of biomass particles. Depending on the shape and size of the feedstock, the
conversion efficiency and energy input requirement can be different. Therefore, optimiza-
tion of accurate particle size can improve the handling, storage, and processing facilities of
thermochemical conversion technology. Maintaining the aspect ratio is necessary for heat
transfer in thermal conversion processing of feedstock. For the characterization of particle
size, different methods like sieving analysis and imaging particle analysis can be performed
(Gera et al. 2002; Cai et al. 2017). Various pretreatment methods can also decrease the par-
ticle size of biomass, which increases the external surface area particle for biochemical
conversion processing so that microorganisms can used for the production of biogas.
6.2.3.2.2 Density
The density of biomass is the measurement of mass of all particles divided by the volume.
The particle density and bulk density are used to characterize the lignocellulosic biomass.
For the calculation of particle density, the pore space volume of the particle is not consid-
ered whereas for the calculation of bulk density, the total volume of particles includes the
pore space volume between and within the biomass particles. This property is important in
designing the system for biomass handling, storage requirements, and transport (Cai et al.
2017) and also for investigation of the behavior of material during thermochemical or bio-
logical processing (McKendry 2002a). Calculation of the bulk density of a biomass sample
can be conducted in accordance with the American Society for Testing and Materials
(ASTM) standard E873‐82 (Cai et al. 2017).
6.2.3.2.3 Grindability
The grindability of a material is a measure of its resistance to grinding. Lignocellulosic
biomass is not easy to grind because it contains very fibrous materials like cellulose and
lignin. Various researchers have reported that there is no standard test for the investigation
of grindability for biomass. However, the Hardgrove Grindability Index (HGI) and Bond
Work Index (BWI), which were developed to determine the resistance of coal and petro-
leum coke, are widely used to find the grindability of biomass. The grindability of biomass
can be greatly improved via the process of torrefaction because it increases brittleness and
reduces cellulose fiber length (Cai et al. 2017).
6.2.3.2.4 Flowability
The investigation of flowability of biomass feedstocks, which measures how biomass flows
from point to point, is also necessary for storage, handling, and transportation (Miccio et al.
2011; Miao et al. 2014). Various researchers have reported that parameters including angle of
response, cohesion coefficient, compressibility index, and flow index (FI) are important to
characterize the flowability of biomass (Lumay et al. 2012; Cai et al. 2017). According to ASTM
standards C144‐00 and D6128‐16, flowability can be calculated in terms of angle of response
(φ) and FI respectively. The angle of response (φ) and FI lie between 0–90° and 1–10°
respectively. Based on these parameters, flowability of biomass can be generally classified as
cohesive, very cohesive, high flowing, medium flowing, and low flowing (Cai et al. 2017).
6.2.3.2.5 Thermal Properties

Thermal conductivity and specific heat are the two important thermal properties of
biomass (Cai et al. 2017) which influence the thermochemical conversion characteris-
tics. Specific heat is a measure of the heat capacity of a material which is influenced by
both temperature and moisture. This property is important for thermodynamic calcu-
lations of processes (Dupont et al. 2014; Cai et al. 2017). Thermal conductivity is the
ability of a material to transfer or conduct heat along and across its fibers, which
depends on temperature, heating direction, moisture, porosity, and density (Cai et al.
2017). Thermal conductivity is used for modeling biomass in storage systems to meas-
ure its ability to conduct heat through an enclosure and also its ability to store heat
energy (Capareda 2014).
6.2.3.2.6 Moisture Sorption

The characterization of water sorption behavior of biomass is an important factor for har-
vesting, handling, transport, and storage. The equilibrium moisture content (EMC) is used
to determine the moisture sorption of biomass which depends on the composition, specific
surface area, porosity, and microstructure. It can be defined as the MC in biomass feedstock
in equilibrium in a particular environment with respect to relative humidity and tempera-
ture (Lin et al. 2016; Cai et al. 2017).
6.3 Pretreatment of Lignocellulosic Biomass
Pretreatment of lignocellulosic biomass is the process by which a complex lignocellulose

can be converted into simpler components such as cellulose, hemicellulose, and lignin
(Kumari and Singh 2018). The selection of pretreatment should consider the compatibility
of feedstock, enzymes, and organisms (Chen et al. 2017) and desired product. It should be
simple, eco‐friendly, cost‐effective, and economically feasible but pretreatments should not
give rise to inhibitory compounds or loss in the fraction of polysaccharide or lignin (Hassan
et al. 2018).
Pretreatments can be classified into different categories: physical, chemical, biological,
physicochemical, and any combination of these. In general, physical and chemical pre-
treatments are considered relatively better but high‐quality and noncorrosive equipment is
required and they also produce severe environment pollution. The biological method pol-
lutes less and has a lower energy requirement for the treatment process than other meth-
ods, but it is costly and enzyme activities for the decomposition of lignocellulose are also
low (Chen et al. 2017). The different pretreatment methods used for biofuel production are
discussed below.
6.3.1 Physical Pretreatments

Physical pretreatments are used to reduce particle size, crystallinity (Onumaegbu et al.
2018), and degree of polymerization of the cellulose present in lignocelluloses (Behera
et al. 2014) during biomass cell disintegration and to increase specific surface area and pore
size of lignocellulosic biomass (Behera et al. 2014; Kumari and Singh 2018). There are
v arious physical pretreatment methods such as grinding, milling, chipping, freezing,

irradiation, etc. or combinations of these can be used to improve the enzymatic digestibility
of biomass waste materials (Kumari and Singh 2018). However, the selection of pretreat-
ment method mainly depends on the characteristics of biomass substrate or feedstock to be
used and hence, it is difficult to compare the applicability and sustainability of such meth-
ods at full scale systematically (Ariunbaatar et al. 2014). Various researchers report that
size reduction is mostly used for the hydrolysis process but many of them are not feasible
economically since energy requirement is high for these methods. Kumari and Singh
(2018) reported that extrusion is an efficient physical pretreatment for reduction of particle
size which also includes heating, mixing and shearing processes resulting in modification
of the physical and chemical properties of the biomass.
6.3.2 Chemical Pretreatments

Chemical pretreatment is used to achieve destruction of the organic compounds of ligno-
cellulosic biomass by means of strong acids, alkalis, organic solvents, and ionic liquids. The
performance of chemical pretreatment also depends on the type of method applied and the
characteristics of the substrates. This pretreatment is not feasible for substrates which are
easily biodegradable or feedstocks which contain high amounts of carbohydrates because
their degradation rate is high and they may accumulate volatile fatty acids, which leads to
failure of methanogenesis (Ariunbaatar et al. 2014; Paudel et al. 2017) although it increases
the biodegradability of lignocellulosic substrates with a high lignin content (Fernandes
et al. 2009).
Alkaline pretreatment of biomass uses various alkali compounds such as calcium
hydroxide, potassium hydroxide, sodium hydroxide, aqueous ammonia, ammonium
hydroxide, and hydrogen peroxide. This method causes swelling of biomass which increases
the internal surface area of the biomass and decreases cellulose crystallinity and degree of
polymerization (Agbor et al. 2011; Behera et al. 2014; Chen et al. 2017). This method
destroys the lignin structure and breaks the linkage between lignin and carbohydrate in
lignocellulosic biomass (Chen et al. 2017; Paudel et al. 2017) which makes the carbohy-
drates in the heteromatrix more accessible (Agbor et al. 2011).
For acid pretreatment of biomass, both organic and inorganic acids such as sulfuric acid,
nitric acid, hydrochloric acid, phosphoric acid, nitrous acid, formic acid, acetic acid etc.
have been used for enzymatic hydrolysis, or for hydrolyzing the lignocellulose to enhance
biofuel production (Behera et al. 2014; Kumari and Singh 2018). Acid treatment can be
performed under either high concentrated acid (e.g., 30–70%) and low temperature (e.g.,
40 °C) or dilute acid (e.g., 0.1%) and high temperature (e.g., 230 °C) (Zheng et al. 2014).
Even though concentrated acid treatment can be more effective for the hydrolysis of cel-
lulose, it is toxic in nature and corrosive and hence nonmetallic materials or alloys which
is very costly are required for the construction reactors (Behera et al. 2014; Kumari and
Singh 2018). Hence, dilute acids (<4% w/w) can be used in acid pretreatment to hydrolyze
biomass (Agbor et al. 2011).
Acid pretreatment may require the use of an alkali to neutralize the hydrolyzate (Kumari
and Singh 2018). Therefore, the use of suitable acids, concentrations, and reaction tem-
peratures should be maintained to decrease the production of inhibitors.
6.3.3 Biological Pretreatments

Biological pretreatment is mostly connected with the action of microorganisms such as
brown, white, and soft rot fungi that are capable of producing enzymes for degradation of
lignin and hemicelluloses (Sindhu et al. 2016). It has many advantages over physical or
chemical pretreatments because the energy requirement is low, it does not generate any
toxic compounds, there is a high yield of desired products (Behera et al. 2014) and it does
not require acids, alkali or any reactive species. However, biological pretreatment is slower
than nonbiological methods and requires careful control for growth and more space to
perform which make this method less attractive on an industrial scale (Agbor et al. 2011;
Behera et al. 2014). However, pretreatment of biomass should be environmentally friendly
so with the development of science and technology, and improved genetic engineering,
bacteria will play an important role in the biological treatment process which makes the
application of biological methods more attractive (Chen et al. 2017).
6.3.4 Physicochemical Pretreatment

Combined pretreatment with chemical and physical processes can improve the accessibil-
ity of the cellulose for hydrolytic enzymes by solubilizing hemicelluloses and destroying
lignin structure (Behera et al. 2014). Rabemanolontsoa and Saka (2016) reported that in
order to enhance lignin removal and increase efficiency, physical parameters such as pres-
sure were added to chemical alkaline pretreatment. Various physicochemical pretreatment
methods such as steam pretreatment, steam explosion, wet oxidation, freeze explosion,
liquid hot water pretreatment, aqueous ammonia pretreatment, organosolv pretreatment,
and CO2 explosion have been performed by recent researchers. The use of ionic liquids and
cellulose solvent‐based lignocellulose fractionation have also been proposed recently
(Agbor et al. 2011). These pretreatments depend on process conditions and the use of sol-
vents which affect the physical and chemical properties of the biomass.
6.3.5 Combined Pretreatment Method

In this method, two or more treatment methods are combined. The combination may be
mechanical crushing‐chemical, physical or biological treatment, mechanical crushing‐
microwave‐chemical processing, mechanical crushing‐chemical treatment‐steam explosion,
mechanical crushing‐electronic radiation‐alkali treatment (Chen et al. 2017). A single pre-
treatment method does not provide effective results because of its limited specific functioning
mode and intrinsic disadvantages (Zheng et al. 2014). It may also have technologic challenges,
produce pollution, have huge energy consumption, be a slow process and corrosive for equip-
ment (Chen et al. 2017). However, combined pretreatment methods have several advantages
over single pretreatments, such as improving the efficiency of hydrolysis, reducing pretreat-
ment severity, increasing methane production, and allowing more complete utilization of bio-
mass. But they may increase the cost of pretreatment, so an economic analysis should be
performed for combined biomass pretreatment methods (Zheng et al. 2014; Chen et al. 2017).
Several pretreatment methods have been investigated by various researchers for enhanc-
ing biofuel production. These processes are still in development. Among these methods,
physical and chemical pretreatments are very costly, have high energy consumption and
produce environmental pollution whereas biological methods consume less energy and
produce less environmental impact but the process is very slow. A comparative study of
various treatment techniques is presented in Table 6.8.
Table 6.8 Comparison of different pretreatment methods for lignocelluloses.
Pretreatment method Advantages Disadvantages
Mechanical Reduce particle size and cellulose Cannot remove lignin and
splintered crystallinity hemicelluloses, high energy
Microwave Simple operation, energy High cost
Physical efficient, short time
pretreatment Ultrasonic Improve accessibility and Negative to enzymatic
reactivity of cellulose hydrolysis
High‐energy Reduce cellulose polymerization High cost
electron degree
radiation
High‐ Decompose cellulose rapidly Energy consumption, low
temperature productivity
pyrolysis
Concentrated High sugar conversion High toxic and corrosive,
acid corrosive to equipment,
high cost
Dilute acid Fast and do not need to recycle High temperature and
Chemical acid pressure, formation of
pretreatment inhibitors
Alkali Room temperature, destroy lignin Less sugar degradation
pretreatment
Oxidation Environmental, remove lignin High cost
pretreatment effectively
Organosolv Obtain pure lignin, cellulose, and High cost, certain effects
pretreatment hemicelluloses on environment and
fermentation
Ionic liquid Environmentally friendly, large High cost
pretreatment temperature range
Steam Lignin transformation, High temperature and
explosion hemicellulose solubilization pressure
AFEX Cost‐effective, increase surface High cost, not efficient for
Physicochemical method CO2 area of cellulose, no inhibition raw high lignin content
pretreatment explosion substances formed material
Electrical Does not produce inhibition High pressure, do not affect
catalysis compounds, cost‐effective, lignin and hemicelluloses
increase surface area, effective Lower efficiency
removal of lignin, clean
Biological Degrade lignin and hemicellulose Low rate of hydrolysis
pretreatment — Low energy consumption
Source: Adapted from Chen et al. (2017) with permission from Elsevier.
AFEX, ammonia fiber expansion.
6.4 Thermochemical Conversion of Lignocellulosic Biomass
Thermochemical decomposition techniques can be applied for energy conversion of all

types of biomass sources but low MC lignocellulosic materials such as herbaceous and
woody biomasses are considered to be the most suitable feedstocks (Demirbas 2001).
The thermochemical conversion process has four different approaches: direct combus-
tion, pyrolysis, liquefaction, and gasification. Figure 6.2 represents the various primary and
secondary products available from these conversion routes. Heat, fuel gas or syngas, liquid
fuel, and charcoal are the primary products from which can be derived secondary products
like electricity, biofuels, chemicals, ammonia, and methanol. Primary products can be
directly utilized as a combustion source for production of heat and power in a power plant
and through various advanced purification techniques, they can be further processed in
higher energy efficient technologies such as gas turbines, internal combustion engines,
molten carbonate fuel cells (MCFC), etc. for production of secondary products (Molino
et al. 2016).
6.4.1 Direct Combustion

Direct combustion can be described as the burning of fuel in the presence of air without
any chemical processing to convert the stored chemical energy into heat along with carbon
Extraction
Chemicals
Charcoal
Upgrading
Pyrolysis
Biofuels
Turbine
Liquid
Liquefaction
Synthesis
Methanol
Gasification
Fuel Gas
Engine
Electricity
Combustion
Heat Fuel Cell
Ammonia
Boiler
Thermochemical Conversion
Primary Products Secondary Product
Process Technology
Figure 6.2 Different thermochemical conversion routes for electricity generation. Source: Adapted
from Molino et al. (2016) with permission from Elsevier.
dioxide and water. Conversion of traditional biomass through combustion is a common

process in most of the underdeveloped nations and plays a crucial role in day‐to‐day life as
it is the main energy source available for cooking and heating. Combustion is a low‐cost,
well‐understood and highly reliable technology available commercially (Zhang et al. 2010).
Any type of biomass can be burned but practically, biomass with MC less than 50% is best
for combustion. Biomass available naturally is rarely in an acceptable form for combustion
and therefore in most cases it undergoes some pretreatment like drying, chopping, grind-
ing, etc. which in turn increases the cost and energy expenditure associated with the overall
conversion process (Lee et al. 2013).
Direct combustion of biomass is a combination of consecutive homogeneous and hetero-
geneous reactions and it depends on the properties of the biomass feedstock, particle size,
operating temperature, and atmosphere of combustion. From the environmental point of
view, biomass combustion is a carbon‐neutral phenomenon but emission of CO2, NOx, and
particulate matter is high and this along with ash handling makes it highly challenging
(Patel et al. 2016). Combustion can be widely used on various scales to convert the stored
chemical energy of biomass into heat and/or mechanical power or electricity through dif-
ferent equipment such as stoves, boilers, turbines, turbo‐generators, etc. Domestic cooking
and space heating are small‐scale applications with heat loss of 30–90% which is very inef-
ficient, but with the use of improved stove technology, this problem can be solved.
Combustion can also be applied on a large scale in boilers to generate steam which is one
option for generating clean electricity from biomass. This approach is discussed in the fol-
lowing sections (Demirbas 2001; Naik et al. 2010).
6.4.2 Liquefaction
Liquefaction is a low‐temperature, high‐pressure thermochemical process used to convert
biomass into liquid fuel in the presence of hydrogen with the help of a catalyst (Demirbas
2001; McKendry 2002b; Goyal et al. 2008). It is also known as hydrothermal or aqueous
liquefaction because water plays a vital role in this conversion. In this process, biomass
disintegrates into small molecules in water or another solvent. These fragments, which are
light, unstable, and reactive in nature, then repolymerize to form oily hydrocarbon com-
pounds with different kinds of molecular weight.
Direct liquefaction is somewhat similar to pyrolysis in terms of target products (i.e., liq-
uid fuel) but there are several differences between them in terms of operational conditions.
In direct liquefaction, a pressure of 5–20 MPa is applied whereas in pyrolysis applied pres-
sure ranges from 0.1 to 0.5 MPa. Another advantage of liquefaction over pyrolysis and gasi-
fication is that dried feedstock is not necessary for liquefaction which reduces energy
consumption as well as the number of operations required to convert biomass into liquid
fuels. Moreover, the presence of water also allows the reaction to occur at a lower tempera-
ture compared to flash pyrolysis. The operating temperature of liquefaction is between 300
and 400 °C and it has a longer residence time than gasification and pyrolysis (0.2–1.0 h)
with a high water:biomass ratio of 3:1–10:1 (Zhang et al. 2010; Patel et al. 2016; Ibarra‐
Gonzalez and Rong 2018). For liquefaction, catalysts are always necessary but for pyrolysis
they are not important. Liquefaction is a less commercial thermochemical conversion pro-
cess because compared to pyrolysis, it requires more complex and expensive equipment
and fuel feeding mechanisms (Zhang et al. 2010). Moreover, bio‐oil viscosity is higher and
production yield is lower in liquefaction in comparison with fast pyrolysis and catalytic
upgradation of these oils could lead to hydrocarbon‐rich organic distillates and useful
chemicals (Naik et al. 2010).
Liquefaction begins with the depolymerization of biomass into monomers which then
repolymerize or condense to form undesirable solid char through high‐order solid‐state
reactions. Addition of solvent slows down this harmful condensation reaction (Zhang et al.
2010). The heavy oil produced from liquefaction is insoluble in water and viscous in nature
which sometimes causes problem during handling. This can be reduced by adding organic
solvents like propanol, butanol, acetone, etc. to the reaction system. Along with the sol-
vents, some solvent‐reducing gases such as CO or H2)and/or catalysts are also necessary.
Presence of a catalyst reduces the temperature required for liquefaction, improves reaction
kinetics and also enhances the oil yield during the process up to 63% whereas with non-
catalytic liquefaction, an average of 31% is reported (Naik et al. 2010). Some solvents and
catalysts used by various researchers are shown in Table 6.9.
6.4.3 Pyrolysis
Pyrolysis is the thermal degradation of biomass by application of heat in the absence of
oxygen/air which in turn results in production of three phases of products: biochar (solid),
bio‐oil (liquid), and fuel gas (gaseous) (Naik et al. 2010). There are various operating condi-
tions such as temperature, heating rate, residence time, particle size, and use of catalyst
which determine the main output products of biomass pyrolysis. Generally, the product of
woody biomass pyrolysis consists mainly of hydrogen (H2) along with CO, CO2 and meth-
ane (CH4). Other organic compounds are also present in the product (Saidur et al. 2011).
Desired products from biomass pyrolysis can be obtained by varying the operational condi-
tions. To obtain a high yield of liquid fuel, low temperature with high heating rate and
short residence time is preferred. For higher yield of charcoal, low temperature and slow
heating rate would be chosen. To obtain the maximum yield of gaseous fuel, high tempera-
ture, long residence time and low heating rate would be used (Demirbas 2001).
Table 6.9 Summary of research on direct liquefaction of biomass.
Feedstock Solvent Catalyst
Crop residues, corn stover, rice straw, Ethylene carbonate, ethylene glycol, Sulfuric acid
wheat straw polyethylene glycol
Pine wood, poplar wood Ethanol and small amount of phenol Sulfuric acid,
sulfonic acids
Sawdusts of white birch, Japanese cedar, Ethylene glycol, ethylene carbonate, Sulfuric acid
Japanese cypress propylene carbonate
Wood meal of birch and aspen, Phenol Alkalis or salt
thermomechanical pulp, kraft pulp,
cotton, jute fiber, kenaf plant meal
Source: Adapted from Zhang et al. (2010) with permission from Elsevier.
Based on the operating conditions, pyrolysis can be categorized into three classes: slow
or conventional pyrolysis, fast pyrolysis, and flash pyrolysis.
6.4.3.1 Slow Pyrolysis

Slow pyrolysis can also be termed conventional pyrolysis during which biomass is con-
verted to a carbon‐rich solid residue under the condition of low heating rate of 0.1–1 K/s
and a long vapor residence time of 45–550 seconds (Naik et al. 2010). On the basis of ther-
mogravimetric analysis of biomass, this pyrolysis process can be differentiated into three
stages. The first stage, known as prepyrolysis, occurs at a temperature range of 120–200 °C.
In this stage, a slight weight loss can be observed due to some internal reorganization such
as breaking of bonds. Free radicals also appear along with the formation of carbonyl groups
and release of CO, CO2 and small amounts of water (H2O). The second stage is the main
transition phase of biomass pyrolysis where a significant amount of weight loss takes place
due to solid decomposition of biomass feedstock. The third stage is followed by the con-
tinuous devolatization of char with a sharp dissection of C─O and C─H bonds. It has been
reported that higher content of lignin and lower content of hemicellulose in biomass
results in higher charcoal yield (Zhang et al. 2010).
6.4.3.2 Fast Pyrolysis

Fast pyrolysis is the process of producing a high yield of liquid fuel. It involves higher oper-
ating temperatures of 580–1250 K, rapid heating rate of 10–200 K/s, and short solid resi-
dence time of 0.5–10 seconds. Unlike slow pyrolysis, it requires fine feedstock particle size
lower than 1 mm.
Fast pyrolysis starts with decomposition of biomass to produce vapors, aerosol, and some
solid char. Then condensation and cooling of these vapor and aerosol form a dark brown
liquid with a lower heating value compared to conventional fuel (Naik et al. 2010). This
bio‐oil is composed of both aqueous and nonaqueous phase (tar) compounds. The aqueous
phase contains various low‐weight organooxygen compounds, whereas the nonaqueous
phase contains higher molecular weight insoluble aromatic organic compounds. Pyrolized
oil has high water content (15–30 wt%), much higher oxygen content (35–40 wt%), low pH
(<3), which is corrosive, and low heating value (16–19 MJ/kg) in comparison with heavy
petroleum oil. At a temperature range of 700–1000 °C, fast pyrolysis produces hydrogen gas
converting methane, vapors of other hydrocarbons and simple aromatics through steam
reforming and water–gas shift reactions (Zhang et al. 2010). Most common reactors used
for fast pyrolysis are bubbling and circulating fluidized bed, entrained bed, wire mesh,
vacuum furnace, rotating and cortex (Goyal et al. 2008).
6.4.3.3 Flash Pyrolysis

Compared to conventional and fast pyrolysis, flash pyrolysis is done at a much higher tem-
perature range of 1050–1300 K with a rapid heating rate of more than 1000 K/s and resi-
dence time less than 0.5 seconds. To maintain this low residence time, a special reactor
arrangement is required. Particle size for this process are very fine, less than 0.2 mm. Bio‐
oil produced from flash pyrolysis can be mixed with char to form bioslurry which can be
another option for gasification with higher conversion efficiency. Moreover, conversion
efficiency of biomass to oil in flash pyrolysis is about 70% and this bio‐oil can be processed
Table 6.10 Product yield of different pyrolysis processes.
Product yield
Conversion process Liquid (wt%) Gas (wt%) Solid (wt%)
Slow pyrolysis 30–50 15–30 12–19

Fast pyrolysis 65–75 13–25 12–19
Flash pyrolysis 60–70 10–15 15–25
Vacuum pyrolysis 45–60 17–27 19–27
Source: Patel et al. (2016) with permission from Elsevier.
in engines and turbines for power production (Naik et al. 2010). Flash pyrolysis can be
done in hydrogen medium which is known as flash hydropyrolysis. In solar flash pyrolysis,
various solar devices such as solar tower, concentrated solar dish, etc. are used for heating.
Vacuum flash pyrolysis conducted in vacuum atmosphere enhances oil yield (Goyal et al.
2008). Table 6.10 presents the product yield of different types of pyrolysis.
6.4.4 Gasification
Gasification is a form of pyrolysis performed in partial oxidation conditions with the appli-
cation of high temperature (800–900 °C or even higher) in order to maximize the production
of combustible gas (Demirbas 2001; McKendry 2002b). The gasification process includes
several chemical reactions of biomass with air/oxygen or steam to form a gaseous product
which is a mixture of CO, H2, CO2, CH4, and N2 generally known as producer gas, syngas or
synthesis gas with respect to the ratios of these gas components (Rowlands et al. 2008).
The gasification process comprises four steps: drying, devolatization/pyrolysis, reduc-
tion, and combustion. In the drying stage, removal of moisture from the biomass feedstock
takes place. Then in the pyrolysis zone, volatiles present in the feedstock are removed
resulting in the formation of tar composed of liquid long chain hydrocarbons along with
light hydrocarbons, CO, and CO2. The operating conditions, nature of feedstock, and gasi-
fying medium highly influence the production of tar. A series of endothermic reactions
occurs to form producer gas in the reduction zone in the presence of gasifying medium
(air/oxygen/steam) which is considered to be the major zone of gasification. Finally, in the
combustion stage, residual char undergoes oxidation to form more gaseous products along
with heat which is utilized in the reaction of reduction zone (Damartzis and Zabaniotou
2011). Gasification composed of various oxidation and reduction reactions is shown in
Table 6.11.
There are several operating parameters which influence the composition of producer gas
obtained from gasification: fuel composition, MC of feedstock, operating pressure and tem-
perature, gasifying medium, and flow direction of gasifying medium. Prediction of the
exact composition of producer gas coming out of a gasifier is difficult but the water–gas
equilibrium concept gives the opportunity to determine the exact producer gas composi-
tion theoretically (Balat et al. 2009b). Two different approaches can be followed to produce
Table 6.11 Series of gasification reactions.
Exothermic reactions
C(s) + O2 ↔ CO2 ΔHR = − 393kJ/mol
1 ΔHR = − 110kJ/mol
Cs O2 ↔ CO
2
1 ΔHR = − 242kJ/mol Combustion reactions
H2 O2 ↔ H2O
2
1 ΔHR = − 283kJ/mol
CO O2 ↔ CO2
2
C(s) + 2H2 ↔ CH4 ΔHR = − 74kJ/mol Hydrogasification reaction
CO + H2O ↔ CO2 + H2 ΔHR = − 42kJ/mol Shift reaction
Endothermic reactions
C(s) + CO2 ↔ 2CO ΔHR = + 173kJ/mol Boudouard reaction
C(s) + H2O ↔ CO + H2 ΔHR = + 132kJ/mol Water‐gas reaction
CH4 + H2O ↔ CO + 3H2 ΔHR = + 206kJ/mol Steam‐methane reforming reaction
syngas from biomass: catalytic and noncatalytic process. The catalytic method operates at
low temperature whereas for the noncatalytic process, higher temperature of about 1300 °C
is required. With the advancement of catalysis, the higher temperature can be expected to
fall to about 900 °C (Naik et al. 2010). Producer gas has very low energy density, about half
that of natural gas, and primarily can be utilized for steam cycle, gas engine, gas turbine
cycle, or fuel cell to produce electricity. It can also act as intermediate feedstock for the
production of various fuels and chemicals which include hydrogen, diesel, methanol,
ammonia, dimethyl ether, etc. (Ibarra‐Gonzalez and Rong 2018).
According to the interaction of gasifying medium and solid fuels, gasifiers can be
classified into three main categories: fixed bed gasifier, fluidized bed gasifier, and
entrained flow gasifier. The fixed bed gasifier can be further classified into three types
on the basis of direction of flow of the gasifying agent into updraft, downdraft, and
cross‐draft gasifiers. Based on the velocity of flow of gasifying agent, fluidized‐bed gas-
ifiers can be distinguished into two different types: circulating fluidized bed (CFB) and
bubbling fluidized bed. Among all the gasifiers, downdraft, updraft, bubbling bed, and
CFB gasifiers are most commonly used in the industry. Commercially, about 75% of
gasifiers used are downdraft, 20% fluidized bed, 2.5% updraft, and 2.5% of the other
types (Jayah et al. 2003). Various operating characteristics of all these types are pre-
sented in Table 6.12.
6.5 Electricity Production from Thermochemical Conversion
The various types of combustible fuel generated from the thermochemical conversion
process can be utilized to generate electricity through different conversion technologies.
On the basis of market potential and overall technology reliability, Molino et al. (2016)
Table 6.12 Characteristics of different types of gasifier.
Gasifier type Characteristics Schematic diagram
Fixed bed Updraft ●● Biomass is fed from the top Advantages

gasifier ●● The gasifying agent flows ●● Effective heat transfers due to countercurrent flow Gas
from the bottom ●● Can process fuel with high moisture
●● The biomass and gas move in ●● Less sensitive to the amount of ash
opposite directions ●● Less complex fuel preparation is required
●● Resulting char falls and burns ●● High cold gas efficiency
to provide heat
●● High methane content in product gas
●● Gas leaves from the top
●● Relatively simple and low‐cost process
●● Tar‐rich gas
Disadvantages
●● The ash falls from the grate at Air Ash Pit
●● Higher amount of tar in the product gas
the bottom
●● Not suitable for scale‐up
●● Limited ability to handle fines from fuel
●● Long feedstock residence time
Downdraft Biomass is fed from the top Advantages

gasifier The gasifying agent flows from ●● Relatively clean fuel production with less tar, ash,
top to bottom and soot
The biomass and gases move in ●● Easier to control
the same direction ●● Low sensitivity to fines from fuel
Biomass is burnt, falling ●● Requires minimal tar clean‐up system
through the throat to ●● Relatively simple and low‐cost process
form a bed of hot charcoal Disadvantages
The gases have to pass through Air Air
●● Fuel specific
a reaction zone
●● Excessive pressure drop
Gas leaves at the bottom, with
●● Drying of fuel to a low moisture content is required
ash collected under the grate Gas
●● Lower efficiency due to lower carbon conversion
Ash Pit
●● Impractical for scale‐up
(Continued)
Cross‐draft ●● Biomass is introduced from the Advantages

gasifier top ●● Compact and can be operated at small scale
●● The gasifying agent is ●● Gas cleaning requirement is less
introduced from one side of ●● Faster start‐up time
the gasifier,
●● The load following ability is quite good
●● the gas is taken out from the
●● No need for grate
other side
Disadvantages Gas
●● Entry of gasifying agent and Air
exit of fuel gas are from the ●● Poor CO2 reduction and high gas velocity
same level ●● Requires low‐ash fuel

Combustion Reducing
●● Oxidation zone is placed near ●● High exit gas temperature reduces thermal efficiency zone zone
the entry of gasifying zone ●● Not suitable for scale‐up Ash pit
●● Reduction zone is placed near
the producer gas exit
●● Higher CO, low H2 and CH4
Fluidized Circulating ●● A bed of fine inert material Advantages
exiting gas
bed bed ●● Oxidant blown upwards ●● High conversion rates and particles
not captured
through it fast enough ●● Low‐quality fuel can be utilized by cyclone
(5–10 m/s) to suspend ●● Very high throughput
material throughout the cyclone
●● Suitable for rapid reactions
gasifier circulating
●● Biomass is fed in from the side ●● High possibility of scale‐up feeding
solid fuel
fluidized
bed
●● High heat and mass transport rates
●● The mixture of producer gas particles returning
and particles is separated ●● Low tar production leg
using a cyclone, Disadvantages

larger particles
●● material is returned into the ●● Temperature gradients occur in the direction of solid bubbling
base of the gasifier flow Air or other gases fluidization
●● Operates at temperatures ●● Minimum transport velocity

below 900 °C to avoid ash ●● Erosion due to high velocity of particles
melting and sticking ●● Heat exchange is less efficient
●● More expensive
Bubbling bed ●● A bed of fine inert material Advantages

Gas
sits at the bottom, ●● High throughput
●● with gasifying agent being ●● Fuel flexibility
Solid
blown upwards through the ●● Tolerates reasonably high moisture
bed just fast enough (1–3 m/s)
●● Less pollution (no NOx and SOx, less CO2) due to
to agitate the material
low temperature operation
●● Biomass is fed in from the
side, mixes, and combusts or Disadvantages
Gas bubble
forms producer gas which ●● Large bubbles may create problem as result of gas
leaves upwards bypassing through the bed Solid par ticle Solid
●● Operates at temperatures ●● The gasification reactions cannot reach their
below 900 °C to avoid ash chemical equilibrium because of relatively short Gas Distributor
melting and sticking residence times
●● Tar formation is more than downdraft gasifier
(without catalyst)
Dual fluidized ●● System has two Advantages
air reactor fuel reactor
bed chambers – gasifier and ●● Gasification and combustion reactions are exhaust exhaust
combustor separated
●● Biomass is fed into the CFB/ ●● Nitrogen‐free product gas is possible
BFB gasification chamber F
●● No external heat supply is required A U
●● The char is burnt in air in the ●● Circulation of hot particles supplies heat for I E
CFB/BFB combustion gasification R L
chamber, heating the
●● Gasification of high volatiles is possible R R
accompanying bed particles
●● Possibility to scale up E E
●● Hot bed material is fed back A Steam A
into the gasification chamber, Disadvantages C
upper
loop seal C
providing the indirect ●● Difficult to maintain continuous circulation of T T
reaction heat particles O lower O
loop seal internal
R R loop seal
●● Temperatures below 900 °C ●● Relatively complex construction makes it expensive
steam
●● Difficult to avoid slight gas mix between chambers
steam
air fuel
(Continued)
Entrained flow ●● Powdered biomass is fed with Advantages Oxygen

pressurized oxygen and/or ●● High throughput and suitable for high capacities
steam ●● High carbon conversion efficiency
A turbulent flame at the top Biomass
●● High‐quality product gas
●●
●● burns some of the biomass, ●● Low tar yield due to high‐temperature operation
providing large amounts of
●● Short residence time
heat, at high temperature
(1200–1500 °C) Disadvantages
●● Very high quality producer ●● Fuel preparation cost is high due to very fine
gas particle More sophisticated reactor design

●● The ash melts onto the walls, ●● Some fuels can form corrosive slag 1300°C
and is discharged as molten ●● Low cold gas efficiency
slag ●● Higher amount of gasifying agent required
Water cooled
radiation screen
Raw Syngas and

Molten Slag
Source: All images in the table are modified from Loha et al. (2018) with permission from Springer Nature.
BFB, bubbling fluidized bed; CFB, circulating fluid bed.
Overall Technology Reliability

High Low
High
Market Potential
Co-firing
Engine
IGCC
Low
Steam cycle
Fuel cell
Biofuels
Figure 6.3 Status of various power generation technology. IGCC, integrated gasification combined
cycle. Source: Adapted from Molino et al. (2016) with permission from Elsevier.
c ategorized different technologies for power generation from thermochemical conversion

as shown in Figure 6.3. This section discusses some of the technologic options for produc-
ing clean electricity.
6.5.1 Steam Cycle

On a large scale, biomass combusted in a boiler to produce steam and then generating elec-
tricity through a steam cycle is a comprehensive substitute for fossil fuel and these plants
are in the range of 100–3000 MW. Sulfur emissions (0.05–0.2 wt%) are much lower and the
formation of particulate can be controlled at the source The efficiencies of large‐scale bio-
mass power generating units with capacity more than 100 MW are in the range of 20–40%,
which is comparable to conventional fossil fuel‐based plants although higher cost is associ-
ated with these systems due to the MC present in biomass. Syngas and cracking oil pro-
duced from other thermochemical process can also be combusted in a boiler and processed
through a steam cycle to generate power in an environmentally friendly way. Syngas‐ or
cracking oil‐fueled power plants are reported to be running without any problem but the
overall electrical efficiency of these types of power plants is reported to be very low, varying
from 15% to 35% with respect to boiler size higher than 150 MW (Bridgwater et al. 2002;
Yassin et al. 2009). Significant improvement in economic status as well as in efficiency for
biomass‐based systems can be achieved through co‐generation, i.e., combined heat‐power
system. Moreover, co‐combustion or co‐firing in coal‐based power plants is considered to
be an attractive option for further improvements in power generation (Demirbas 2001;
Naik et al. 2010).
6.5.2 Co-Firing
Co‐firing or co‐combustion of biomass and coal has three different approaches: direct co‐
firing, indirect co‐firing and parallel co‐firing, as shown in Figure 6.4. In direct co‐firing,
blending the biomass and coal occurs in the fuel handling system and is then fed to the boiler.
In indirect co‐firing, syngas produced from biomass gasification is either burned with coal in
a boiler or utilized in a combine gas turbine cycle for power generation. In parallel co‐firing,
biomass handling and feeding is done separately without hampering the coal handling sys-
tem and then injected to the boiler (Saidur et al. 2011). These methods are important from the
standpoint of achieving maximum use of biomass for electricity generation. For this technol-
ogy, very few modifications are required in existing coal‐based plants which makes it cost‐
effective compared to other thermochemical processes of biomass conversion, including
single biomass‐based combustion (Zhang et al. 2010). Out of these three types, direct co‐fir-
ing is the most common approach because it is low cost, simple and, most importantly, with-
out any significant expenditure; approximately 3% (energy basis) co‐firing is possible. On the
other hand, indirect co‐firing enables a high degree of fuel flexibility. Moreover, combustion
of cleaned syngas does not affect the performance of the boiler (Saidur et al. 2011).
A significant improvement has been achieved with this technology with an overall electrical
efficiency of about 35% if the fuel replacement by biomass‐derived primary product is restricted
to 5–15 wt% (Molino et al. 2016). This is mainly due to the low‐grade properties of biomass
such as low bulk densities, high MC, etc. Co‐combustion of biomass and coal results in reduc-
tion of fouling and corrosion compared to single biomass combustion because the alkali met-
als present in biomass are diluted and consumed during the interaction with sulfur or silica
present in coal (Ericsson 2007). Fuel gas derived from low‐grade biomass (with higher MC
>60%) can also be co‐fired with natural gas in the indirect co‐firing approach which is another
way of utilizing bioenergy (Rodrigues et al. 2003; Fiaschi and Carta 2007). Table 6.13 presents
the co‐firing combination of various biomass feedstocks with coal and natural gas.
6.5.3 Gasifier IC Engine Combined Technology

Producer gas derived from biomass gasification coupled with an internal combustion (IC)
engine is a viable technology for small‐scale heat and power generation (Martínez et al.
(a) (b) (c)

To steam turbine To steam turbine To steam turbine
Biomass
Biomass Syngas
Gasifier
Biomass Coal
Coal Coal
Boiler Boiler Boiler Boiler
Figure 6.4 Co-firing of coal and biomass. (a) Direct co-firing. (b) Indirect co-firing. (c) Parallel
co-firing.
Table 6.13 Summary of the feedstocks used in previous co-firing studies.
Type of biomass feedstock Coal or natural gas
Wheat straw, sewage sludge, wood chip, and WPOS (woody matter Federal and Bellambi coals
from olive stones)
Foot cake (waste from the olive oil industry) Coal (lignite and anthracite)
Sludge and hog fuel Subbituminous coal
Paper mill sludge Coal
Gasified sugarcane residues Natural gas
Cellulose biomass and nonhazardous waste (institutional/ Natural gas
household waste with plastics and food‐related paper components)
Source: Adapted from Zhang et al. (2010) with permission from Elsevier.
2012). Producer gas can be processed in both a spark ignition engine as a single fuel mode
and a compression ignition (CI) engine as a dual fuel mode. The gasifier combined IC
engine system can deliver an overall electrical energy conversion in the range of 35–45%
(Yassin et al. 2009). A schematic diagram of a gasifier system coupled with an engine is
shown in Figure 6.5. The complete conversion technology comprises a biomass feeding
mechanism, gasifier, gas cooling and cleaning system, engine fuel injection system, engine,
and generator.
Prior to the utilization of producer gas in an engine, it is essential to optimize the operation
of the engine so that successful implementation of the system can be achieved. For the well‐
matched performance of producer gas in an IC engine, biomass with density of >300 kg/m3,
MC up to 20%, particle size of 20–50 mm must be used, and also the impurities in the pro-
ducer gas must be lowered down to acceptable levels (dust, tar, and acids <50 mg/m3) (Yaliwal
et al. 2014). Moreover, the gaseous fuel must be cleaned before it is injected into the engine.
Wet scrubbing is preferred where the gas is cooled to below 150 °C before it is passed to the
scrubbing stage. Wet scrubbing is an established cleaning technology which removes alkali
metals, ammonia, particulates, and tar. In order to retain the chemical energy of the gaseous
Biomass
Gas Tank Air Tank
Gas Mixer
Fuel Tank
Gasifier Exhaust Gas
To Loading
Engine Generator System
Filter
Cooler
Figure 6.5 Schematic diagram of gasifier system coupled with engine. Source: Adapted from
Ramadhas et al. (2008) with permission from Elsevier.
fuel, incorporation of catalytic or thermal cracking of the tar before wet scrubbing is prefer-
able (Bridgwater et al. 2002).
Producer gas has a better knocking property compared with other combustible gases
which makes it suitable to be utilized in an IC engine. This is because the presence of H2 in
producer gas increases the laminar flame velocity which reduces the knocking tendency.
Another reason for lower knocking is the suppression of preflame reaction due to the pres-
ence of higher fractions of inactive gases such as CO2 (12–15%) and N2 (48–50%) in producer
gas (Sridhar et al. 2005). A host of researchers have found that application of producer gas
in a CI engine under dual fuel mode has a significant effect on the performance, combus-
tion, and emission characteristics of the engine. Under dual fuel mode, the producer gas
(derived from biomass) is used as primary fuel and diesel is used as the pilot liquid fuel,
thereby replacing diesel with a renewable energy source. A dual fuel engine shows better
performance along with lower smoke and NOx emissions at higher load in comparison
with a diesel engine (Yaliwal et al. 2014).
The major challenge with this technology is that current engines are designed for either
petrol or diesel due to which utilization of producer gas results in derating of the system.
Further modification of the injection system can improve the engine performance.
Moreover, producer gas can also be mixed with biogas produced from anaerobic digestion.
6.5.4 Gasifier-Gas Turbine Combined Technology

The gas turbine is known for its high efficiency and low emissions, simple operation and
reliability with a low specific capital cost. Gas turbine technology is one of the known path-
ways of biomass gasification power generation which gives an electrical efficiency of about
40%, higher than a steam power cycle. This technology is mostly utilized for large‐scale
power generation units. Upstream limitations of this technology include feeding prepared
biomass into the gasifier and the downstream process includes supply of filtered low heat-
ing value producer gas to the gas turbine fuel injection system. Components of this technol-
ogy include the biomass feeding mechanism, gasifier, moderate gas cooling, cleaning of hot
gas and the gas turbine.
Supply of clean fuel gas to the turbine is the major issue in this technology in order to
avoid rapid deterioration of turbine blades as gas turbines are highly sensitive to the quality
of fuel gas. Therefore, contaminants present in the gas must be removed. In this system, the
temperature of the fuel gas need not to be cooled down to atmospheric temperature but it
is cooled moderately to a temperature of 500 °C to remove particulate matter and to
condense alkali metals. The fuel gas is delivered at a temperature of 450 °C which is rela-
tively high and allows the tar present in the gas to be in the vapor state which can be burned
along with the fuel gas in the turbine. The quality requirements of producer gas for this
technology are calorific >4 MJ/Nm3, tar = 10 mg/Nm3, metals = 0.025–0.1 ppmw,
particulates = 2.4 mg/Nm3 and H2S = 20 ppmv (Bridgwater et al. 2002; Molino et al. 2016).
6.5.5 Biomass Integrated Gasification Combined Cycle

Gas turbine combustion efficiency can be enhanced through a biomass integrated gasifica-
tion combined cycle (BIGCC) in which the combustor of the gas turbine cycle is replaced
by the gasifier and in addition, the exhaust heat of the gas turbine is utilized for producing
steam to run the conventional steam turbine. In this technology, gas and steam turbines
operate in combination which holds the promise of clean, efficient, and cost‐effective
technology for power production from biomass (Krigmont 2001; Balat et al. 2009a). The
BIGCC comprises three processes: gasification process, gas power cycle, and steam power
cycle. A schematic diagram of the BIGCC is shown in Figure 6.6.
The gasification processes have already been discussed in previous sections. In the later
stage, the hot and clean fuel gas from the gasifier enters the gas turbine fuel injection sys-
tem and burns to produce electricity. Exhaust heat coming from the gas turbine is pro-
cessed through a heat recovery system to produce superheated steam which is expanded
in the steam turbine, resulting in combined cycle power generation without using any
additional fuel (Krigmont 2001). There are several issues to be considered for running a
gas turbine with a low heating value gaseous fuel including the optimum mixing of air
and fuel to achieve the correct temperature at the inlet of the turbine, operation and con-
trol of the gas turbine compressor, limitations of emission, limitations of fuel injection
system and also possible redesign of early stages of the gas turbine in order to meet the
increased flow rate which will rise due to firing of gas with low heating value (Bridgwater
et al. 2002).
Syngas
Gas Cleanup
Biomass
Air/oxygen Gasifier
Clean Fuel Gas
Combustor
Steam
To Gasifier
Ash
Gas
Compressor Turbine
Air
Water
Exhaust Heat
Chimney
Recovery Unit
Steam Steam
Turbine
Flue Gas
Water
To Gasifier
Figure 6.6 Schematic diagram of a biomass integrated gasification combined cycle (BIGCC).
The BIGCC has several advantages when considering operational, economic, and social
concerns. It shows excellent environmental performance in terms of removal of NOx, SO2,
and particulate matter. Sulfur present in the coal is removed to 99%, NOx reduced to about
90–35% and CO2 reduction is achieved. Another attractive feature of BIGCC power genera-
tion is that it allows the application of various types of fuel such as coal, natural gas, oil, etc.
This feature enables the plant to run even during unavailability or disruption of fuel supply,
as the plant can be switched from coal to natural gas or even to oil. This technology also
allows repowering of already existing fossil fuel‐based plants which are low efficiency and
highly polluting. BIGCC plants allow integration of additional stages in blocks ranging
from 100 to 450 MW. An efficiency of about 42–52% could be achieved with this technology
in the near future which would be higher compared to coal‐based plants. Furthermore, the
amount of water required for the BIGCC is only 50–70% that of a pulverized coal‐based
plant. Application of the BIGCC could provide a new direction for the problems associated
with other wood combustion technologies and also holds promise for the effective utiliza-
tion of waste woody biomass at lower cost. Combination of a BIGCC with a fuel cell is
another future technology which could enable further reductions of CO2 emissions per
unit of electricity (Krigmont 2001).
6.5.6 High-Temperature Fuel Cell

A fuel cell is an electrochemical device which converts the chemical energy of fuel into
electricity along with heat and water vapor. Fuel cell technology is a one‐step process for
energy conversion with much higher efficiency than other multistage thermomechanical
technologies (chemical‐thermal‐mechanical).
A fuel cell encompasses three main components: a catalytic fuel electrode (anode), a
catalytic oxygen electrode (cathode), and an electrolyte which restricts the electronic con-
tacts of the electrodes (Ud Din and Zainal 2016). The high‐temperature fuel cell is another
efficient way of utilizing combustible gas produced from gasification. Modern high‐
temperature fuel cells can be operated with hydrogen, carbon monoxide, and methane and
therefore integration of a biomass gasifier could be a suitable option for power generation.
Moreover, the application of producer gas in this type of cell can also reduce the combus-
tion process which relatively decreases the environmental impact. The main advantage
with this technology is that more 40% electrical conversion efficiency can be obtained.
There are two types of fuel cells available for this conversion: solid oxide fuel cell (SOFC)
and molten oxide carbonate fuel cell (MCFC). One advantage of these fuel cells is that
they can be made of CO or CO2 in addition to hydrogen (H2) and methane (CH4) (Molino
et al. 2016).
Chemical energy of gasifier‐based fuel gas can be directly converted to electrical energy
by MCFCs at constant temperature through electrochemical reactions using an alkaline
solution as electrolyte which mainly consists of potassium and lithium. This fuel cell oper-
ates at a temperature range of 600–650 °C and pressure of 3.5–4 bar. Higher operating
temperatures help to decrease the presence of noble metals as catalyst which reduces the
cost. MCFCs can be processed with other cells which requires shifting of CO to H2.
Moreover, as the methane steam reforming is done on site, there is no poisoning problem
due to catalysts which reduces the fuel cleaning operation simultaneously. Due to low
operating temperatures compared to SOFCs, this type of fuel cell can be considered to be
the most promising fuel cell which can directly convert producer gas at higher efficiency.
But MCFCs demand purification of producer gas up to a certain level in order to prevent
material deterioration from tar or particulate matter (Iaquaniello and Mangiapane 2006;
Molino et al. 2019).
On the other hand, in SOFCs, solid oxide ceramic electrolytes are used which are made
of zirconium oxide (ZrO2) stabilized with yttrium oxide (Y2O3) and are considered to have
immense potential for being utilized in a combined cycle. SOFCs works at a temperature
range of 900–1000 °C. This higher temperature is required due to ionic conductivity values
and this in turn abolishes the evaporation and corrosion possibilities of solid components,
which are more prevalent for the cells that work on liquid electrolytes. Internal reforming
of fuel gas is possible in SOFCs because of their high temperature operation which allows
reforming of hydrocarbons directly in the anode chamber. Therefore, handling of a wide
range of hydrocarbons is possible for this type. Hence, suitability for higher temperature,
high efficiency, solid‐state design, and ability to reform fuel gas internally make this type
more promising than MCFCs (Kempegowda et al. 2012; Molino et al. 2019).
From the above description, it is obvious that due to the ability to perform direct reform-
ing of hydrocarbon without any limit to CO and CO2 content, MCFCs and SOFCs are the
most reliable and promising option for producing electric power from renewable sources.
6.6 Conclusion
Lignocellulosic biomass is regarded as a sustainable and reliable renewable energy source

which has the potential to mitigate the dual challenge of diminishing fossil fuel and green-
house gas emission. Moreover, biomass‐derived fuels can also fulfill the growing energy
demand of the modern era of industrialization and urbanization. This chapter covers all
aspects of lignocellulosic biomass conversion processes, including types of feedstocks, charac-
terization, physiochemical properties, pretreatment process and especially the thermochemi-
cal conversion technologies and also discusses their applications for electricity generation.
Characterization of biomass feedstock is crucial for selecting the proper conversion
route. It also determines the applicability of a feedstock for energy use. Understanding of
physiochemical properties of lignocellulosic biomass plays a significant role in terms of
design and implementation of any conversion technology and it also helps in optimizing
various operating parameters. Pretreatment of lignocelluloses improves the performance
of conversion processes and also enables high yield with desired properties of products.
Lignocellulosic biomass can be transformed to useable fuel through numerous conversion
processes. Selection of these processes depends on many factors such as feedstock availabil-
ity and type, end‐use of derived products, economic aspects, environmental standards, etc.
Direct combustion is the simplest mode of biomass conversion to heat and this heat can be
further utilized for electricity generation through a steam cycle. Biomass can also be co‐fired
directly or indirectly with coal or natural gas in order to increase the heating value of com-
bustible gas which can simultaneously enhance the overall performance of the system.
Pyrolysis and liquefaction are two thermochemical processes through which liquid fuel
can be generated from biomass. In comparison with pyrolysis, liquefaction process is
c onsidered to be more expensive and also produces highly viscous bio‐oil compared to fast
pyrolysis. Depending upon the operating temperature, heating rate and residence time,
pyrolysis can deliver three different forms of fuel. Slow pyrolysis is preferable for the pro-
duction of char and fast and flash pyrolysis are considered to be more attractive options for
generating liquid fuel with higher production yield. Bio‐oil generated from pyrolysis has
comparative chemical characteristics to those of conventional petroleum oil and can be
used as an alternative for engine application.
Biomass gasification can be considered as the most multipurpose thermochemical conver-
sion method as the derived gaseous fuel, producer gas, can be utilized in many directions. Based
on operation principles, gasifiers are classified in three types. Fixed‐bed gasifiers are generally
used for small‐scale operations whereas fluidized‐bed and entrained‐bed gasifiers are suitable
for large‐scale application. Producer gas can directly be injected into an IC engine or can be
utilized in a CI engine with dual fuel mode for generating power. Moreover, electricity can be
generated through a BIGCC where an additional steam cycle can be operated with a gas turbine
exhaust heat recovery unit. Gasification can also be combined with fuel cell technology in order
to enhance electrical efficiency as compared to engine or gas turbine technologies.
Utilization of lignocellulosic biomass for energy generation has several socioeconomic
advantages such as creation of market value for biomass, local skill development and job
opportunities, rural development through environmentally friendly technologies, proper utili-
zation and control of local resources, etc. Proper utilization of biomass through various con-
version processes can lead to production of high‐quality fuel which could mitigate dependence
on fossil fuel and also could help in improving the economic structure of developing nations.
Acknowledgment
The authors gratefully acknowledge the financial support extended by the Science and
Engineering Research Board, Government of India (Grant number: ECR/2016/001830),
and the Centre for Energy, IIT Guwahati, for providing all research facilities.
R
eference
Agbor, V.B., Cicek, N., Sparling, R. et al. (2011). Biomass pretreatment: fundamentals toward
application. Biotechnology Advances 29 (6): 675–685.
Al Arni, S., Bosio, B., and Arato, E. (2010). Syngas from sugarcane pyrolysis: an experimental
study for fuel cell applications. Renewable Energy 35 (1): 29–35.
Ali, G., Nitivattananon, V., Abbas, S., and Sabir, M. (2012). Green waste to biogas: renewable
energy possibilities for Thailands green markets. Renewable and Sustainable Energy Reviews
16 (7): 5423–5429.
Ariunbaatar, J., Panico, A., Esposito, G. et al. (2014). Pretreatment methods to enhance
anaerobic digestion of organic solid waste. Applied Energy 123: 143–156.
Balat, M., Balat, M., Kirtay, E., and Balat, H. (2009a). Main routes for the thermo‐conversion of
biomass into fuels and chemicals. Part 1: pyrolysis systems. Energy Conversion and
Management 50 (12): 3147–3157.
Balat, M., Balat, M., Kirtay, E., and Balat, H. (2009b). Main routes for the thermo‐conversion of
biomass into fuels and chemicals. Part 2: gasification systems. Energy Conversion and
Management 50 (12): 3158–3168.
Behera, S., Arora, R., Nandhagopal, N., and Kumar, S. (2014). Importance of chemical
pretreatment for bioconversion of lignocellulosic biomass. Renewable and Sustainable
Bilgen, S., Kaygusuz, K., and Sari, A. (2004). Renewable energy for a clean and sustainable
future. Energy Sources 26 (12): 1119–1129.
Brandt, A., Gräsvik, J., Hallett, J.P., and Welton, T. (2013). Deconstruction of lignocellulosic
biomass with ionic liquids. Green Chemistry 15 (3): 550–583.
Bridgwater, A.V., Toft, A.J., and Brammer, J.G. (2002). A techno‐economic comparison of
power production by biomass fast pyrolysis with gasification and combustion. Renewable
and Sustainable Energy Reviews 6 (3): 181–248.
British Petroleum (2018). BP Statistical Review of World Energy. www.bp.com/en/global/
corporate/energy‐economics/statistical‐review‐of‐world‐energy.html.
Cai, J., He, Y., Yu, X. et al. (2017). Review of physicochemical properties and analytical
characterization of lignocellulosic biomass. Renewable and Sustainable Energy Reviews 76:
309–322.
Capareda, S.C. (2014). Introduction to Biomass Energy Conversions. New York: CRC Press.
Channiwala, S.A. and Parikh, P.P. (2002). A unified correlation for estimating HHV of solid,
liquid and gaseous fuels. Fuel 81 (8): 1051–1063.
Chen, H., Liu, J., Chang, X. et al. (2017). A review on the pretreatment of lignocellulose for
high‐value chemicals. Fuel Processing Technology 160: 196–206.
Cotana, F., Cavalaglio, G., Petrozzi, A., and Coccia, V. (2015). Lignocellulosic biomass feeding
in biogas pathway: state of the art and plant layouts. Energy Procedia 81: 1231–1237.
Damartzis, T. and Zabaniotou, A. (2011). Thermochemical conversion of biomass to second
generation biofuels through integrated process design‐a review. Renewable and Sustainable
Energy Reviews 15 (1): 366–378.
de Bhowmick, G., Sarmah, A.K., and Sen, R. (2017). Lignocellulosic biorefinery as a model for
sustainable development of biofuels and value added products. Bioresource Technology 247
(June): 1144–1154.
Demirbas, A. (1997). Calculation of higher heating values of biomass fuels. Fuel 76: 431–434.
Demirbas, A. (2001). Biomass resource facilities and biomass conversion processing for fuels
and chemicals. Energy Conversion and Management 42 (11): 1357–1378.
Demirbas, A. (2004). Combustion characteristics of different biomass fuels. Progress in Energy
and Combustion Science 30 (2): 219–230.
Dhyani, V. and Bhaskar, T. (2018). A comprehensive review on the pyrolysis of lignocellulosic
biomass. Renewable Energy 129: 695–716.
Dupont, C., Chiriac, R., Gauthier, G., and Toche, F. (2014). Heat capacity measurements of
various biomass types and pyrolysis residues. Fuel 115: 644–651.
Effendi, A., Gerhauser, H., and Bridgwater, A.V. (2008). Production of renewable phenolic
resins by thermochemical conversion of biomass: a review. Renewable and Sustainable
Energy Reviews 12 (8): 2092–2116.
Elbersen, W., Lammens, T.M., Alakangas, E.A. et al. (2017). Lignocellulosic biomass quality:
matching characteristics with biomass conversion requirements. In: Modeling and
Optimization of Biomass Supply Chains (ed. C. Panoutsou), 55–78. London: Academic

Press.
Ericsson, K. (2007). Co‐firing‐a strategy for bioenergy in Poland? Energy 32 (10): 1838–1847.
Feng, Q. and Lin, Y. (2017). Integrated processes of anaerobic digestion and pyrolysis for
higher bioenergy recovery from lignocellulosic biomass: a brief review. Renewable and
Fernandes, T.V., Klaasse Bos, G.J., Zeeman, G. et al. (2009). Effects of thermo‐chemical pre‐
treatment on anaerobic biodegradability and hydrolysis of lignocellulosic biomass.
Fiaschi, D. and Carta, R. (2007). CO2 abatement by co‐firing of natural gas and biomass‐
derived gas in a gas turbine. Energy 32 (4): 549–567.
Gera, D., Mathur, M.P., Freeman, M.C., and Robinson, A. (2002). Effect of large aspect ratio of
biomass particles on carbon burnout in a utility boiler. Energy and Fuels 16 (6): 1523–1532.
Goyal, H.B., Seal, D., and Saxena, R.C. (2008). Bio‐fuels from thermochemical conversion of
renewable resources: a review. Renewable and Sustainable Energy Reviews 12 (2): 504–517.
Hamelinck, C.N. and Faaij, A.P.C. (2006). Outlook for advanced biofuels. Energy Policy 34 (17):
3268–3283.
Hassan, S.S., Williams, G.A., and Jaiswal, A.K. (2018). Emerging technologies for the
pretreatment of lignocellulosic biomass. Bioresource Technology 262: 310–318.
Iaquaniello, G. and Mangiapane, A. (2006). Integration of biomass gasification with MCFC.
International Journal of Hydrogen Energy 31 (3): 399–404.
Ibarra‐Gonzalez, P. and Rong, B.‐G. (2018). A review of the current state of biofuels production
from lignocellulosic biomass using thermochemical conversion routes. Chinese Journal of
Chemical Engineering https://doi.org/10.1016/j.cjche.2018.09.018.
Jayah, T.H., Aye, L., Fuller, R.J., and Stewart, D.F. (2003). Computer simulation of a downdraft
wood gasifier for tea drying. Biomass and Bioenergy 25 (4): 459–469.
Kaur, R.R. and Luthra, A. (2018). Population growth, urbanization and electricity – challenges
and initiatives in the state of Punjab, India. Energy Strategy Reviews 21: 50–61.
Kempegowda, R.S., Skreiberg, O., and Tran, K.Q. (2012). Cost modeling approach and
economic analysis of biomass gasification integrated solid oxide fuel cell systems. Journal of
Renewable and Sustainable Energy 4: 043109.
Kobayashi, N. and Fan, L.S. (2011). Biomass direct chemical looping process: a perspective.
Biomass and Bioenergy 35 (3): 1252–1262.
Krigmont, H.V. (2001). Integrated Biomass Gasification Combined Cycle (IBGCC) Power
Generation Concept: The Gatway to A Cleaner Future. www.semanticscholar.org/paper/
INTEGRATED‐BIOMASS‐GASIFICATION‐COMBINED‐CYCLE‐(‐)‐Krigmont/6c43b7dbbf
c0b861b939f58f2d17adb1bddad4b1.
Kumari, D. and Singh, R. (2018). Pretreatment of lignocellulosic wastes for biofuel production:
a critical review. Renewable and Sustainable Energy Reviews 90: 877–891.
Lee, U., Balu, E., and Chung, J.N. (2013). An experimental evaluation of an integrated biomass
gasification and power generation system for distributed power applications. Applied Energy
101: 699–708.
Lin, G., Yang, H., Wang, X. et al. (2016). The moisture sorption characteristics and modelling
of agricultural biomass. Biosystems Engineering 150: 191–200.
Loha, C., Karmakar, M.K., De, S., and Chatterjee, P.K. (2018). Gasifier: types, operational
principles and commercial forms. In: Coal and Biomass Gasification: Recent Advances and
Future Challenges (eds. S. De, A.K. Agarwal, V.S. Moholkar and B. Thallada), 63–91.
Singapore: Springer Nature.
Lumay, G., Boschini, F., Traina, K. et al. (2012). Measuring the flowing properties of powders
and grains. Powder Technology 224: 19–27.
Mahmoud, A., Shuhaimi, M., and Abdel Samed, M. (2009). A combined process integration
and fuel switching strategy for emissions reduction in chemical process plants. Energy 34
(2): 190–195.
Martínez, J.D., Mahkamov, K., Andrade, R.V., and Silva Lora, E.E. (2012). Syngas production in
downdraft biomass gasifiers and its application using internal combustion engines.
Renewable Energy 38 (1): 1–9.
McKendry, P. (2002a). Energy production from biomass (part 1): overview of biomass.
Bioresource Technology 83: 37–46.
McKendry, P. (2002b). Energy production from biomass (part 2): conversion technologies.
Miao, Z., Grift, T.E., Hansen, A.C., and Ting, K.C. (2014). Flow performance of ground biomass
in a commercial auger. Powder Technology 267: 354–361.
Miccio, F., Silvestri, N., Barletta, D., and Poletto, M. (2011). Characterization of woody biomass
flowability. Chemical Engineering Transactions 24: 643–648.
Molino, A., Chianese, S., and Musmarra, D. (2016). Biomass gasification technology: the state
of the art overview. Journal of Energy Chemistry 25: 10–25.
Molino, A., Migliori, M., Larocca, V. et al. (2019). Power production by biomass gasification
technologies. In: Current Trends and Future Developments on (Bio‐) Membranes (eds. A.
Basile, A. Cassano and A. Figoli), 293–318. Amsterdam: Elsevier.
Naik, S.N., Goud, V.V., Rout, P.K., and Dalai, A.K. (2010). Production of first and second
generation biofuels: a comprehensive review. Renewable and Sustainable Energy Reviews 14
(2): 578–597.
Nunes, L.J.R., Matias, J.C.O., and Catalão, J.P.S. (2016). Biomass combustion systems: a review
on the physical and chemical properties of the ashes. Renewable and Sustainable Energy
Reviews 53: 235–242.
Onumaegbu, C., Mooney, J., Alaswad, A., and Olabi, A.G. (2018). Pre‐treatment methods for
production of biofuel from microalgae biomass. Renewable and Sustainable Energy Reviews
93 (May): 16–26.
Panwar, N.L., Kothari, R., and Tyagi, V.V. (2012). Thermo chemical conversion of biomass – eco
friendly energy routes. Renewable and Sustainable Energy Reviews 16 (4): 1801–1816.
Patel, M., Zhang, X., and Kumar, A. (2016). Techno‐economic and life cycle assessment on
lignocellulosic biomass thermochemical conversion technologies: a review. Renewable and
Paudel, S.R., Banjara, S.P., Choi, O.K. et al. (2017). Pretreatment of agricultural biomass for
anaerobic digestion: current state and challenges. Bioresource Technology 245 (September):
1194–1205.
Paul, S. and Dutta, A. (2018). Challenges and opportunities of lignocellulosic biomass for
anaerobic digestion. Resources, Conservation and Recycling 130: 164–174.
Rabemanolontsoa, H. and Saka, S. (2016). Various pretreatments of lignocellulosics.

Ramadhas, A.S., Jayaraj, S., and Muraleedharan, C. (2008). Dual fuel mode operation in diesel
engines using renewable fuels: rubber seed oil and coir‐pith producer gas. Renewable Energy
33 (9): 2077–2083.
Raveendran, K., Ganesh, A., and Khilar, K.C. (1995). Influence of mineral matter on biomass
pyrolysis characteristics. Fuel 74 (12): 1812–1822.
Reddy, B.R. and Vinu, R. (2018). Feedstock characterization for pyrolysis and gasification. In:
Coal and Biomass Gasification: Recent Advances and Future Challenges (eds. S. De, A.K.
Agarwal, V.S. Moholkar and B. Thallada), 3–36. Singapore: Springer Nature.
Rodrigues, M., Walter, A., and Faaij, A. (2003). Co‐firing of natural gas and biomass gas in
biomass integrated gasification/combined cycle systems. Energy 28 (11): 1115–1131.
Rowlands, W.N., Master, A., and Maschmeyer, T. (2008). The biorefinery – challenges,
opportunities, and an Australian perspective. Bulletin of Science, Technology & Society 28 (2):
149–158.
Saidur, R., Abdelaziz, E.A., Demirbas, A. et al. (2011). A review on biomass as a fuel for boilers.
Renewable and Sustainable Energy Reviews 15 (5): 2262–2289.
Sawatdeenarunat, C., Surendra, K.C., Takara, D. et al. (2015). Anaerobic digestion of
lignocellulosic biomass: challenges and opportunities. Bioresource Technology 178: 178–186.
Sellin, N., Krohl, D.R., Marangoni, C., and Souza, O. (2016). Oxidative fast pyrolysis of banana
leaves in fluidized bed reactor. Renewable Energy 96: 56–64.
Sindhu, R., Binod, P., and Pandey, A. (2016). Biological pretreatment of lignocellulosic
biomass – an overview. Bioresource Technology 199: 76–82.
Song, C., Hu, H., Zhu, S. et al. (2004). Nonisothermal catalytic liquefaction of corn stalk in
subcritical and supercritical water. Energy and Fuels 18 (1): 90–96.
Sridhar, G., Sridhar, H.V., Dasappa, S. et al. (2005). Development of producer gas engines.
Proceedings of the Institution of Mechanical Engineers, Part D: Journal of Automobile
Engineering 219 (3): 423–438.
SuÁrez, J.A., Luengo, C.A., Felfli, F.F. et al. (2002). Thermochemical properties of cuban
biomass. Energy Sources 22 (10): 851–857.
Ud Din, Z. and Zainal, Z.A. (2016). Biomass integrated gasification‐SOFC systems: technology
overview. Renewable and Sustainable Energy Reviews 53: 1356–1376.
Wang, S., Dai, G., Yang, H., and Luo, Z. (2017). Lignocellulosic biomass pyrolysis mechanism:
a state‐of‐the‐art review. Progress in Energy and Combustion Science 62: 33–86.
Xu, R., Ferrante, L., Briens, C., and Berruti, F. (2009). Flash pyrolysis of grape residues into
biofuel in a bubbling fluid bed. Journal of Analytical and Applied Pyrolysis 86 (1): 58–65.
Yaliwal, V.S., Banapurmath, N.R., Gireesh, N.M., and Tewari, P.G. (2014). Production and
utilization of renewable and sustainable gaseous fuel for power generation applications: a
review of literature. Renewable and Sustainable Energy Reviews 34: 608–627.
Yassin, L., Lettieri, P., Simons, S.J.R., and Germanà, A. (2009). Techno‐economic performance
of energy‐from‐waste fluidized bed combustion and gasification processes in the UK
context. Chemical Engineering Journal 146 (3): 315–327.
Yin, C.‐Y. (2011). Prediction of higher heating values of biomass from proximate and ultimate
analyses. Fuel 90: 1128–1132.
Zabed, H., Sahu, J.N., Suely, A. et al. (2017). Bioethanol production from renewable sources:
current perspectives and technological progress. Renewable and Sustainable Energy Reviews
71: 475–501.
Zhang, L., Charles, C., and Champagne, P. (2010). Overview of recent advances in thermo‐
chemical conversion of biomass. Energy Conversion and Management 51: 969–982.
Zhang, K., Chang, J., Guan, Y. et al. (2013). Lignocellulosic biomass gasification technology in
China. Renewable Energy 49: 175–184.
Zhao, X., Liu, W., Deng, Y., and Zhu, J.Y. (2017). Low‐temperature microbial and direct
conversion of lignocellulosic biomass to electricity: advances and challenges. Renewable and
Zheng, Y., Zhao, J., Xu, F., and Li, Y. (2014). Pretreatment of lignocellulosic biomass for
enhanced biogas production. Progress in Energy and Combustion Science 42: 35–53.
Zhu, J.Y. and Pan, X.J. (2010). Woody biomass pretreatment for cellulosic ethanol production:
technology and energy consumption evaluation. Bioresource Technology 101 (13): 4992–5002.
125
Biopolymers from Lignocellulosic Biomass

Feedstocks, Production Processes, and Applications
Grazielle Machado1, Fernando Santos2, Rogério Lourega1, Jaqueline Mattia2,
Douglas Faria1, Paulo Eichler1, and Angenor Auler2
1
Polytechnic School, Pontifical Catholic University of Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil
2
Study Center in Biorefinery, State University of Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil
7.1 Introduction
In the current scenario, there is a great demand for plastics manufacturing since these
materials are used in a variety of applications, especially for packaging, leading to the
generation of enormous amounts of waste. Most of the plastics used in commercial appli-
cations are derived from petroleum, which is a nonrenewable resource. Besides that, after
their use, usually only a small fraction is recycled, while the largest fraction of plastic resi-
dues is disposed of, often improperly, leading to serious environmental hazards (Osman
et al. 2016; Emadian et al. 2017). In this context, an emerging trend is the conversion of
lignocellulosic biomass into high value‐added products, including biopolymers. Research
involving new materials development has been growing in recent years, focusing mainly
on high‐performance eco‐friendly materials at affordable costs, such as biopolymers
(Satyanarayana et al. 2009; Grewal and Khare 2018).
Biopolymers are polymers synthesized by living organisms, such as plants, fungi, bacte-
ria, and archaea, which are a promising alternative to replace conventional plastics. The
advantages of employing biopolymers include that they are biodegradable, reducing accu-
mulation of waste in the environment, while their biodegradation normally leads to the
formation of less toxic products. In addition, in most cases, smaller amounts of pollutants
are generated during their production process, and they can be produced from waste or
by‐products (Fasciotti 2017). There are many biopolymers that can be obtained from ligno-
cellulosic biomass; some examples are methylcellulose, carboxymethylcellulose, cellulose,
cellulose acetate, polylactic acid (PLA), acrylic acid, polyhydroxybutyrate (PHB), polyeth-
ylene (PE), furfural resins, and phenolic resins (Boneberg et al. 2016).
This chapter is focused on biopolymers obtained from lignocellulosic biomass. In addi-
tion, other important topics including types of biopolymers, biodegradability, production

processes, feedstocks, properties, applications, biocomposites, and the current scenario in

biopolymer commercial production are discussed.
7.2 Biopolymers in General
Polymers are macromolecules constituted of small and simple repeating structural units
called monomers. There are a few different definitions postulated by acknowledged organi-
zations regarding biopolymers. According to the International Union of Pure and Applied
Chemistry( IUPAC), a biopolymer is a polymer produced by living organisms, including
proteins, polysaccharides, and nucleic acids (DNA and RNA). Biopolymers are the “build-
ing blocks of life,” that is, they are the most important components of cell infrastructure
and play essential roles in cell regulation and replication. The three main classes of biopol-
ymers synthesized by living organisms are proteins (formed by amino acids), polysaccha-
rides (sugar polymeric chains), and nucleic acids (formed by nucleotides) (McNaught and
Wilkinson 1997).
Another important definition is regarding biodegradable polymers which, according to
the American Society for Testing and Materials (ASTM) D883 regarding Standard
Terminology Relating to Plastics (2019), are “a degradable plastic in which the degradation
results from the action of naturally occurring microorganisms such as bacteria, fungi, and
algae.” In this context, biodegradable polymers are materials whose degradation in natural
processes leads to their conversion in CO2, H2O, and nontoxic inorganic compounds (ASTM
2019). Regarding this standard, there are two categories of degradable polymers: polymers
which are degraded by microorganisms, so‐called biodegradable polymers, and oxo‐degra-
dable polymers, which contain inorganic additives that activate polymer degradation
initiated by oxygen (Boneberg et al. 2016).
Moreover, the European Bioplastics Organization classifies bioplastics (biopolymers)
in two different groups: bio‐based and biodegradable polymers. According to this defini-
tion, bio‐based or partially bio‐based bioplastics are polymers that possess the same
properties as conventional forms; that is, they are technically equivalent to oil‐based
plastics but are produced from renewable resources. Whereas biodegradable plastics are
polymers derived from either renewable or nonrenewable resources, which are certified
as compostable according to international standards, such as EN13432. Figure 7.1 shows
some examples of biodegradable, bio‐based, and both biodegradable and bio‐based
bioplastics (E‐Bioplastics 2018).
7.3 Biodegradability
Biodegradation of polymers generally occurs through various processes involving biologi-
cal, chemical, and mechanical activities which lead to a modification in the polymer chem-
ical structure, resulting in the formation of simple metabolites, as illustrated in Figure 7.2
(Boneberg et al. 2016).
Biopolymers from Lignocellulosic Biomass 127
Bioplastics
Biodegradable
plastics
PES PE
PHB
PCL PLA NY II
PBS Starch AcC
Bio-based
plastics
Figure 7.1 Bioplastics including biodegradable and bio-based plastic. AcC, acetylcellulose; NY II,
nylon II; PBS, polybutylene succinate; PCL, polycaprolactone; PE, polyethylene; PES, polyethylene
succinate; PHB, polyhydroxybutryrate; PLA, polylactic acid. Source: Adapted from Boneberg et al.
(2016); open access article, copyright permisison provided under Creative Commons Attribution 4.0
International License.
Mechanical
strain H2O/O2 Microorganisms
Biomass
Polymer + CO2 + H2O
volatiles
Sunlight Hydrolysis/ Biodegradation
oxidation
Figure 7.2 Polymer biodegradation involving mechanical, chemical, and biological processes.
Source: Adapted from Boneberg et al. (2016); open access article, copyright permisison provided
under Creative Commons Attribution 4.0 International License.
There are three main steps involved in polymer biodegradation: biodeterioration, bio-
fragmentation, and assimilation. Biodeterioration is the modification of mechanical,
chemical, and physical properties of the polymer due to microbial growth on the surface
or inside the polymer. Biofragmentation is the conversion of polymers into oligomers
and monomers by microbial metabolism. Assimilation is the final step in which micro-
organisms, supplied by carbon, energy and nutrient sources from the fragmentation
step, are able to convert these components to carbon dioxide, water, and biomass
(Emadian et al. 2017).
Factors that influence polymer biodegradation include chemical structure, polymer
chain, crystallinity, and complexity of polymeric formula. The degradation carried out by
microorganisms involves enzymatic reactions, so that a specific enzyme is required for
each functional group in the polymeric structure. In this context, polymers which have a
shorter chain, more amorphous regions and a less complex formula are easy to degrade by
microorganisms. Also, environmental conditions play a significant role in biodegradation,
including pH, temperature, moisture, and oxygen content (Emadian et al. 2017).
7.4 Types of Biopolymer
There are four main types of biopolymer that can be obtained from lignocellulosic biomass:
(i) natural biopolymers (e.g., cellulose, hemicellulose, lignin); (ii) biopolymers obtained
through modification of naturally occurring polymers (e.g., nanocellulose and carboxym-
ethylcelullose); (iii) biopolymers synthesized by microorganisms (e.g., polyhydroxyal-
kanoates); (iv) biopolymers obtained through polymerization of biomonomers derived
from lignocellulosic feedstock (e.g., PLA) (Satyanarayana et al. 2009). Figure 7.3 shows
examples of some biopolymers and Figure 7.4 shows the routes used to obtain all these
biopolymers from lignocellulosic feedstock (Brodin et al. 2017).
7.4.1 Natural Biopolymers

Cellulose, hemicelluloses, and lignin are the most abundant natural biopolymers, since
they are the compounds present in the highest amount in lignocellulosic biomass. Cellulose
(Figure 7.5) is a linear homopolysaccharide composed of β‐cellobiose repeating units,
which consist of two β‐d‐glucopyranose monomers connected by a β‐1,4‐glycosidic bond.
In this polymer, the linearity of polymeric chains allows neighbor molecules to connect by
hydrogen and van der Waals interactions, which provide high strength and high crystallin-
ity to the polymer. Hemicelluloses (Figure 7.6) are a group of complex heteropolysaccha-
rides containing different types of sugars in their chains, including glucose, xylose,
galactose, mannose, and arabinose. These biopolymers have amorphous structures and
may exhibit substituent branches, which enable bridge formation with lignin and cellulose
fibers, leading to biomass recalcitrance, that is, the formation of a rigid structure of cellu-
lose, hemicelluloses, and lignin. Lignin (Figure 7.7) is the aromatic heteropolymer most
abundant in nature composed of the phenylpropanoid units p‐coumaryl alcohol, coniferyl
alcohol, and sinapyl alcohol, which comprise p‐hydroxyphenyl (H), guaiacyl (G), and
Biodegradable
Polymers
Renewable Chemical Microbial Biopolymers

sources synthesized synthesized blends
- Polysaccharides: - Polyacids; - PHAs; - Starch blends;

e.g. cellulose and - Poly(vinyl - Bacterial - Polyester blends;
hemicelluloses; alcohols); cellulose; - Caseinate
- Proteins: e.g. - Polyesters - Xanthan blends
casein and collagen;
- Lipids: e.g.
tryglicerides
Figure 7.3 Classification of biopolymers. PHA, polyhydroxyalkanoates. Source: Adapted from

Satyanarayana et al. (2009) with permission from Elsevier.
Chemical Biobased plastic,
modification e.g. cellulose
acetate
Biotechnological Biobased plastic,

Polysaccharide
conversion e.g. PHA
Chemical or Sugar, e.g.

enzymatic glucose,
degradation xylose
Intermediate, Monomer,
Lignocellulosic e.g. lactic Chemical e.g. ethylene,
Pretreatment Fermentation
feedstock acid, ethanol conversion lactide
Monomer or Biobased or
Lignin or Chemical Polymerization
precursor, Polymerization bioenriched plastic,
tannin conversion
e.g. phenolic e.g. PF, PUR
Biobased plastic,
Possible e.g. PE, PLA
additional
monomer
Figure 7.4 The main routes for production of biopolymers from lignocellulosic feedstock. PE, polyethylene; PF, phenol formaldehyde resin; PHA,
polyhydroxyalkanoates; PLA, polylactic acid; PUR, polyurethane. Source: Adapted from Brodin et al. (2017) with permission from Elsevier.
OH OH OH
OH HO OH
HO HO O
O O O O O O
O O O
O HO O
HO
HO OH OH OH
OH OH OH n
Figure 7.5 Cellulose structure. Source: Adapted from Machado et al. (2016); open access article,
copyright permisison provided under Creative Commons Attribution 4.0 International License.
HO2C
O
H3CO
HO Xylose
OH
O H
HO HO OH
O
O O O O
O HO O
OH
n
Figure 7.6 An example of hemicellulose structure. Source: Adapted from Machado et al. (2016);
open access article, copyright permisison provided under Creative Commons Attribution 4.0
International License.
CHO
CH
CH
H3CO OCH3
H3CO
CH2OH O
CH2OH
O CH HC CH2OH
CHOH HC CH
CI CHOH
CH2OH
CH2OH OCH3
HC
HC O OH OCH3
C O CH2OH HC O
HC
CHOH
H3CO CH2OH H3CO OCH3 CH2OH
O CH O CH
O CH2OH H3CO OCH3 CH2
HC O
O CH
H3CO OCH3 H3CO OCH3
OH CH2OH CH2OH O
OCH3 HC CH
O CH CH
H3CO H3CO OCH3

OH O
Figure 7.7 Lignin structure. Source: Adapted from Boneberg et al. (2016); open access article,
copyright permisison provided under Creative Commons Attribution 4.0 International License.
HO HO HO
OCH3 H3CO OCH3

OH OH OH
G S
H
Figure 7.8 Building blocks of lignin p-hydroxyphenyl (H), guaiacyl (G), and syringyl (S) residues.
Source: Adapted from Sen et al. (2015) with permission from the Royal Society of Chemistry.
syringyl (S) residues (Figure 7.8) in this natural polymer respectively (Faria et al. 2016;
Machado et al. 2016; Santos et al. 2017a).
7.4.2 Biopolymers Obtained through Modification of Natural Polymers

7.4.2.1 Cellulose Derivatives
Since cellulose is the most abundant renewable polysaccharide available in the world, it is
an important source for the production of different materials. Many cellulose properties
depend on its degree of polymerization, because the high number of hydroxyl groups
present on its structure allows extensive substitutions by other groups, forming cellulose
derivatives, resulting in property changes including durability, biocompatibility, crystallin-
ity, and rigidity. The presence of three hydroxyl groups in each monomer and their high
reactivity may modify properties such as hydrophobicity and chirality. Thus, hydroxyl
groups are modification facilitators allowing the obtaining of different cellulose deriva-
tives. Some examples of these derivatives are mentioned in Table 7.1 (Kanmani et al. 2017;
Shaghaleh et al. 2018).
7.4.2.2 Hemicellulose Derivatives

The investigation of hemicellulose potential in different applications has been increasing in
recent years, since these polysaccharides are biodegradable, biocompatible, and show some
interesting physicochemical properties (Peng and She 2014). Moreover, hemicelluloses are
the most complex polymers of plant cell walls, once they are formed by different sugar mon-
omers or by hydrogen interactions with cellulose, covalent bonds with lignin and ester link-
ages with acetyl units and hydroxycinnamic acids. These characteristics lead to a
heteropolymer with different properties according to the type of plant and extraction condi-
tions. Thus, through different modification reactions that will be presented in the following
sections, a variety of biopolymers derived from hemicelluloses can be produced, which can
be applied in different situations. Some examples of biopolymers derived from hemicellu-
loses and their applications/properties are presented in Table 7.2 (Sun et al. 2003).
7.4.2.3 Lignin Derivatives

Lignin can be obtained from lignocellulosic biomass using different extraction methods,
and its chemical structure is normally modified during these processes. In this sense, lignin
is not only classified based on its origin (hardwood, softwood, or grass lignin), but mostly
Table 7.1 Biopolymers derived from cellulose.
Biopolymer Application/properties References
Microcrystalline Pharmaceutical industry due to its chemical Kallakunta et al.

cellulose (MCC) inertia, high absorption, nontoxicity, and (2018); Koskela et al.
good compressibility in low pressures. Food (2018)
industry in emulsifiers, stabilizers, and
gelling agents in dairy products
Carboxymethylcellulose Food, textile, paper, and pharmaceutical Alam and
(CMC) industries due to its properties of stabilizing Christopher (2018);
emulsions and altering viscosity Jeong et al. (2018)
Nanocrystalline Foams, aerogels, building block for perm‐ Brinchi et al. (2013)
cellulose (NCC) selective membranes, adhesives, etc.
Cellulose acetate (CA) Cigarette filters, textiles, spectacle frames, Boneberg et al. (2016)
film media
Methylcellulose (MC) Food as emulsifier; cosmetics as thickener; Nasatto et al. (2015)
pharmaceuticals as additive; ceramics and
construction materials as thickener and
providing better flow properties; adhesives
Table 7.2 Biopolymers derived from hemicelluloses.
Biopolymer Application/properties References
Carboxymethyl chitosan‐ Recyclable and effective adsorbent Wu et al. (2017)

hemicellulose resin for the removal and recovery of
metal ions from waste water
Hemicelluloses‐g‐AA hydrogels Adsorption of heavy metal ions Peng and She (2014)
(hemicelluloses and acrylic Pd2+, Cd2+, and Zn2+ from aqueous
acid co‐polymer) solutions
Acetylated hemicellulose Increase of hydrophobicity, water Sun et al. (2003)
resistance, and film flexibility
Blends of hemicelluloses Food package Mendes et al. (2017)
from Caesalpina pulcherrima
and Tamarindus indica
Hemicelluloses esterified with Increase in hydrophobicity and high Farhat et al. (2017)
fatty acids moisture barrier characteristics
Etherified hemicelluloses Solubility regulation, stability Farhat et al. (2017)
against microorganisms, or to
produce film‐forming abilities
based on the extraction method used to isolate it. Based on the most used extraction meth-
ods, lignin can be classified as kraft lignin, lignosulfonate, lignin by enzymatic hydrolysis,
and organosolv lignin. These different types of lignin can be modified to produce several
types of biopolymers. Some examples of biopolymers that can be obtained by lignin
modification are shown in Table 7.3 (Agrawal et al. 2014; Naseem et al. 2016).
Table 7.3 Biopolymers derived from lignin.
Lignin classification Biopolymer Application/properties References
Lignosulfonate Polyaniline‐lignosulfonate/epoxy Coatings for corrosion protection Gupta et al. (2013)

Starch‐sodium lignosulfonate Antioxidant activity in food and cosmetics Shogren and Biswas (2013)
Kraft lignin Sulfonated kraft lignin Corrosion inhibitor for iron‐based materials Abu‐Dalo et al. (2013)
Cationic softwood Kraft lignin Flocculant for dye removal Kong et al. (2015)
Sodium‐lignosulfonate Lignopolyurethanic‐sodium lignosulfonate Flexural moduli that are significantly higher de Oliveira et al. (2015)
Organosolv lignin Phenol‐formaldehyde resol‐organosolv Lignin thermal stability increased Wang et al. (2009)
pine lignin
Carbon fibers based on organosolv Carbon fiber used as a reinforcement in Ding et al. (2016)
lignin/polyacrylonitrile blends composite application
Enzymatic hydrolysis Bio‐phenol‐hydroxymethyl furfural Molded products, coatings, and Zhang et al. (2016)
lignin adhesives
c07.indd 133 11/12/2019 6:36:55 PM

7.4.3 Biopolymers Synthesized by Microorganisms

A variety of biopolymers can be synthesized by microorganisms through fermentation pro-
cesses using lignocellulosic biomass as a substrate. Dextran, xanthan, and polyhydroxyal-
kanoates (PHAs) are some examples of these biopolymers. Xanthan gum (Figure 7.9) is an
exopolysaccharide that can be obtained by fermentation of sugars obtained from biomass,
widely used in the food, pharmaceutical, cosmetics, paint, textile, agricultural products,
and petroleum industries (Pu et al. 2018). Dextran, also an exopolysaccharide, is produced
through fermentation using lactic acid bacteria such as Leuconostoc mesenteroides,
Lactobacillus brevis, Streptococcus mutans, and Weissela confusa. This biopolymer is com-
monly used in the food, pharmaceutical, and medical industries (Rosca et al. 2018).
Another group of biopolymers obtained through fermentation are the PHAs. Their prop-
erties are very similar to those of synthetic polymers so they are an environmentally friendly
option to replace conventional plastics. PHAs are a heterogeneous group of polyesters
which can be divided into two main groups, according to the number of carbons in the
polymeric chain: short‐chain length (SCL) consists of 3–5 carbon atoms and medium‐chain
length (MCL) consists of 6–14 carbon atoms. The major difference in the polymeric chain
formation is the specificity of substrate metabolization by microorganisms (Hahn et al.
1995; Khanna and Srivastava 2005; Kumar et al. 2012; Singh et al. 2013). Figure 7.10 shows
the structural unit repeated in PHAs. SCL‐PHAs are thermoplastics presenting a high
degree of crystallinity while MCL‐PHAs are elastic polymers showing low crystallinity
degree and low melting temperature. One of the most attractive SCL‐PHAs is polyhydroxy-
butyrate (PHB), a biopolymer with crystallinity above 50%, melting temperature of 180 °C
and glass temperature of 2 °C, whose repeating unit is presented in Figure 7.10 (Kumar
et al. 2012; Singh et al. 2013).
CH2OH CH2OH
O O
OH
O
O n
O
OH OH
CH2OCCH3
O
OH
OH
COO–M+
H2 O O
C
OH
+M–OOC O O O
OHOH
C OH
H3C O
Figure 7.9 Xanthan gum structure. Source: Reprinted from Pu et al. (2018) with permission from
Elsevier.
Figure 7.10 PHA and PHB repeating units.

O O
H2 H H2 H
O C C C O C C C
R CH3
n n
PHA repeating unit PHB repeating unit
7.5 Biopolymers Obtained through Polymerization

of Biomonomers Derived from Lignocellulosic Feedstock
The lignocellulosic feedstock can be converted to a wide range of high value‐added prod-
ucts through biorefinery processes. Many of the chemicals derived from lignocellulosics
are known as building blocks or platform chemicals as a variety of other products, includ-
ing biopolymers, can be produced from them. For example, glucose, derived from cellulose,
can be fermented to ethanol by yeast and bacteria in anaerobic conditions, which can be
further dehydrated to ethylene, the monomer for polyethylene, or it can be dehydrogenated
and dehydrated to produce 1,3‐butadiene, the monomer for synthetic rubber. Another
example is that glucose can be converted to lactic acid by lactic acid bacteria, which is the
monomer to produce PLA. Some examples of high value‐added products obtained from the
main lignocellulosic components are shown in Figure 7.11 (Liu 2012).
7.5.1 Lignocellulosic Feedstock

Lignocellulosic biomass is the most abundant renewable raw material available in the
world, which is composed of cellulose, hemicelluloses and lignin as major components,
and pectin, waxes, proteins, lipids, ash, pigments and extractive compounds as minor com-
ponents. This type of biomass is a promising feedstock for biofuels, bioenergy, biomaterials
and biochemicals since it is a renewable material available in large amounts (Lee et al.
2014; Santos et al. 2017a).
Lignocellulosic feedstock can be divided into four different groups: energy crops, agricul-
tural residues, forestry residues, and industrial and municipal wastes. Energy crops are
normally short rotation, high productivity, low cost and low maintenance, including
grasses, wood, and agricultural energy crops. Some examples of energy crops are bamboo,
wheatgrass and alfalfa hay, as grasses; silver maple, black walnut and hybrid poplar, as
wood; sunflower, olive and oilseed rape, as agricultural energy crops. Agricultural residues
are by‐products of commercial crops such as stalks, leaves, and husks. The most common
agricultural residues are sugarcane bagasse, sugarcane straw, rice husk, rice straw, corn
stover, and wheat straw. Forestry residues include biomass not harvested from commercial
logging sites and waste generated during forest management. Lastly, industrial waste
includes residues of wood pulping, as black liquor, and municipal waste includes residen-
tial, commercial, and institutional wastes that contain plant‐derived organic materials. The
composition of lignocellulosic feedstock varies with the type of biomass (e.g., energy crop
or agricultural residue), type of plant, part of the plant, cultivation conditions, and plant
age. Table 7.4 shows some examples of different types of lignocellulosic biomass (Maity
2015; Santos et al. 2017a; Machado et al. 2018).
Biomass
Feedstock
Platform
Conversion/refining
Cellulose Hemicellulose Lignin
Uronic acids Phenolic resins

Esters Solid fuels
Glucose
Ethers
Phenol, Vanillin,
Acetic acid
Lignin Oxidized Oils
Enzymes Hydroxymethyl Xylose; Glucose; Arabinose;
furfural Mannose; Galactose. Gas synthesis
Nylon
Vitamin c Sorbitol resins PHAs
Furfural Methanol
Levulinic acid Polyesters

SCP Lisin A Glutamic acid
Organic acids
Combustible
Ethanol Resins and liquids
Polyhydroxyalkanoates Plastics Furan
Acetone
Chemicals
Butadiene Rubbers
Xylitol SCP
Butanol Sorbitol
Acetone Mannitol
Esters Galactol
Glycerol Enzymes
Citric acid Arabitol
Butyric acid
Glutamic acid Ethylene Plastics
Lactic acid
Gluconic acid
Succinic acid
Figure 7.11 Some high value-added products obtained from lignocellulosic feedstock. Source:
Adapted from Boneberg et al. 2016; open access article, copyright permisison provided under
Creative Commons Attribution 4.0 International License).
7.5.2 Biopolymer Production from Lignocellulosic Biomass

With increasing technologic advances, biopolymer production is rapidly becoming eco-
nomically interesting, especially using lignocellulosic biomass and agro‐industrial waste as
raw material. The production of biopolymers is increasingly attracting broad interest since
they not only come from a renewable source, contributing to the replacement of some oil‐
based products, but they are also biodegradable and have great potential in medical appli-
cations, as they have a higher chance of being biocompatible. A great variety of biopolymers
are found in nature (plants and animals) and can also be produced by microorganisms
(polysaccharides, polyesters, polyamides, polyamines, etc.) (Fasciotti 2017).
Biopolymer production can be divided into two types, depending on the biorefinery method
used for production. First is direct extraction of biopolymer from biomass or agro‐industrial
residue, where the product can be directly obtained or refined through physical and chemical
processes, which can be followed by some chemical modification, as shown in the following
sections. The second is biopolymer production through biological processes, which includes
the use of biological catalysts (enzymes) or microorganisms. For the first method, biopolymer
can be already present in the raw material, which is extracted through a series of processes in
the renewable feedstock until the biopolymer is obtained. In this case, processes can include
Table 7.4 Chemical composition of different types of lignocellulosic biomass (Lee et al. 2014;
Boneberg et al. 2016; Machado et al. 2016; Santos et al. 2017a).
Cellulose Hemicelluloses Lignin

Type of biomass Lignocellulosic biomass (%) (%) (%)
Energy crop Switchgrass 45 31 12

Cotton 95 2 0.3
Empty fruit bunch 41 24 21.2
Agricultural Nut shells 25–30 25–30 30–40
residue Sugarcane straw 40–44 30–32 22–25
Sugarcane bagasse 32–48 19–24 23–32
Corn stover 35 25 35
Rice straw 43.3 26.4 16.3
Forestry residue Hardwood stems 40–55 24–40 18–25
Leaves 15–20 80–85 0
Softwood stems 45–50 25–30 25–35
Industrial/ Waste papers from chemical pulps 60–70 10–20 5–10
municipal waste Recycled paper sludge 60.8 14.2 8.4
Paper residues 76 13 11
Organic compound from 8–15 0 0
wastewater solids
physical (mechanical breaking of fibers), chemical (acid hydrolysis), and biological (enzyme
hydrolysis) treatments and purification of the polymer of interest. For the second method,
the feedstock needs to have enough resources for the microbial transformation of simple
compounds into the polymer. Polymerization is done by a microorganism or a biocatalyst
(enzyme) in a controlled environment, and a final separation step of the polymer from other
compounds in the medium is required (Kawaguchi et al. 2016).
In Table 7.5, there are some examples of biopolymers obtained by each type of conver-
sion method used for production of biopolymers, along with their possible current indus-
trial application. Technology development around new biopolymers and new applications
of renewable products brings great promise in this industry for the near future (Rehm 2010;
Brodin et al. 2017).
7.6 Chemical and Structural Modifications of Extracted

Lignocellulosic Components
7.6.1 Cellulose Derivatives

Cellulose has been used for centuries as an energy source, as raw material for paper and
textile industries, as well as for building materials in its natural form, and many industries
continue to apply this polymer in their processes. However, in order to obtain high
Table 7.5 Examples of biopolymers split by type of production process and their applications.
Production process Biopolymer Application
Extraction and Cellulose Paper products, textiles, drugs, emulsifier, biofuel,

chemical and biomaterials
polymerization Hemicelluloses Thickeners, films, binders, food stabilizers,
cosmetics, and drugs
Lignin Antioxidants, bioplastics, cosmetics, asphalt,
paper, and biochemicals
Starch Food, paper, adhesives, textiles, biofuel, and oil
exploration
Pectin Food, drugs, cosmetics, animal feedstock additive,
adhesive, and stabilizer
Biological Xanthan Food additive (thickener), emulsifier, oil
polymerization exploration, and drugs
Alginate Biomaterial, tissue scaffold, and drug delivery
Nanocellulose Food, wound dressing, and drug delivery
Hyaluronic acid Cosmetics, viscosupplementation, tissue repair,
and drug delivery
Dextran Blood plasma extender and chromatography media
v alue‐added products from cellulose, chemical and structural modifications need to be

accomplished. These modifications are required because natural cellulose has some disad-
vantageous properties such as incompatibility with hydrophobic polymer matrixes,
tendency to form aggregates during processing, and water‐swellable nature, especially in
its amorphous regions (Brinchi et al. 2013).
Chemical modifications that can be performed on cellulose include esterification,
including acetylation, acylation and sulfation, and etherification, including carboxymeth-
ylation, ethylation, and propylation (Granström 2009). Some examples of cellulose
chemical modifications are shown in Table 7.6. Besides derivatives obtained through
chemical modifications, it is also possible to obtain microcrystalline cellulose (MCC) and
nanocellulose from lignocellulosic feedstock. MCC is obtained from purified and partially
depolymerized cellulose, and is a commercial material largely applied in the food and
pharmaceutical industries, whose dimensions are in the range of 10–50 μm. MCC is the
starting material for nanocellulose that presents at least one dimension in nanoscale
(Brinchi et al. 2013; Trache et al. 2016).
Microcrystalline cellulose production consists of the pretreatment and pulping of the
lignocellulosic feedstock, followed by a separation, normally a hydrolysis process. The
mechanism of acid hydrolysis is shown in Table 7.6. After separation, the next steps
involved in MCC production are neutralization, washing, filtration, and drying. Although
there are various ways to isolate MCC, the most common commercial process is acid
hydrolysis. Acid hydrolysis is widely applied because it requires a shorter reaction time,
the isolation process is continuous, a small amount of acid is employed, and small cellu-
lose particles are produced. Also, hydrolysis under controlled conditions of temperature,
Table 7.6 Chemical and structural modification in cellulose.
Modification Structure References
Esterification of cellulose OH OAc Konwar et al. (2016)

with acetic anhydride to HO OH Excess Ac2O AcO OAc
O O
cellulose acetate O
HO O
O
O O
AcO O
O
O
OH Sulfonated carbon catalyst OAc
OH n OAc n
80°C, 9–12h
Cellulose Solvent free conditions
Cellulose acetate
Solid state reaction
mechanical stirring
Carboxymethylation of OH
Aqueous NaOH
OR Ali et al. (2013)
cellulose with aqueous O HO OH ClCH2COONa
O O
O O
NaOH and ClCH2COONa HO O
O RO O
OCH2COONa
OH
OH n n
Cellulose
R = H or R =CH2COONa
Sodyum Carboxymethyl Cellulose
Microcrystalline cellulose OH OH Trache et al. (2016)

(MCC) isolation – acid O HO OH H+
O HO OH
O O
hydrolysis HO O O
O
HO O+ O
O
OH OH H
OH n OH n
OH OH
HO OH H2O HO OH
O O O O O
HO O HO O
HO OH HO O
OH OH2
OH n OH n
(Continued)
c07.indd 139 11/12/2019 6:36:58 PM

Modification Structure References
Nanocellulose production Brinchi et al. (2013)

Pulping Bleaching
from lignocellulosic
biomass Chipping
Cellulose
Milling
Acid
Lignocellulosic
Steam hydrolysis
biomass Fractionation
explosion
NCC Glucose
Sonication Dialysis Centrifugation NCC
suspension Acid
Source: Images in this table are adapted from mentioned references with permission from respective publishers.
time, acid concentration, and acid/cellulose proportion leads to a partial rupture in the
polymer, releasing cellulosic microcrystals presenting a linear rigid arrangement due to
acid penetration in amorphous regions. The most used acids are sulfuric and hydrochloric
acid, and combinations of these two acids are also used in order to produce polymers with
different physical and chemical properties. Although acid hydrolysis is the most common
isolation method, other processes like alkali hydrolysis, steam explosion, extrusion, and
radiation‐enzymatic processes can also be employed (Trache et al. 2016; Ismail et al. 2018;
Kian et al. 2018).
Nanocellulose is categorized into two groups: nanocrystalline cellulose (NCC) and
nanofibrillated cellulose (NFC). Both types exhibit similar chemical characteristics but the
difference is the physical characteristic of their colloidal forms – NCC resembles cellulosic
“rice” while NFC resembles cellulosic “spaghetti.” Nanocellulose production processes are
very similar to those involved in MCC production. A scheme of steps for nanocellulose
production from lignocellulosic biomass is shown in Table 7.6 (Lee et al. 2014).
7.6.2 Hemicellulose Derivatives

Chemical modifications in hemicelluloses are generally performed to improve the hydro-
phobic, thermoplastic, and functional properties of these polysaccharides so that different
biopolymers can be obtained for various applications (Peng and She 2014). Since hemicel-
luloses possess some free hydroxyl groups, these biopolymers have a hydrophilic character,
while synthetic polymers largely used in the industry have a hydrophobic character. Besides
that, hemicelluloses do not have a uniform chemical structure, since they are composed of
different monomers linked by different types of glycosidic bonds forming branched and
amorphous structures. These characteristics limit the industrial application of hemicellu-
lose in its natural form, but some chemical modifications can enable its utilization in a
variety of applications (Sun et al. 2003).
Modifications mostly focus on providing a hydrophobic character to hemicelluloses,
since their hydrophilic characteristics result in poor mechanical properties due to poor
interfacial adhesion when these biopolymers are blended with conventional plastics.
Hemicelluloses have been modified by a variety of reactions including oxidation, reduc-
tion, esterification, cross‐linking, and etherification. Some examples of the most important
chemical modification used in hemicelluloses are shown in Table 7.7 (Farhat et al. 2017).
7.6.3 Lignin Derivatives

Lignin is the aromatic biopolymer most abundantly found in Nature, so it has huge poten-
tial to become the main renewable resource to produce aromatic compounds in the chemi-
cal industry, such as phenolic resins, surfactants, epoxy resins, adhesives, polyester, and
many other products (Agrawal et al. 2014). However, some undesirable characteristics of
lignin restrict their utilization at large scale. The fact that lignin changes its properties sig-
nificantly when relative moisture changes and has poor mechanical properties at mild tem-
peratures are examples of properties that limit its utilization for most industrial applications.
Another disadvantage presented by lignin is its dark brown color, which persists even
when it is submitted to a bleaching process. Free radicals present in lignin slowly react with
Table 7.7 Chemical and structural modifications in hemicelluloses.
Modification Method Structure References
Esterification General – reaction O Hemicelluloses Farhat et al.

O O
between an alcohol (2017)
HO OH n
and carboxylic Dimethylaminopyridine as catalyst
acid, acid chlorides
or acid anhydrides
N N O
+ C
N R N R O R
C O C
O O
C O
Acid anhidride R
or acid derivatives
O O N
O O O O
C + O +
HO R HO n HO n N
O H O
C C
R O R O
O
N C
O R
N R
C
O O O
O Hemicelluloses ester derivatives
O n
O
C C
R O R O
c07.indd 142 11/12/2019 6:36:58 PM

Transesterification O

O O
O O O Peng et al.
HO
OH n O O + HO (2012)
Xyl
Xylose Transesterification O
Acetylation O O

O O H 3C O CH3 O O O Farhat et al.
O HO
HO O (2017)
OH C O
n
CH3 n
Hemicelluloses
Acetylated hemicelluloses
Benzylation of
O
hemicelluloses O O
O
CH2Cl O
O
O Junli et al.
with benzyl HO
OH CH2
O
n (2012)
n
chloride NaOH CH2
Hemicelluloses
Benzylated hemicelluloses
Fluorination Reaction with O O

gaseous O O
O CF3 O CF3 O O
O Farhat et al.
trifluoroacetic HO
OH
O
O (2017)
C O n
anhydride n
C
CF3 O
Hemicelluloses CF3
Fluorinated Hemicelluloses
(Continued)
c07.indd 143 11/12/2019 6:37:00 PM

Cross‐ Thermo cross‐ O

OH Wu et al.
linking linking between HOOC
(2017)
carboxymethyl H3CO O HOH2C OH
HO
chitosan and OCH2COOH OCH2COOH OH
hemicelluloses NHCOCH3 NHCOCH3
HO OH HO OH HO HO
O O
O
O O O
O O O O +HO O O O
O O O O O O
NH2 CH2OH OH OH OH OH OH
NH2 CH OH
2
CMC HC
O
OH
HOOC
H3CO O HOH2C OH
HO OCH2COOH
OH
OH HO OH HO HO O N O
HO O O O
HO O O O O
O O O
OH OH OH OH OH
Thermo-crosslinking O HOH2C NHCOCH3
OH O
HOOC O
H3CO O HOH2C OH
HO OCH2COOH
OH
OH OH HO HO O N O
HO O HO O O
HO O O O O O
O O
OH OH OH OH OH
HOH2C NHCOCH3
O
O
CMCH
Source: Images in this table are adapted from mentioned references with permission from respective publishers.
c07.indd 144 11/12/2019 6:37:00 PM

atmospheric oxygen after the bleaching process which results in the lignin darkening to a
black or brown color, as time goes by (Naseem et al. 2016).
In order to overcome these limitations, chemical and structural modifications have been
proposed in several studies. Some of the modifications applied to lignin that allow its use
in a wider range of applications include alkylation and alkoxylation, esterification, co‐poly-
mer in polyurethane synthesis, lignin‐based phenol‐formaldehyde resin synthesis, and
grafting. Some examples of the most important chemical modifications used in lignin are
shown in Table 7.8 (Sen et al. 2015).
7.7 Microbial Conversion of Biomass into Biopolymers
Biopolymer production can be done by microbial or enzyme polymerization. Microbial

polymerization can utilize various enzymes (inside their cells) to perform substrate trans-
formation and further polymerization. The polymer production process mainly depends on
the nutrients needed for microbial growth and appropriate conditions for polymer produc-
tion. The optimization highly depends on the environment with controlled temperature,
pressure, humidity, and pH (Rehm 2009).
Biopolymer production via fermentation is a huge advance in bioprocess technology and
has environmental benefits. First, the production of a biodegradable polymer reduces the
accumulation of waste in Nature; second, the production processes tend to generate less
pollution, comparing to oil‐based polymers. Also, it is possible to use biomass or waste as
nutrient material, adding value to agro‐industrial by‐products. And finally, the biopoly-
mers thus produced are generally less toxic, having potential applications in the food and
medical industries (Basnett and Roy 2010).
Some examples of biopolymers, their main components, microorganisms generally used
in their production and industrial applications are shown in Table 7.9. Production of
biopolymers can be intracellular or extracellular, which influences the subsequent purifi-
cation processes. Extracellular polymers are directly purified after fermentation whereas to
obtain intracellular polymers, there is a need to first remove it from within the cell, which
can increase production costs. Various microorganisms under specific growth and produc-
tion conditions produce these polymers. For example, PHAs are produced as storage mol-
ecules under nutrient stress conditions by bacteria such as Cupriavidus necator and Bacillus
megaterium, and have great commercial potential as bioplastic. Another example is PLA, a
biodegradable polymer produced by some lactic bacteria of the Lactobacilli genera with
great industrial applications in the packaging sector. Another example of polysaccharide
production is extracellular xanthan, which is synthesized by the bacterium Xanthomonas
campestris, and used in food, pharmaceuticals, cosmetics, materials and even the petro-
leum industry (Basnett and Roy 2010; Jamshidian et al. 2010).
Production of biopolymers from renewable feedstocks by microbial fermentation pre-
sents an alternative to petroleum‐based polymers. As mentioned by Kawaguchi et al.
(2016), several biopolymers have been used as an alternative to existing oil‐based polymers
such as polybenzoxazoles and aromatic polyimides. Also, some biopolymers (xanthan,
alginate, dextran, and PHA) have well‐established markets, with increasing research into
Table 7.8 Chemical and structural modifications in lignin.
Modification Method Scheme Property changes References
Alkylation Methylation of L L Reduction in Tg since

lignin using (A) the intramolecular Sen et al.
Me2SO4, NaOH, H2O
dimethyl sulfate hydrogen bonds are (2015)
and (B) methyl H3CO rt, 80°C, 2h H3CO eliminated after
iodide. OH OCH3
complete
derivatization of
softwood lignin.
L L
Alkylation completely
MeI, K2CO3, DMF
removes the possibility
rt,18h
of radical initiated
H3CO H3CO
self‐polymerization of
OH OCH3 the KL
Alkoxylation Reaction initiated L L

by base attack on + OH– + H2C
O
CH2 Reduction in Tg due to
the phenolic H3CO H3CO increase of ethylene or Meister
hydroxyl groups to OH O– propylene oxide in the (2002)
form phenoxide lignin structure.
groups, which Primary hydroxyl
then attack the groups inserted in
polar epoxide ring lignin structure make
L
it more reactive
H3CO
OCH2CH2O–
Esterification Synthesis of OH

lignin‐based Improved flow
thermoplastic properties. Saito et al.
polyesters O O
(2012)
co‐polymerized O
n
O
with carboxy R2 R1
KOH
terminated OH
telechelic 1, 4-Dioxane,100°C, 24h
+
polybutadiene R2 R1 R1 R2
HOOC COOH OH OH
n
c07.indd 146 11/12/2019 6:37:01 PM

Polyurethane Synthesis of OH
Polybutadiene diisocyanate

synthesis block co‐polymers Lignin segment
O
using lignin and O
provides mechanical Saito et al.
soft elastomeric + N O O N strength to the (2013)
H3CO H H
material to form NCO polymer and the soft
O
thermoplastic 1, 4-dioxane
synthetic polymer
L
elastomers Dibutyltindilaurate delivers rubbery flow
24h characteristics
O O O O
O N N O O N N O
H H H H
n
H3CO OCH3
O O
L L
Synthesis of Reaction of free Lignin Lignin

Lignin Lignin
Lignin replaces phenol
lignin‐based phenolic hydroxyl HCHO
in these resins, which Sen et al.
+
phenol‐ groups of the ortho is a fossil‐derived (2015)
formaldehyde positions with H3CO
H3CO chemical whose cost
H3CO OCH3
resins formaldehyde OH
OH has been growing
OH OH
(Continued)
c07.indd 147 11/12/2019 6:37:02 PM

Grafting Free radical Polimerization reaction

polymerization of Lignin Biodegradable Hollmann
acrylamide onto NH2
alternative to and
lignin O conventional acrylic Arends
O2 OH
R
O
O
Polimerization homopolymer (2012)
OH
Laccase
Lignin
H2O O H
R O
O
OH
O
Grafting
O R
H2N nO
O OH
O O NH2
+ O R
H2N n O O OH
Lignin O NH2
Lignin
c07.indd 148 11/12/2019 6:37:03 PM

Table 7.9 Examples of commercially available biopolymers, their main components, usual producer, and industrial applications.
Polymer class Biopolymer Main components Producer Industrial application
Polysaccharide Xanthan Glucose, mannose and Xanthomonas spp. Food additive (thickener),
glucuronate drugs, and emulsifier
Polysaccharide Dextran Glucose Leuconostoc spp. and Streptococcus spp. Blood plasma extender and
chromatography media
Polysaccharide Nanocellulose D‐glucose Gluconacetoobacter spp., Rhizobium spp., Food additive, diaphragms of
Agrobacterium tumefaciens and Sarcina acoustic transducers, wound
ventriculli dressing, and drugs
Polysaccharide Alginate Mannuronic acid and Pseudomonas spp. Biomaterial, drug delivery, and
glucuronic acid and Azotobacter tissue scaffold
spp.
Polyester Polylactic acid Lactic acid Lactobacillus spp. Packaging, plastics and
mulching films
Polyester Polyhydroxyalkanoates (R)‐β‐hydroxyacyl CoA Cupriavidus necator, Pseudomonas spp. and Bioplastic, biomaterial and
Bacillus megaterium matrices for displaying or
binding proteins
Polyanhydrides Polyphosphate ATP Alcaligenes spp., Trypanosoma spp. and Replacement of ATP in
Saccharomyces cerevisiae enzymatic synthesis and flavor
enhancer
Source: Adapted from Rehm 2010 with permission from Elsevier.
c07.indd 149 11/12/2019 6:37:03 PM

new applications. This continued advance in technologies aiming to increase and optimize
biopolymer production is expected to facilitate the establishment of sustainable biorefinery
processes. Also, research focused on utilization of agro‐industrial waste for biopolymer
production will further encourage its use in industrial processes, decreasing costs in pro-
duction and environmental impacts.
7.8 Lignocellulosic Biopolymer Properties

Several types of biopolymers can be obtained from lignocellulosic biomass by a variety of
production processes. These different routes of biopolymer production lead to products
with a variety of molecular and chemical structures that result in materials with different
properties. Since it is not possible to generalize properties of lignocellulosic biomass‐
derived biopolymers, in this section the main properties of some of the most common
biopolymers produced from lignocellulosics are discussed (Boneberg et al. 2016).
7.8.1 Hydroxyethyl Cellulose (HC)

A biopolymer with a high molar mass and fairly rigid backbone. Solutions containing this
biopolymer are usually tolerant to temperature and mechanical shearing. Also, HC has
properties like salinity durability and stability in neutral and basic pH (Pu et al. 2018).
7.8.2 Methylcellulose (MC)

A biopolymer with higher thermal stability and higher solubility in water due to an increase
in the degree of substitution (DS) when compared to cellulose. This polymer has the char-
acteristic of forming a gel. The gelation process consists of the disruption of hydrogen
bonds between the polymer and the surrounding solvent, as the temperature of the solu-
tion containing the polymer increases. Gelation temperature depends on salt concentration
in solution; as salt concentration increases, water molecules tend to surround the salt and
consequently gelation temperature decreases (Boneberg et al. 2016).
7.8.3 Carboxymethylcellulose (CMC)

This is a biopolymer whose properties are highly dependent on the DS of hydroxyl groups
on the anhydroglucose unit and distribution of carboxymethyl substituent. Some examples
of polymer properties affected by substitution of hydroxyl groups in cellulose structure are
CMC solubility in water, which depends on the DS of hydroxyl groups on anhydroglucose
by carboxymethyl groups, and rheology of CMC solutions that are dependent on the distri-
bution of carboxymethyl substituents in the polymer structure (Pu et al. 2018).
7.8.4 Polyhydroxyalkanoates
A class of biopolymers exhibiting different characteristics based on type of biopolymer, raw
material used as feedstock, and microorganism that synthesized the biopolymer. PHAs
produced from Bacillus species have properties more similar to propylene than PHAs
roduced from other genera, such as better melting stability, crystallinity, tensile
p
strength, and elongation‐to‐break (Mohapatra et al. 2017). Some examples of PHAs that
present different properties are polyhydroxybutyrate (PHB) and polyhydroxybutyrate‐
co‐hydroxyvalerate (PHBV). Regarding mechanical properties, PHB is stiffer and more
brittle than PHBV. Also, their chemical properties differ; PHB solvent resistance is inferior
but it has better natural resistance to ultraviolet (UV) weathering. PHB’s thermochemical
properties can be related to the polymer molecular weight. Melting temperatures, crystal-
lization, and degradation initially increase with a higher molecular weight, but higher
molecular weights exhibit cold crystallization and reduction in melting temperature
and melting enthalpy, suggesting difficulty of crystallization. The properties of PHBV are
closely related to valerate content present in its structure; once its presence reduces poly-
mer crystallinity and melting point, it can lead to a decrease in stiffness but a higher tough-
ness or impact resistance (Bastioli 2005; Domínguez‐Díaz et al. 2015).
7.8.5 Polylactic Acid (PLA)

This is a rigid thermoplastic biopolymer whose properties mainly depend on its optical
purity, since its crystallinity and melting temperature reduce with a decrease of optical
purity of the L‐unit (Bastioli 2005). This polymer has some undesirable properties which
limit its application such as brittleness, poor thermal resistance, limited gas barrier proper-
ties, and poor UV light barrier properties (Boneberg et al. 2016).
7.8.6 Polyethylene (PE)

Polyethylene is a biopolymer that can have different amounts of crystalline and amorphous
regions, whose properties are highly dependent on this feature. Regarding its chemical
properties, PE is inert to most chemicals due to its paraffinic nature, high molecular weight,
and partially crystalline structure (Boneberg et al. 2016).
7.9 Biocomposites
Biocomposites are defined as biocompatible and/or eco‐friendly composites. They are a

combination of two or more materials or phases, in which one of the components is pre-
sent in a higher concentration forming a matrix, while the material present in smaller
amounts acts as a reinforcement inducing enhanced performance, such as improving
some properties (Fernandes et al. 2013; Haraguchi 2015). The combined use of a biopoly-
mer with a natural fiber filler in a biocomposite may improve the material’s properties,
enabling its use in other applications. Table 7.10 shows some examples of biocomposites
containing biopolymers and fillers obtained from lignocellulosic feedstock and their
possible applications (Sánchez‐Safont et al. 2018). Lignocellulosic fibers are interesting
bio‐based reinforcement/fillers because of some characteristics they provide to the
biocomposite, such as nonbrittle fracture on impact, same performance of the biopolymer
with a lower weight, stronger for the same weight, lower cost than the base resin, among
others (Satyanarayana et al. 2009).
Table 7.10 Biocomposites obtained from lignocellulosic biomass.
Biocomposites Application/properties References
PHB/lignocellulosic Food packaging/better permeability and Sánchez‐Safont

waste mechanical properties et al. (2018)
PLA/bentonite and Packaging/reduction in oxygen permeability Petersson and
MCC and amount of light transmitted Oksman (2006)
HC/CMC sodium salt Bulking agent for stomach; treatment of Fernandes et al.
overweight; drug delivery (2013)
Lignin/xanthan gum Biodegradable hydrogel‐films Fernandes et al.
(2013)
CMC/NFC powder Human nucleus pulposus Fernandes et al.
(2013)
CMC/polyacrylic acid Removal of Pb(II) ions in waste water Wang et al. (2017)
PLA/NCC and silver Food packaging, antibacterial effects Fortunati et al.
nanoparticles (2012)
Corn starch/cellulose Composite foams formed; tensile Lawton et al. (2004)
fiber strength increased with increasing
fiber content up to 15 wt% fiber
Polyester amide/hemp The higher the amount of fiber, the faster Keller (2003)
the degradation
Hemicelluloses/chitosan Drug delivery Karaaslan et al.
(2010)
PLA and pine resin/açaí Flexural strength slightly increased Santos et al. (2017b)
short fibers
PHB and co‐polymer Bending modulus increased Shanks et al. (2004)
PHV/flax
CMC, carboxymethylcellulose; NCC, nanocrystalline cellulose; NFC, nanofibrillated cellulose; PHB,

polyhydroxybutyrate; PHV, polyhydroxyvalerate; PLA, polylactic acid.
7.10 Advantages and Challenges
Global polymer consumption has been continuously rising in recent decades, since these
materials are widely applied in various industries, for example in the production of chemi-
cals, electronic equipment, automotive supplies, packages, and many others. Conventional
plastics are petroleum based, a nonrenewable resource, contribute to greenhouse gas emis-
sions, and as they are not biodegradable, the waste generates a problem, since its accumula-
tion and degradation resistance cause environmental hazards. A huge amount of
conventional plastics is improperly disposed into landfill which not only could be used as
carbon feedstock to generate energy by burning theses residues, but also can harm the
environment by potentially causing groundwater pollution by leaching of toxic additives
(Osman et al. 2016; Trache et al. 2016; Dietrich et al. 2017).
In this scenario, the replacement of conventional polymers by bio‐based polymers is an

excellent choice, since biopolymers fulfill the principles of sustainability, eco‐efficiency,
industrial ecology, green chemistry and engineering, and have the potential of reducing
carbon footprint and greenhouse gas emissions. Regarding economic aspects, biopolymers
derived from lignocellulosic biomass can contribute to the economic growth and develop-
ment of new areas, especially rural areas due to the use of agricultural residues and energy
crops; improvement in energy security, by reducing dependence on nonrenewable
resources; and economic partnership between agricultural and industrial sectors (Boneberg
et al. 2016; Trache et al. 2016).
Since lignocellulosic biomass is a heterogeneous material, a wide range of different
biopolymers can be produced from this feedstock. Besides that, lignocellulosic materials
are largely available natural resources whose cost is relatively low, making their use in
biopolymer production feasible. Biopolymers produced from distinct components of bio-
mass through different production processes present different properties, allowing their
application in a variety of industry sectors (Iles and Martin 2013).
One of the challenges of producing biopolymers at large scale to replace conventional
plastics is how to integrate this sustainable innovation in the chemical industry business
models. Although reducing costs, increasing yields, and developing better feedstock sup-
plies may contribute to the success of bioplastics insertion in the market, there are some
drawbacks in replacing conventional plastics, including the high investment to switch pro-
duction of conventional plastics to bioplastics; lack of quality standards; and uncertain
customer response (Iles and Martin 2013; Dietrich et al. 2017).
In this sense, it is important to develop a sustainable model of bioplastics production, in
which the processes are improved continuously, in such a way that biopolymer businesses
not only can maintain competitiveness but can guarantee a more environmentally friendly
option to replace conventional plastics (Iles and Martin 2013).
7.11 Conclusion
Lignocellulosic biomass is a promising raw material for the production of biopolymers

aiming to replace conventional plastics. Polymers derived from petroleum present disad-
vantages including greenhouse gas emissions, dependence on a nonrenewable resource,
and generation of huge amounts of nonbiodegradable waste. Thus, their replacement by
biopolymers, which are biodegradable and environmentally friendly, is a favorable
alternative.
Since lignocellulosic biomass is a heterogeneous material, a wide range of different
biopolymers can be produced from this feedstock. Besides that, lignocellulosic materials
are widely available natural resources whose cost is relatively low, making their use in
biopolymers production feasible. Biopolymers produced from distinct components of bio-
mass through different production processes present different properties, allowing their
application in a variety of industry sectors. In this sense, it is increasingly important to
support research focused on making biopolymer production from lignocellulosic biomass
commercially feasible and lucrative.
R
eferences
Abu‐Dalo, M.A., Al‐Rawashdeh, N.A.F., and Ababneh, A. (2013). Evaluating the performance
of sulfonated Kraft lignin agent as corrosion inhibitor for iron‐based materials in water
distribution systems. Desalination 313: 105–114.
Agrawal, A., Kaushik, N., and Biswas, S. (2014). Derivatives and applications of lignin– an
insight. Scitech Journal 01: 30–36.
Alam, M.N. and Christopher, L. (2018). Natural cellulose‐chitosan cross linked superabsorbent
hydrogels with superior swelling properties. ACS Sustainable Chemistry & Engineering 6:
8736–8742.
Ali, Z.M., Mughal, M.A., Laghari, A.J. et al. (2013). Polymeric cellulose derivative:
carboxymethyl‐cellulose as useful organic flocculants against industrial waste waters.
International Journal of Advance Research and Technology 2: 14–20.
ASTM (2019). ASTM D883‐19b: Standard Terminology Relating to Plastics. www.astm.org/
Standards/D883.htm.
Basnett, P. and Roy, I. (2010). Microbial production of biodegradable polymers and their role in
cardiac stent development. Current Research, Technology & Education Topics in Applied
Microbiology & Microbial Biotechnology 2: 1405–1415.
Bastioli, C. (2005). Handbook of Biodegradable Polymers. Crewe, UK: Rapra Technology
Limited Press.
Boneberg, B.S., Machado, G.D., Santos, D.F. et al. (2016). Biorefinery of lignocellulosic
biopolymers. Revista Eletrônica Científica Da UERG 2: 79.
Brinchi, L., Cotana, F., Fortunati, E., and Kenny, J.M. (2013). Production of nanocrystalline
cellulose from lignocellulosic biomass: technology and applications. Carbohydrate Polymers
94: 154–169.
Brodin, M., Vallejos, M., Opedal, M.T. et al. (2017). Lignocellulosics as sustainable resources
for production of bioplastics – a review. Journal of Cleaner Production 162: 646–664.
Dietrich, K., Dumont, M.J., del Rio, L.F., and Orsat, V. (2017). Producing PHAs in the
bioeconomy: towards a sustainable bioplastics. Sustainable Production & Consumption 9:
58–70.
Ding, R., Wu, H., Thunga, M. et al. (2016). Processing and characterization of low‐cost
electrospun carbon fibers from organosolv lignin/polyacrylonitrile blends. Carbon N.Y. 100:
126–136.
Domínguez‐Díaz, M., Meneses‐Acosta, A., Romo‐Uribe, A. et al. (2015). Thermo‐mechanical
properties, microstructure and biocompatibility in poly‐β‐hydroxybutyrates (PHB) produced
by OP and OPN strains of Azotobacter vinelandii. European Polymer Journal 63: 101–112.
E‐Bioplastics (2018). Bioplastics: Industry Standards & Labels. https://docs.european‐
bioplastics.org/publications/fs/EUBP_FS_Standards.pdf.
Emadian, S.M., Onay, T.T., and Demirel, B. (2017). Biodegradation of bioplastics in natural
environments. Waste Management 59: 526–536.
Farhat, W., Venditti, R.A., Hubbe, M. et al. (2017). A review of water‐ resistant hemicellulose‐
based materials: processing and applications. ChemSusChem 10: 305–323.
Faria, D., Machado, G., Eichler, P. et al. (2016). Cenários e perspectivas das principais
culturas do Rio Grande do Sul em processos de biorrefinaria. Revista Eletrônica Científica
Da UERGS 2: 291.
Fasciotti, M. (2017). Perspectives for the use of biotechnology in green chemistry applied to
biopolymers, fuels and organic synthesis: from concepts to a critical point of view.
Sustainable Chemistry and Pharmacy 6: 82–89.
Fernandes, E.M., Pires, R.A., Mano, J.F., and Reis, R.L. (2013). Bionanocomposites from
lignocellulosic resources: properties, applications and future trends for their use in the
biomedical field. Progress in Polymer Science 38: 1415–1441.
Fortunati, E., Armentano, I., Zhou, Q. et al. (2012). Multifunctional bionanocomposite films of
poly(lactic acid), cellulose nanocrystals and silver nanoparticles. Carbohydrate Polymers 87:
1596–1605.
Granström, M. (2009). Cellulose derivatives: synthesis, properties and applications. Academic
dissertation submitted to University of Helsinki, Finland.
Grewal, J. and Khare, S.K. (2018). One‐pot bioprocess for lactic acid production from
lignocellulosic agro‐ wastes by using ionic liquid stable Lactobacillus brevis. Bioresource
Gupta, G., Birbilis, N., Cook, A.B., and Khanna, A.S. (2013). Polyaniline‐lignosulfonate/epoxy
coating for corrosion protection of AA2024‐T3. Corrosion Science 67: 256–267.
Hahn, S.K., Chang, Y.K., and Lee, S.Y. (1995). Recovery and characterization of poly(3‐
hydroxybutyric acid) synthesized in Alcaligenes eutrophus and recombinant Escherichia coli.
Applied and Environmental Microbiology 61: 34–39.
Haraguchi, K. (2015). Biocomposites. In: Encyclopedia of Polymeric Nanomaterials (eds. S.
Kobayashi and K. Müllen), 2–19. Berlin, Germany: Springer.
Hollmann, F. and Arends, I.W.C.E. (2012). Enzyme initiated radical polymerizations. Polymers
(Basel) 4: 759–793.
Iles, A., Martin, and AN (2013). Expanding bioplastics production: sustainable business
innovation in the chemical industry. Journal of Cleaner Production 45: 38–49.
Ismail, I., Sa’adiyah, D., Rahajeng, P. et al. (2018). Extraction of cellulose microcrystalline from
galam wood for biopolymer. AIP Conference Proceedings 1945: 020072. https://doi.
org/10.1063/1.5030294.
Jamshidian, M., Tehrany, E.A., Imran, M. et al. (2010). Poly‐lactic acid: production,
applications, nanocomposites, and release studies. Comprehensive Reviews in Food Science
and Food Safety 9: 552–571.
Jeong, D., Joo, S.W., Hu, Y. et al. (2018). Carboxymethyl cellulose‐based superabsorbent
hydrogels containing carboxymehtyl β‐cyclodextrin for enhanced mechanical strength and
effective drug delivery. European Polymer Journal 105: 17–25.
Junli, R., Xinwen, P., Linxin, Z. et al. (2012). Novel hydrophobic hemicelluloses: synthesis and
characteristic. Carbohydrate Polymers 89: 152–157.
Kallakunta, V.R., Tiwari, R., Sarabu, S. et al. (2018). Effect of formulation and process variables
on lipid based sustained release tablets via continuous twin screw granulation: a
comparative study. European Journal of Pharmaceutical Sciences 121: 126–138.
Kanmani, P., Aravind, J., Kamaraj, M. et al. (2017). Environmental applications of chitosan
and cellulosic biopolymers: a comprehensive outlook. Bioresource Technology 242:
295–303.
Karaaslan, A.M., Tshabalala, M.A., and Buschle‐Diller, G. (2010). Wood hemicellulose/
chitosan‐based semi: interpenetrating network hydrogels: mechanical, swelling and
controlled drug release properties. BioResources 5: 1036–1054.
Kawaguchi, H., Hasunuma, T., Ogino, C., and Kondo, A. (2016). Bioprocessing of bio‐based
chemicals produced from lignocellulosic feedstocks. Current Opinion in Biotechnology 42:
30–39.
Keller, A. (2003). Compounding and mechanical properties of biodegradable hemp fibre
composites. Composites Science and Technology 63: 1307–1316.
Khanna, S. and Srivastava, A.K. (2005). Recent advances in microbial polyhydroxyalkanoates.
Process Biochemistry 40: 607–619.
Kian, L.K., Jawaid, M., Ariffin, H., and Karim, Z. (2018). Isolation and characterization of
nanocrystalline cellulose from roselle‐derived microcrystalline cellulose. International
Journal of Biological Macromolecules 114: 54–63.
Kong, F., Parhiala, K., Wang, S., and Fatehi, P. (2015). Preparation of cationic softwood Kraft
lignin and its application in dye removal. European Polymer Journal 67: 335–345.
Konwar, L.J., Mäki‐Arvela, P., Thakur, A.J. et al. (2016). Sulfonated carbon as a new, reusable
heterogeneous catalyst for one‐pot synthesis of acetone soluble cellulose acetate. RSC
Advances 6: 8829–8837.
Koskela, J., Morton, D.A.V., Stewart, P.J. et al. (2018). The effect of mechanical dry coating with
magnesium stearate on flowability and compactibility of plastically deforming
microcrystalline cellulose powders. International Journal of Pharmaceutics 537: 64–72.
Kumar, S., Zafar, M., Kumar, S., and Dhiman, A.K. (2012). Modeling and optimization of
poly(3hydroxybutyrateco‐3hydroxyvalerate) production from cane molasses by
Azohydromonas lata MTCC 2311 in a stirred‐tank reactor: effect of agitation and aeration
regimes. Journal of Industrial Microbiology and Biotechnology 39: 987–1001.
Lawton, J.W., Shogren, R.L., and Tiefenbacher, K.F. (2004). Aspen fiber addition improves the
mechanical properties of baked cornstarch foams. Industrial Crops and Products 19: 41–48.
Lee, H.V., Hamid, S.B.A., and Zain, S.K. (2014). Conversion of lignocellulosic biomass to
nanocellulose : structure and chemical process. Scientific World Journal 2014: 1–20.
Liu, S. (2012). Bioprocess Engineering: Kinetics, Biosystems, Sustainability, and Reactor Design,
2e. Amsterdam, The Netherland: Elsevier.
Machado, G., Leon, S., Santos, F. et al. (2016). Literature review on furfural production from
lignocellulosic biomass. Natural Resources 7: 115–129.
Machado, G., Santos, F., Faria, D. et al. (2018). Characterization and potential evaluation of
residues from the sugarcane industry of Rio Grande do Sul in biorefinery processes. Natural
Resources 9: 175–187.
Maity, S.K. (2015). Opportunities, recent trends and challenges of integrated biorefinery: part
II. Renewable & Sustainable Energy Reviews 43: 1446–1466.
McNaught, A.D. and Wilkinson, A. (1997). Compendium of Chemical Terminology. Research
Triangle Park, NC, USA: International Union of Pure and Applied Chemistry.
Meister, J.J. (2002). Modification of lignin. Journal of Macromolecular Science Part C: Polymer
Reviews 42 (2): 235–289.
Mendes, F.R.S., Bastos, M.S.R., Mendes, L.G. et al. (2017). Preparation and evaluation of
hemicellulose films and their blends. Food Hydrocolloids 70: 181–190.
Mohapatra, S., Maity, S., Dash, H.R. et al. (2017). Bacillus and biopolymer: prospects and
challenges. Biochemistry & Biophysics Reports 12: 206–213.
Nasatto, P.L., Pignon, F., Silveira, J.L.M. et al. (2015). Methylcellulose, a cellulose derivative
with original physical properties and extended applications. Polymers (Basel) 7: 777–803.
Naseem, A., Tabasum, S., Zia, K.M. et al. (2016). Lignin‐derivatives based polymers, blends and
composites: a review. International Journal of Biological Macromolecules 93: 296–313.
de Oliveira, F., Ramires, E.C., Frollini, E., and Belgacem, M.N. (2015). Lignopolyurethanic
materials based on oxypropylated sodium lignosulfonate and castor oil blends. Industrial
Crops and Products 72: 77–86.
Osman, Y., Abd Elrazak, A., and Khater, W. (2016). Microbial biopolymer production by
microbacterium WA81 in batch fermentation. Egyptian Journal of Basic Applied and Sciences
3: 250–262.
Peng, P. and She, D. (2014). Isolation, structural characterization, and potential applications of
hemicelluloses from bamboo: a review. Carbohydrate Polymers 112: 701–720.
Peng, X., Ren, J., Zhong, L. et al. (2012). Glycidyl methacrylate derivatized xylan‐rich
hemicelluloses: synthesis and characterizations. Cellulose 19: 1361–1372.
Petersson, L. and Oksman, K. (2006). Biopolymer based nanocomposites: comparing layered
silicates and microcrystalline cellulose as nanoreinforcement. Composites Science and
Pu, W., Shen, C., Wei, B. et al. (2018). A comprehensive review of polysaccharide biopolymers
for enhanced oil recovery (EOR) from flask to field. Journal of Industrial and Engineering
Rehm, B.H.A. (2009). Microbial Production of Biopolymers and Polymer Precursor. New
Zealand: Caister Academic Press.
Rehm, B.H.A. (2010). Bacterial polymers: biosynthesis, modifications and applications. Nature
Reviews Microbiology 8: 578–592.
Rosca, I., Petrovici, A.R., Peptanariu, D. et al. (2018). Biosynthesis of dextran by Weissella
confusa and its in vitro functional characteristics. International Journal of Biological
Macromolecules 107: 1765–1772.
Saito, T., Brown, R.H., Hunt, M.A. et al. (2012). Turning renewable resources into value‐added
polymer: development of lignin‐ based thermoplastic. Green Chemistry 14: 3295.
Saito, T., Perkins, J.H., Jackson, D.C. et al. (2013). Development of lignin‐based polyurethane
thermoplastics. RSC Advances 3: 21832. https://doi.org/10.1039/c3ra44794d.
Sánchez‐Safont, E.L., Aldureid, A., Lagarón, J.M. et al. (2018). Biocomposites of different
lignocellulosic wastes for sustainable food packaging applications. Composites Part B:
Engineering 145: 215–225.
Santos, F., Machado, G., Faria, D. et al. (2017a). Productive potential and quality of rice husk
and straw for biorefineries. Biomass Conversion Biorefinery 7: 117–126.
Santos, N.S., Silva, M.R., and Alves, J.L. (2017b). Reinforcement of a biopolymer matrix by
lignocellulosic agro‐waste. Procedia Engineering 200: 422–427.
Satyanarayana, K.G., Arizaga, G.G.C., and Wypych, F. (2009). Biodegradable composites based
on lignocellulosic fibers‐an overview. Progress in Polymer Science 34: 982–1021.
Sen, S., Patil, S., and Argyropoulos, D.S. (2015). Thermal properties of lignin in copolymers,
blends, and composites: a review. Green Chemistry 17: 4862–4887.
Shaghaleh, H., Xu, X., and Wang, S. (2018). Current progress in production of biopolymeric
materials based on cellulose, cellulose nanofibers, and cellulose derivatives. RSC Advances
8: 825–842.
Shanks, R.A., Hodzic, A., and Wong, S. (2004). Thermoplastic biopolyester natural fiber
composites. Journal of Applied Polymer Science 9: 2114–2121.
Shogren, R.L. and Biswas, A. (2013). Preparation of starch‐sodium lignosulfonate graft

copolymers via laccase catalysis and characterization of antioxidant activity. Carbohydrate
Polymers 91: 581–585.
Singh, G., Kumari, A., Mittal, A. et al. (2013). Poly β ‐hydroxybutyrate production by Bacillus
subtilis NG220 using sugar industry waste water. BioMed Research International 2013:
952641. https://doi.org/10.1155/2013/952641.
Sun, R., Sun, X.F., and Tomkinson, J. (2003). Hemicelluloses and their derivatives. In:
Hemicellulose Science and Technology (eds. P. Gatenholm and M. Tenkanen), 2–22.
Washington DC, USA: ACS Publications.
Trache, D., Hussin, M.H., Hui‐Chuin, C.T. et al. (2016). Microcrystalline cellulose: isolation,
characterization and bio‐composites application: a review. International Journal of Biological
Macromolecules 93: 789–804.
Wang, M., Leitch, M., and Xu, C. (2009). Synthesis of phenol‐formaldehyde resol resins using
organosolv pine lignins. European Polymer Journal 45: 3380–3388.
Wang, J., Qian, W., He, Y. et al. (2017). Reutilization of discarded biomass for preparing
functional polymer materials. Waste Management 65: 11–21.
Wu, S.P., Dai, X.Z., Kan, J.R. et al. (2017). Fabrication of carboxymethyl chitosan–
hemicellulose resin for adsorptive removal of heavy metals from wastewater. Chinese
Chemical Letters 28: 625–632.
Zhang, Y., Yuan, Z., Mahmood, N. et al. (2016). Sustainable bio‐phenol‐ hydroxymethylfurfural
resins using phenolated de‐polymerized hydrolysis lignin and their application in bio‐
composites. Industrial Crops and Products 79: 84–90.
159
Sustainable Production of Biosurfactants and

Their Applications
Paulo Ricardo Franco Marcelino, Fernanda Gonçalves, Itzcóatl Muñoz Jimenez,
Bruna Curry Carneiro, Bruno Bosquiroli Santos, and Silvio Silvério da Silva
Department of Biotechnology, Engineering School of Lorena (EEL), University of São Paulo (USP), Lorena, São Paulo, Brazil
8.1 Introduction
Surfactants are molecules with amphipathic structures (one polar and one apolar region)
and for this reason, they are responsible for two extremely important phenomena: the
reduction of surface and interfacial tensions and emulsification of liquids with different
polarities. The development of modern society is totally dependent on these substances
since they are applied in distinct industrial sectors, from the production of agro‐chemicals
to formulations in the pharmaceutical industry, such as sulfonated olefins, ethoxylated
alkyl phenols, ethoxylated alcohols, alcohol sulfates, alkylbenzene sulfonates, and salts of
fatty acids (soaps). Despite the requirements for these products, synthetic surfactants have
been the subject of criticism, especially in by environmental lobby, since they have low
biodegradability and high toxicity and their synthesis is dependent on petroleum and its
derivatives. Generally, these synthetic products have long carbonic chains, branched or
aromatic groups that hinder biodegradation and lead to numerous environmental prob-
lems, such as eutrophication of rivers and lakes, increase of phosphate concentration, solu-
bilization of toxic organic compounds, and formation of foam (Marchant and Banat 2012;
Kosaric and Vardar‐Sukan 2015). Figure 8.1 represents the schematic structure of the dif-
ferent regions present in biosurfactants.
To overcome the environmental problems caused by synthetic surfactants, production of
biological surfactants, or biosurfactants, using enzymatic and microbiological pathways has
attracted a great deal of attention all over the world. The concept of biosurfactants has become
more popular nowadays which can be clearly seen from the increase in the publication of
scientific papers in the last two decades (Figure 8.2). The eco‐friendly and superior physico-
chemical characteristics of biosurfactants, such as high biodegradability, low toxicity, envi-
ronmental compatibility, possibility of production from agro‐industrial by‐products, higher
foaming, specific activity under extreme conditions such as temperature, pH, and salinity,
make them potential substitutes for synthetic surfactants (Sudhanshu et al. 2015).

Presence of apolar groups
In synthetics: hydrocarbons of long

Presence of polar groups chain.
In synthetics: sulfates, sulfonates, In naturals: long chain fatty acids and

salts derived from organic nitrogen triterpenic or steroidal chains.
compounds, carboxyl and others.
In naturals: carbohydrates, amino

acids, peptides, proteins, phosphate
groups and others.
Figure 8.1 Amphipathic structures of surfactants.
500
450
Number of published articles
400
350
300
250
200
150
100
50
0
00
02
20 3
20 4
20 5
20 6
20 7
20 8
09
01
12
13
14
15
16
17
18
10
11
0
0
0
0
0
0
20
20
20
20
20
20
20
20
20
20
20
20
20
Year
Figure 8.2 Number of articles published on biosurfactants in the period between 2000 and 2018
Source: Elsevier’s Scopus (2018).
Biosurfactants are synthesized by plants (saponins), microorganisms (biosurfactants)

and also by higher organisms such as vertebrates (bile salts) (Nitschke and Pastore 2002).
Among the biological surfactants mentioned, the microbial origin has been highlighted in
the market and in academic research. Biosurfactants produced by microorganisms (bacte-
ria, yeasts, and filamentous fungi) are generally classified according to their chemical
composition.
Sustainable Production of Biosurfactants and Their Applications 161
For the production of biosurfactants, carbon and nitrogen sources are used in the culture
medium. The main sources of carbon are carbohydrates and lipids. With the arrival of the
concept of biorefinery, many renewable sources are being used as raw material in the pro-
duction processes. More specifically, lignocellulosic sources such as sugarcane bagasse
have been used. Some studies report the use of lignocellulosic biomass as a source of car-
bon in the production of biosurfactants, for example the use of a hemicellulosic hydrolyz-
ate of sugarcane bagasse as a source of carbon for the production of glycolipids (Marcelino
et al. 2017). In this chapter, we present the basic concepts of biosurfactants, their synthesis,
raw materials, industrial applications, and sustainable production in the context of ligno-
cellulosic biorefineries.
8.2 Biosurfactants: Types, Structures, and Functions
The major classes of biosurfactants include glycolipids, lipopeptides/lipoproteins, phos-

pholipids/fatty acids, polymeric surfactants, and particulates (Table 8.1).
Among microbial biosurfactants, the glycolipid and lipopeptide/lipoprotein groups are
well known (Figure 8.3). In the glycolipid group, rhamnolipids (rhamnose lipids) (RMLs),
sophorolipids (sophorose lipids) (SPLs), and mannosylerythritol lipids (MELs) are the
most common, whereas among the lipopeptide/lipoprotein group, surfactin is the most
popular.
The RMLs are glycolipids that contain rhamnose (a monosaccharide of the pentose
type – 6‐deoxy‐L‐mannose) linked to a fatty acid tail, generally β‐hydroxydecanoic acid
(C10) (Cortés‐Sánchez et al. 2013). The structures of these biosurfactants can be classified
according to the number of rhamnose groups: monorhamnolipids (one rhamnose group in
the structure) and dirhamnolipids (two rhamnose groups in the structure). The RMLs are
the best known biosurfactants in the world but their applications still have some restric-
tions, since they are obtained by pathogenic bacteria (Pseudomonas aeruginosa). RMLs are
commonly applied in environmental processes such as bioremediation of soils and water
contaminated by petroleum, metals, and other xenobiotic compounds (Roy et al. 2015; Liu
et al. 2018a). They are also used in food processing, protein folding, microbial fuel cells, and
nanoparticle synthesis (Andersen and Otzen 2014; Zheng et al. 2015; Kiran et al. 2016).
Sophorolipids are another type of glycolipidic biosurfactants that contain sophorose (a
disaccharide with two glucose residues connected by a glycosidic linkage β‐1,2) linked to a
chain of hydroxylated fatty acids, generally hydroxy‐hexadecanoic (C16:0, C16:1 and C16:2)
and hydroxy‐octadecanoic (C18:0, C18:1 and C18:2) acids (Kurtzman et al. 2010; Ma et al. 2011;
Cortés‐Sánchez et al. 2013). These compounds are structurally classified as acidic (having
a carboxyl group in their structure) and lactonic (having a cyclic ester in their structure). In
industry, they are used in products for beauty and personal care (bath products, acne pads,
antidandruff products, pencil‐shaped lip rouge, lip cream, lipsticks, and toothpaste),
household cleaning products, and biopesticides (Shete et al. 2006).
Mannosylerythritol lipids are also glycolipidic biosurfactants, generally mannose and
erythritol (4‐O‐β‐D‐mannopyranosyl‐meso‐erythritol) linked by acylation to fatty acids of
different hydrocarbon chains (Cortés‐Sánchez et al. 2013). They are classified according to
the degree of acetylation (mono‐, di‐ or triacetylated) and the type of polyol found in the
Table 8.1 Major classes of biosurfactants and their composition and properties.
Biosurfactant Composition and characteristics Examples
Glycolipids ●● Composed of carbohydrates Rhamnolipids,

(monosaccharides, oligosaccharides, and sophorolipids, cellobiose
polyols) and lipids (fatty acids) lipids, mannosylerythritol
●● Present ether or ester linkages connecting lipids, trehalose lipids, and
glycidic to lipid moiety liamocins (polyol lipids)
●● Exhibit low molecular mass (628–826 Da)
●● Critical micellar concentration (CMC)
between 20 and 366 mg/L (Siñeriz et al.
2009; Kulakovskaya and Kulakovskaya 2014)
●● Reduce water surface tension from 72 to
20 mN/m
Lipopeptides/ ●● Composed of peptides or proteins and lipids Iturin, surfactin, fengycin,
lipoproteins ●● Usually present between 7 and 10 amino viscosin, amphisin, and
acid residues linked to a hydrophobic fatty putisolvin
acid structure (Smyth et al. 2010)
●● Exhibit higher molecular mass (≥1000 Da)
●● CMC around 10 μmol/L or 23 mg/L
●● Water surface tension reduction from 72 to
20 mN/m (Wu et al. 2017)
Phospholipids/ ●● Composed of phosphate group linked to Phosphatidylcholine,
fatty acids fatty acid chains of 5–12 carbon atoms phosphatidylethanolamines
●● CMC varying from 58.9 mM to 0.007 mM (PEs), and
●● Reduction of water surface tension from 72 phosphatidylglycerols
to 21 mN/m (Weschayanwiwat et al. 2005)
Polymerics ●● The composition of these biosurfactant types Alasan, liposan,
is complex with varied structures lipomannan, emulsan, and
(heteropolysaccharides) other polysaccharide–
●● Exhibit higher molecular mass (≥1000 Da) protein complexes
●● Reduction of water surface tension from 72
to 30 mN/m (Amoabediny et al. 2010;
Shekhar et al. 2014)
Particulates ●● Vesicles of proteins, phospholipids, and Extracellular membrane
lipopolysaccharides with size between 20 vesicles
and 50 nm (Kappeli and Finnerty 1979;
Santos et al. 2016b)
structure (erythritol, mannitol, ribitol or arabitol) (Morita et al. 2015). Unlike SPLs, MELs
biosurfactants are mainly applied in the formulation of beauty products and personal care.
Among the lipopeptide group, surfactin is the most popular biosurfactant. It is an amphi-
pathic cyclic heptapeptide that can be formed by L‐glutamine, L‐leucine, D‐leucine, L‐
valine, L‐asparagine, D‐leucine, and L‐leucine residues linked by peptide bonds and a fatty
acid chain with 13–16 carbons, resulting in a molecular weight of 1036 Da (Vedaraman and
Venkatesh 2011; Wu et al. 2017). Surfactin has potential biomedical applications due to its
antimycoplasmal, antiviral, antibacterial, and antitumoral activities. In addition, it induces
Mannosylerythritol lipid
OH H3C
CH3
HO
O O CH3
H3C n OH O
O H2N O
OAc O HO N
N
n O H3C NH H HN CH3
O
OH R NH H HN NH2
O HO
O CH3
O O O H3C O O
AcO O O
NH NH NH
O O H3CO
O NH H
n = 6 - 10 NH CH3
H H3C H N
N N H2N N
Rhamnolipid HO O O O CH3
OH H3C OH
O OH O
O
H3C O
O O OH
NH2
OH CH3
HO O O Surfactin
O O Iturin NH2
HO
H3C OH
CH3 OH O O O
HO H H H
HO N N N
N N
H H
CH3 CH3 O O O
O NH
O
Sophorolipids O OH H3C
O
N
HO HO O O
O CH3 O O HN
HO HO H H
HO O HO O N
HO H3C N
O HO O H
O O H3C O O
O HO H3C
HO HO H2N
OH OH HO
C Fengycin
O
HOOC
Lactonic form Acidic form
Figure 8.3 Structures of the main microbial biosurfactants.

formation of ion channels in lipid bilayer membranes, has antiinflammatory activity, and
inhibits biofilm formation due to its antiadhesive properties. This biosurfactant is also
commonly used for environmental bioremediation, enhanced hydrocarbon biodegrada-
tion, heavy oil transportation, and biocontrol (Chen et al. 2015; Wu et al. 2017).
In addition to these biosurfactants, obtained naturally or by chemical synthesis, there are
those that may be obtained by enzymatic/chemical transformation in a preexisting struc-
ture, called semi‐synthetic or modified biosurfactants (Recke et al. 2013). These modifica-
tions not only allow the structural change of the molecule but also help in the modification
of their physicochemical and biological properties. Lang and Wagner (1987) and Zinjarde
and Ghosh (2010) reported the important role of enzymatic systems, such as lipases and
glycosidases, in the structural modification of biosurfactant precursors, like carbohydrates
and lipids. Delbeke and collaborators (2015) synthesized a quaternary ammonium
sophorolipid from a sophorolactone produced by fermentation; organic reactions were per-
formed in order to insert an amine group into the biosurfactant molecule. The modified
sophorolipid obtained was tested against pathogenic bacteria, proving to have excellent
antibacterial properties.
8.3 Biosurfactant Production by Fermentation:

Microorganisms, Raw Materials, Substrates, Metabolism,
Ways of Conducting the Process, and Bioreactors
The biosurfactants are applied in several industrial sectors, but the reasons why the micro-
organisms produce these compounds are still unknown. In many cases, biosurfactants
have physiologic functions such as emulsification, solubilization and intracellular trans-
port of aqueous insoluble compounds, cell release in biofilms, antimicrobial activity, and
quorum sensing (Nitschke and Pastore 2002).
Currently, biosurfactants are produced by fermentation processes, both in laboratories
and in industries. The success of the production of these bioproducts depends on the
microorganism chosen as the fermenting agent, the culture medium, and the way of con-
ducting the process, as described below.
8.3.1 Microorganisms
Bacteria and yeasts are the microorganisms generally used for the production of biosur-
factant. The bacterial members of the genus Bacillus (B. subtilis, B. mojavensis, B. pumilus, B.
brevis, B. salmalaya, B. licheniformis, and B. amyloliquefaciens) (Mulligan et al. 2001;
Fernandes et al. 2007; Sudhanshu et al. 2015), Pseudomonas genus (P. aeruginosa, P. fluores-
cens, P. chlororaphis, P. marginalis, and P. maltophilia) (Desai and Banat 1997; Jadhav et al.
2011), Thiobacillus thiooxidans (Christofi and Ivshina 2002), Gluconobacter spp., Paenibacillus
polymyxa, Flavobacterium spp. (Bodour et al. 2004), Rhodococcus erythropolis (Christofi and
Ivshina 2002), Brevibacterium brevis, Agrobacterium spp., Acinetobacter calcoaceticus (Choi
et al. 1996), Leuconostoc mesenteroides and Lactobacillus fermentum are responsible for the
production of lipopeptides, glycolipids, polymeric surfactants, fatty acids, phospholipids,
neutral lipids, and particulate biosurfactants (Sudhanshu et al. 2015; Sarwar et al. 2018).
Although bacteria are well reported as biosurfactants producers, some authors consider
yeasts the best fermentation agents to produce these biomolecules. According to Monteiro
et al. (2009), when compared to bacteria, yeasts have cell structures that are more resistant
to the biosurfactants secreted in the culture medium. In addition to cellular resistance, the
yeasts used in biosurfactant production in majority have GRAS status (Generally Regarded
As Safe), eliminating risks of toxicity and pathogenicity and allowing applications in the
food and pharmaceutical industries without restrictions (Barth and Gaillardin 1997; Fontes
et al. 2008). The most used yeasts in biosurfactant production include members of the
genus Candida (C. bombicola) (Gobbert et al. 1984; Felse et al. 2007), C. utilis (Shepherd
et al. 1995), C. ingens (Amezcua‐Vega et al. 2007), C. batistae, C. stellata, C. bogoriensis, C.
riodocensis, C. tropicalis, C. apicola (Hommel et al. 1994), C. lipolytica (Cirigliano and
Carman 1984) and C. antarctica (Kitamoto et al. 1992), Starmerella (S. bombicola)
(Marcelino et al. 2019) and Pseudozyma (P. rugulosa, P. aphidis, P. siamensis, P. fusiformata,
P. parantarctica, and P. tsukubaensis). However, in the literature there are reports of biosur-
factant production by Rhodotorula, Pichia, Debaromyces, Aureobasidium (Brumano et al.
2017), Kluyveromyces, Issatchenkia, Cryptococcus, Yarrowia lipolytica (Zinjarde et al. 1997;
Beopoulos et al. 2009), Trichosporon (Christofi and Ivshina 2002; Das et al. 2008;
Kulakovskaya et al. 2010), Cutaneotrichosporon, and Ustillago (Christofi and Ivshina 2002).
It should be noted that, recently, the yeasts described as oleaginous (accumulators of intra-
cellular lipids) and positive lipase yeasts have been highlighted as the best biosurfactant
producers.
In addition to bacteria and yeasts, there are some reports in the literature of filamentous
fungi (Penicillium chrysogenum, P. spiculisporum and Aspergillus versicolor) and microalgae
(Porphiridium cruentum, Spirulina platensis, S. maxima, Dunaliella bardawil, Chlorella
minutissima, Phaeodactylum tricornatum, Cylindrotheca closterium, Stichococcus bacillaris
and Scenedesmus spp.) being used for biosurfactant production (Szuhaj 1989; Shepherd
et al. 1995; Bigogno et al. 2002; Xue et al. 2002; Lin et al. 2003; Gao et al. 2011).
8.3.2 Raw Materials, Substrates, Metabolism, and Culture Conditions

After microorganism selection and a primary study on its physiology, it is necessary to for-
mulate the culture medium. The selection of carbon and nitrogen sources is extremely
important for the fermentative process of biosurfactant production.
The main carbon sources used in biosurfactant production are carbohydrates and lipids.
However, some studies, mainly those focused on bioremediation, also use hydrocarbons.
Raw materials such as industrial and agro‐industrial by‐products can be excellent sugar
and lipid sources for biosurfactant production. Some studies report the use of agro‐indus-
trial waste such as sugarcane bagasse, wheat straw, cassava bagasse, cassava flour wastewa-
ter, rice straw, soy hull, corn and sugarcane molasses, potato, sweet potato, soybean, sweet
sugar beet and sorghum residues, crude glycerol from biodiesel refinery, and animal fat
(Nitschke and Pastore 2003; Nitschke et al. 2004; Moldes et al. 2007; Portilla‐Rivera et al.
2007; Joshi et al. 2008; Moldes et al. 2013; Marcelino et al. 2017; Marcelino et al. 2019; de
Souza Monteiro et al. 2012; Sousa et al. 2014; Walaa et al. 2016; Makkar et al. 2011).
Other residues that can also be used as raw materials include those from coffee process-
ing (coffee pulp, coffee husks), dairy (curd whey, cheese whey, whey waste), distillery, food
processing (frying edible oils and fats, olive oil, potato peels, rape seed oil, sunflower, veg-
etable oils), fruit processing (banana waste, apple and grape pomace, carrot industrial
waste, pineapple) and oil processing mills (coconut cake, canola meal, olive oil mill waste
water, palm oil mill waste water, peanut cake effluent, soybean cake, soap stock, waste from
lubricating oil) (Fox and Bala 2000; Thompson et al. 2000; Thompson et al. 2001; Das and
Mukherjee 2007; Colak and Kahraman 2013; Makkar et al. 2011; Thavasi et al. 2007; Banat
et al. 2014; Felix et al. 2019).
Sugars and lipids used as substrates by the microorganisms are directed to the metabolic
pathways responsible for cell growth and biosurfactant biosynthesis. Studies have reported
that hydrophobic substrates act as inducers in biosynthesis of microbial surfactants. Thus,
oils and hydrocarbons can be used strategically in the production of these bioproducts and
even regulate the anabolic pathways (Desai and Desai 1993). Other studies showed satisfac-
tory yields in biosurfactant production when carbon sources with different polarities were
mixed (Fontes et al. 2008). It should be noted that in the microbial synthesis of biosur-
factants there are complex processes that require integration of metabolic pathways, since
intermediates and products of catabolic pathways, such as those obtained in glycolysis,
pentoses‐phosphate pathway, amino acid degradation and β‐oxidation of fatty acids, will be
the precursors of biosurfactant synthesis reactions.
In biosurfactant biosynthesis, four possibilities are accepted (Siñeriz et al. 2009).
1) Hydrophilic and hydrophobic moieties are synthesized de novo via independent
pathways.
2) The hydrophilic moiety is synthesized de novo and the substrate induces the hydropho-
bic moiety.
3) The synthesis of the hydrophilic moiety is substrate dependent while the hydrophobic
moiety is synthesized de novo.
4) The synthesis of both residues depends on the carbon substrate used.
The carbon source used in biosurfactant production is of extreme importance because it
influences the structure of the molecule and consequently the physicochemical properties
(Santos et al. 2016a). Besides the carbon sources, nitrogen sources are essential for biosur-
factant production and nucleic acids, amino acids, and protein/enzyme biosynthesis are
needed for cellular metabolism. Several compounds can be used as nitrogen sources in the
production of biosurfactants, such as corn steep liquor, urea, peptone, yeast extract, meat
extract, malt extract, rice bran extract, soybean meal extract, ammonium sulfate, ammo-
nium nitrate, and sodium nitrate (Marcelino et al. 2017). The nitrogen source commonly
used in biosurfactant production is yeast extract and its concentrations vary according to
the culture medium and microorganism (Fontes et al. 2008).
In the selection of nitrogen source for biosurfactant production, it is recommended to
use organic compounds with a complex constitution, since their consumption does not
cause large changes in pH. The use of inorganic salts can cause hydrolysis of cations or
anions and alter the pH of the culture medium, thus impairing the fermentation process.
In biosurfactant production, the C:N ratio is generally high. Thus, the nitrogen source is
used strategically as a limiting factor in this bioprocess and for biosynthesis regulation.
According to Albrecht et al. (1996), high C:N stresses the microbial cells and causes a
decrease in the activity of the enzyme isocitrate dehydrogenase, responsible for isocitrate
oxidation to α‐ketoglutarate in the Krebs cycle. Thus, there is isocitrate and citrate accumu-
lation in the mitochondria and both are transported to the cytosol, where the citrate is
cleaved by the enzyme citrate lyase, originating acetyl‐CoA, the precursor metabolite of
fatty acids, increasing biosurfactant production.
In addition to carbon and nitrogen sources, micronutrients are also required in culture
media to obtain biosurfactants. Generally, multivalent cations act as co‐factors in various
enzymatic reactions and may favor biosurfactant biosynthesis (Fontes et al. 2008). Iron
(Fe2+) limitation stimulates biosurfactant production in P. aeruginosa (Guerra‐Santos et al.
1986; Ramana and Karanth 1989). Recently, Niu et al. (2017) reported the significative
influence of manganese (Mn2+) and calcium (Ca2+) cations in MEL production by the fun-
gal endophyte Ceriporia lacerate.
Physicochemical properties and operational factors such as temperature, pH, agitation,
and aeration can also interfere with biosurfactant production. Temperature and pH may
impair the enzymatic activities of microorganisms used as biosurfactant producers.
Generally, these microorganisms produce biosurfactants in temperatures between 25 and
40 °C. Kitamoto et al. (1992) demonstrated that C. antarctica increased MEL production
when grown at 25 °C. Recently, Cryptococcus, Rhodotorula, Candida, and Tremella yeasts,
isolated from the Antarctic continent, have been reported to produce biosurfactants at
25 °C (Chaves 2017). The P. aeruginosa strain 44 T1 produced RMLs at 37 °C (Robert et al.
1989). Perfumo and collaborators (2018) reported that several species of psychrophilic bac-
teria and yeasts are capable of producing biosurfactants in a temperature range between 4
and 35 °C. As for the effect of pH, RML production by Pseudomonas spp. was highest when
the pH was maintained between 6.0 and 6.5; above pH 7.0, production decreased to low
levels (Guerra‐Santos et al. 1984). According to van Bogaert et al. (2011), in the case of
sophorolipids produced by yeasts, pH values between 7 and 7.5 can disorganize their
structure.
Agitation and aeration are important parameters in biosurfactant production since,
beyond culture medium homogenization, they also supply the oxygen demand, being
directly related to the rate of oxygen transfer in culture medium (represented by the volu-
metric coefficient of oxygen transfer – kLa). The kLa is important for definition of the
metabolic pathways adopted by the cell, favoring cell growth, through the respiratory
chain, or catabolic and anabolic pathways of interest for biosurfactant production.
According to Yeh et al. (2006), biosurfactant production by yeasts increased when the agita-
tion and aeration rates were high. However, vigorous aeration and agitation during fermen-
tation may cause problems with foaming (Joshi et al. 2013).
8.3.3 Types of Fermentations and Bioreactors Involved in Biosurfactant

Production
In addition to studies of the nutritional, physicochemical, and operational factors, optimi-
zation of biosurfactant production is also dependent on the ways of conducting the bio-
process and bioreactor configurations. Most biosurfactant production studies conduct
processes in submerged fermentations in batch and fed batch modes (Trummler et al. 2003,
Rau et al. 2005; Fontes et al. 2008; Müller et al. 2010; Zhu et al. 2012; Maddikeri et al. 2015).
With the need for industrial biosurfactant production, studies with bioreactors have been
performed on bench and pilot scale (BBEPP 2015; Coppe 2009). Submerged fermentations
in stirred tank reactors (STR) are the most popular way of obtaining biosurfactants
(Table 8.2).
Although STRs are the most common reactors used in biosurfactant production by sub-
merged fermentation, in this kind of reactor there is the possibility of foam formation
which can be a major problem (Figure 8.4). The excess of foam during fermentation is a
limiting factor for biosurfactant production by the submerged process since, together with
the foam, there is loss of nutrients, products, and biomass, reducing productivity or, in
extreme cases, making fermentation unfeasible (Winterburn and Martin 2012).
In order to mitigate foaming in submerged processes of biosurfactant production in STR
bioreactors, strategies such as the use of mechanical, acoustic and ultrasonic methods,
“switchable foam control” (using the pH sensitivity of certain biosurfactants), membranes
and foam fractionation are adopted (Stocks et al. 2005; Winterburn and Martin 2008, 2012;
Pereira et al. 2013). In addition, solid‐state fermentation (SSF) has also been used in biosur-
factant production in order to avoid foaming, due to its simplicity and cost. SSF can be
defined as the growth of a microorganism on a solid substrate, with a minimum amount of
free water in the system. In other words, the water forms a thin layer on the substrate sur-
face (Pandey 2003; Pinto et al. 2005). Therefore, the water retained in the matrix must be
enough to allow the growth and metabolism of microorganisms. However, it cannot affect
the matrix maximum retention capacity (Muso‐Cachumba 2017).
Table 8.2 Types of bioreactors used in the production of biosurfactants.
Biosurfactant Production
Reactor Microorganism type (g/L) References
Stirred‐tank Candida lipolytica Glycolipid 40 Santos et al.

bioreactor (2016b)
Stirred‐tank Candida sphaerica Glycolipid 21.49 Luna et al.
bioreactor UCP 0995 (2014)
Stirred‐tank Starmerella Glycolipid 342 Liu et al.
bioreactor bombicola ATCC (2018b)
22214
Three‐phase inverse Bacillus subtilis Lipopeptide 58 Fahim et al.
fluidized bed ATCC 21332 (2013)
Stirred‐tank Bacillus pumilus 2lR Lipopeptide 1.06 Fooladi et al.
bioreactor (2018)
Stirred‐tank Bacillus subtilis Lipopeptide 335.77 Chenikher
bioreactor ATCC 6633 et al. (2010)
Stirred‐tank Rhodococcus sp. Glycolipid 2.7 Bages et al.
bioreactor PML 026 (2018)
Stirred‐tank Pseudomonas Glycolipid 15.9 Benincasa
bioreactor aeruginosa et al. (2002)
Raimbault column Bacillus pumilus Lipopeptide 0.808 Slivinski
bioreactor UFPEDA 448 et al. (2012)
Figure 8.4 Foam formation in biosurfactant production by Cutaneotrichosporon mucoides during

batch fermentation in a STR reactor.
In SSF, microorganism growth occurs in a similar way to its natural habitat which leads
to better adaptability and higher metabolite production. Moreover, inhibition caused by
substrate is not likely to occur. Another relevant fact is that agro‐industry by‐products can
be used as substrate in SSF, which reduces production costs, lowering the price of the final
product and also avoiding environmental problems (Durand 2003; Ashok et al. 2017; Soccol
et al. 2017). Apart from this, the costs involved in recovery will be lower, due to more con-
centrated products, and there will be decreased generation of effluents (Pandey 2003;
Krishna 2005; Rodríguez‐Couto and Sanromán 2005).
Most recent work regarding SSF focuses on the production of enzymes, mainly cellu-
lases and xylanases (Soccol et al. 2017). However, this is a promising method for biosur-
factant production, because it can solve the foam formation problem, a great advantage
over submerged fermentation (Pandey et al. 2000; Pinto et al. 2005; Jiménez‐Peñalver
et al. 2018). Different studies have shown the potential of biosurfactant production in
SSF. Slivinski et al. (2012) described production of surfactin with yield of 809 mg/L by
employing B. pumilus using a mixture of okra and sugarcane bagasse as a solid matrix in
a column reactor with forced aeration. Ghribi et al. (2012a) reported biosurfactant pro-
duction of 20.8 mg/g using millet as substrate and B. subtilis as fermentative agent in
flasks. Moreover, Camilios‐Neto et al. (2011) studied rhamnolipid production by P. aer-
uginosa using a mixture of sugarcane bagasse and corn bran as substrate, yielding 45 g/L
of biosurfactant.
For yeasts, Jiménez‐Peñalver et al. (2018) tested biosurfactant production in cylindrical

polyvinylchloride packed‐bed bioreactors by S. bombicola using polyurethane foam as inert
support and stearic acid and molasses as carbon sources, yielding 0.211 g sophorolipids/g
substrate. Brumano et al. (2017) reported biosurfactant yield of 1.8 g/L employing sugar-
cane bagasse as a solid matrix, sucrose as carbon source and the yeast Aureobasidium pul-
lulans. Velioglu and Urek (2016) applied sunflower seed as substrate for the fungus
Pleurotus djamor, yielding 10.2 g/L of biosurfactant. This suggests that biosurfactant pro-
duction in SSF may vary according to the microorganism and substrate employed.
8.4 Biosurfactant Production in Biorefineries:

Use of Agro‐Industrial By‐Products as Raw Materials
Recently, the European Commission has enforced the idea of a “circular economy” that
aims to prevent waste generation or, when this cannot be avoided, that waste should be
used for bioeconomical purposes (Satpute et al. 2017). Considering this background, in the
last few years, with the arrival of the biorefinery concept, a broad range of products have
been developed using renewable sources as raw material, more specifically from lignocel-
lulosic sources like sugarcane bagasse and various sorts of brans.
In Brazil, for example, there is a vast sugarcane bagasse surplus, that is not used by the
sugar mills for energy production. Therefore, researchers like Marcelino et al. (2017) have
focused on converting this biomass into biosurfactants. These authors used hemicellulosic
sugarcane bagasse hydrolyzate as a raw material for glycolipid production by Scheffersomyces
stipitis NRRL Y‐7124, obtaining an emulsifying index of 70%, and evaluated its larvicidal
effect against Aedes aegypti.
Chen et al. (2019) applied xylose‐rich corncob hydrolyzate as substrate for surfactin pro-
duction by B. subtilis BS‐37, achieving 0.523 g/L and, beyond that, the microorganism was
able to tolerate several inhibitory compounds from the hydrolyzate. Samad et al. (2014)
reported sophorolipid production (3.6 g/L) by the microorganism Candida (Starmerella)
bombicola growing on hydrolyzates from sweet sorghum bagasse. The same authors
reported that addition of soybean oil can promisingly increase production to 84.6 g/L.
Mercadé et al. (1993) analyzed rhamnolipid production by Pseudomonas sp. JAMM using
olive oil mill effluent as the carbon source in a 2‐litre bioreactor with yield of 1.4 g/L.
Monteiro et al. (2007) also evaluated rhamnolipid production, obtaining a yield of 3.9 g/L
using P. aeruginosa DAUPE 614 as fermenting agent and glycerol as substrate. Another
study showed that P. aeruginosa was able to produce 3.2 g/L of rhamnolipids using corn
steep liquor (10% [v/v]) and molasses (10% [w/v]) as culture medium (Gudiña et al. 2015).
Kim et al. (2005) employed soybean dark oil as a carbon source for Candida bambicola (ATCC
22214); the microorganism produced 90 g/L of sophorolipid. Thavasi et al. (2009) studied the
conversion of peanut oil cake by Azotobacter chroococcum that yielded 4.6 g/L of lipopeptide.
8.5 Markets for Biosurfactants
Biosurfactants are considered versatile molecules due to their physicochemical and bio-
logical properties, guaranteeing a wide range of industrial applications. These bioproducts
can be used in the oil, chemical, food, pharmaceutical, and agricultural industries, among
Table 8.3 Main companies producing biosurfactants.
Companies Country Products
TeeGene Biotech United Kingdom RMLs and lipopeptides

AGAE Technologies LLC United States RMLs
Jeneil Biosurfactant Co. LLC United States RMLs
Paradigm Biomedical Inc. United States RMLs
Rhamnolipids Companies, Inc. United States RMLs
Fraunhofer IGB Germany SPLs, cellobiose lipids, and MELs
Saraya Co. Ltd. Japan SPLs (Sophoron®)
Ecover Belgium Belgium SPLs
Groupe Soliance France SPLs
MG Intobio Co. Ltd. South Korea SPLs (Sopholine®)
Synthezyme LLC. United States SPLs
Allied Carbon Solutions (ACS) Ltd. Japan SPLs
Henkel Germany SPLs, RMLs, and MELs
Kaneka Co. Japan SPLs
Evonik Germany SPLs and RMLs
Source: Sekhon Randhawa and Rahman (2014).

MEL, mannosylerythritol lipid; RML, rhamnose lipid; SPL, sophorose lipid.
others (van Dyke et al. 1991; Nitschke and Pastore 2002; Nitschke and Costa 2007; Lourith
and Kanlayavattanakul 2009; Gharaei‐Fathabad 2011; Campos et al. 2013; Sachdev and
Cameotra 2013; Silva et al. 2015).
The market for biosurfactants is constantly expanding. According to forecasts, up to 2020
a 4.3% increase will occur in production, reaching 462 000 tons. and approximately 2.2 bil-
lion dollars will be collected. Developed countries, such as Europe and North America,
account for 75–80% of global consumption of biosurfactants, while the emerging countries
and the Asia‐Pacific region have 20–25% consumption (GVR 2015). Table 8.3 shows the
main companies producing biosurfactants around the world.
8.6 Applications of Biosurfactants
Biosurfactants are reported to have a wide range of applications in various fields, including
bioremediation, oil recovery, medicine, cosmetic, food industries, biorefineries, etc. All
these are briefly discussed below.
8.6.1 Bioremediation
One of the most studied applications of biosurfactants is bioremediation of water and soil. It
has been noted that these molecules enhance the contaminant removal rate compared to
normal methods that do not add them, reaching high rates within 24 hours (Silva et al. 2018).
This is possible due to an increase of bioavailability of hydrocarbons, organic and inorganic

compounds. Two different mechanisms have been elucidated: increasing the solubility of the
pollutant or interaction with the cell membrane, altering its hydrophobicity and facilitating
substrate entry into cytoplasm (Płociniczak et al. 2011).
8.6.2 Oil Recovery

In addition to increased bioavailability, a wide variety of other processes can benefit from
biosurfactant properties, especially in the petroleum industry (Shekhar et al. 2014). The
recovery of polluted soils may require washing before bioremediation. The addition of bio-
surfactants to this step effectively removes hydrocarbons, such as polycyclic aromatics, and
apolar organic compounds from the soil, due to their ability to increase mobility and solu-
bility (Lai et al. 2009; Xiaohong et al. 2009). Chemical enhanced oil recovery (EOR) also
uses biosurfactants to improve the efficiency of crude oil recovery.
8.6.3 Antibiofilms and Antimicrobials

Contamination and microbial resistance are of major concern in the food processing and
biomedical fields. Biofilms are known to protect bacteria from antibiotics and other lethal
agents, also making it difficult to clean equipment (Stewart 2002; Sharma and Saharan
2016). By altering the hydrophobicity of a surface, biosurfactants can prevent biofilm for-
mation and cell adherence, as well as causing biofilm disruption where it has already
grown (Sharma and Saharan 2016; Karlapudi et al. 2018). Natural surfactants may also
display antimicrobial properties depending on the producing organism and the growing
conditions, which lead to different types of products. Biosurfactants have been found to
present antibacterial, antifungal, antitumoral, and antiviral effects (Cameotra and Makkar
2004; Garg and Chatterjee 2018; Karlapudi et al. 2018).
8.6.4 Medicinal Applications

Biosurfactants also have several applications in the medical field, including antifilm coat-
ing of medical materials, immune‐modulator agents, and enzyme inhibitors. These bio-
molecules are seen as an alternative to synthetic medicines, although their effect on human
cells needs further research (Rodrigues et al. 2006). The need to safely and efficiently intro-
duce an exogenous gene into an animal cell has led scientists to use cationic liposomes as
one possible method. Biosurfactant‐based liposomes have shown increased efficiency com-
pared to cationic ones (Gharaei‐Fathabad 2011). In addition, biosurfactants could possibly
be used as drug transport agents, supplements of pulmonary surfactant, and adjuvants for
vaccines (Rodrigues et al. 2006).
8.6.5 Cosmetics
Modern consumers are giving preference to products containing natural components.
Therefore, the cosmetic industry has given special attention to replacing synthetic sur-
factants with microbial ones. Their applications are extensive in this field, as they can act
as detergents, emulsifiers, foaming, wetting and dispersion agents (Ferreira et al. 2017;
Vecino et al. 2017). The addition of biosurfactant to sunscreen formulations containing
mica greatly increases the sunscreen protection factor (SPF) in comparison to those with-
out this biomolecule (Rincón‐Fontán et al. 2018).
8.6.6 Food and Beverage Industry

In the food and beverage industry, biosurfactants can be applied as emulsification and dee-
mulsification agents. Moreover, they can be used to improve stability, consistency, conserva-
tion, and texture of a vast range of foods (Nitschke and Costa 2007; Muthusamy et al. 2008).
8.6.7 Insecticides
Agriculture also benefits from biosurfactants. Apart from soil remediation, the application
of these biomolecules increases the bioavailability of nutrients for plant‐associated micro-
organisms and eliminates phytopathogens, which enhances soil quality and productivity.
Biosurfactants can act as substitutes for other surfactants which are commonly added to
pesticides, fungicides, and herbicides (Sachdev and Cameotra 2013). Furthermore, biosur-
factants themselves have been described with insecticidal properties. Many species from
the aphid superfamily and lepidopteran order have been reported as susceptible to these
molecules (Kim et al. 2011; Ghribi et al. 2012b; Mnif et al. 2013). As far as larvicidal activity
is concerned, biosurfactants have been reported to be efficient in eliminating A. aegypti
larvae, by desroying the waxy cuticle, and emerging as a green pesticide for vectors of
neglected tropical diseases such as dengue and zika (Marcelino et al. 2017).
8.6.8 Other Applications

A wide variety of novel and interesting applications of biosurfactants can be found both in
industry and the literature. Microbes isolated from polar environments produce molecules
able to perform at very low temperatures which have applications for ice–water interfaces, gas
hydrate exploitation, and biodiesel enhanced flow properties at low temperatures (Perfumo
et al. 2018). In the field of bioenergy, biosurfactants have been added to microbial fuel cells
(MFC) to improve electron transfer and electricity generation (Zheng et al. 2015; Zhang et al.
2017). These versatile molecules have been successfully applied in the mining and textile
industries, for paint removal and to enhance extraction of molecules such as antibiotics from
cells (Akbari et al. 2018; Martins and Martins 2018; Bastrzyka et al. 2019; Chuo et al. 2019).
8.7 Conclusion
The search for sustainable and eco‐friendly products has increased interest in the develop-
ment of biosurfactants. Although sugars are considered a good source of carbon, their pro-
duction potential is low. Therefore, it is necessary to continue the search for alternatives to
reduce the cost of production. In this context, the use of renewable raw materials such as
lignocellulosic biomass in the production of biosurfactants is very promising.
Acknowledgments
CAPES, CNPq and FAPESP (2015/06238‐4; 2016/10636‐8; 2016/14852‐7) for financial

support.
References
Akbari, S., Abdurahman, N.H., Yunus, R.M. et al. (2018). Biosurfactants – a new frontier for social
and environmental safety: a mini review. Biotechnology Research and Innovation 2: 81–90.
Albrecht, A., Rau, U., and Wagner, F. (1996). Initial steps of soforose lipid biosynthesis by
Candida bombicola ATCC 22214 grown on glucose. Applied Microbiology and Biotechnology
73: 46–67.
Amezcua‐Vega, C., Poggi‐Varaldo, H.M., Esparza‐Garcia, F. et al. (2007). Effect of culture
conditions on fatty acid composition of a biosurfactant produced by Candida ingens and
changes of surface tension of culture media. Bioresource Technology 98: 237–240.
Amoabediny, G., Rezvani, M., Rashedi, H. et al. (2010). Application of a novel method for
optimization of bioemulsan production in a miniaturized bioreactor. Bioresource Technology
101 (24): 9758–9764.
Andersen, K.K. and Otzen, D.E. (2014). Folding of outer membrane protein A in the anionic
biosurfactant rhamnolipid. FEBS Letters 588: 1955–1960.
Ashok, A., Doriya, K., Rao, D.R.M., and Kumar, D.S. (2017). Design of solid state bioreactor for
industrial applications: an overview to conventional bioreactors. Biocatalysis and
Agricultural Biotechnology 9: 11–18.
Bages, S., White, D.A., Winterburn, J.B. et al. (2018). Production and separation of a
trehalolipid biosurfactant. Biochemical Engineering Journal 139: 85–94.
Banat, I.M., Satpute, S.K., Cameotra, S.S. et al. (2014). Cost effective technologies and
renewable substrates for biosurfactants production. Frontiers in Microbiology 5 (697): 1–18.
Barth, G. and Gaillardin, C. (1997). Physiology and genetics of the dimorphic fungus Yarrowia
lipolytica. FEMS Microbiology Reviews 19 (4): 219–237.
Bastrzyka, A., Fiedot‐Tobolab, M., Polowczyksa, I. et al. (2019). Effect of a lipopeptide
biosurfactant on the precipitation of calcium carbonate. Colloids and Surfaces B:
Biointerfaces 174: 145–152.
Benincasa, M., Contiero, J., Manresa, M.A., and Moraes, I.O. (2002). Rhamnolipid production
by Pseudomonas aeruginosa LBI growing on soapstock as the sole carbon source. Journal of
Food Engineering 54: 283–288.
Beopoulos, A., Chardot, T., and Nicaud, J.M. (2009). Yarrowia lipolytica: a model and a tool to
understand the mechanisms implicated in lipid accumulation. Biochimie 91 (6): 692–696.
Bigogno, C., Khozin‐Goldberg, I., Boussiba, S. et al. (2002). Lipid and fatty acid composition of
the green oleaginous alga Parietochloris incisa, the richest plant source of arachidonic acid.
Phytochemistry 60 (5): 497–503.
Bio Base Europe Pilot Plant (2015). TeeGene Biotech Ltd (UK) successfully conducted
biosurfactant manufacturing process at Bio Base Europe pilot plant. www.bbeu.org/
pilotplant/eegene‐biotech‐ltd‐successfully‐conducted‐biosurfactant‐manufacturing‐
process‐at‐bio‐base‐europe‐pilot‐plant.
Bodour, A.A., Guerrero‐Barajas, C., Jiorle, B.V. et al. (2004). Structure and characterization of
flavolipids, a novel class of biosurfactants produced by Flavobacterium sp. strain MTN11.
Applied and Environmental Microbiology 70 (1): 114–120.
Brumano, L.P., Antunes, F.A.F., Souto, S.G. et al. (2017). Biosurfactant production by
Aureobasidium pullulans in stirred tank bioreactor: new approach to understand the
influence of important variables in the process. Bioresource Technology 243: 264–272.
Cameotra, S.S. and Makkar, R.S. (2004). Recent applications of biosurfactants as biological and
immunological molecules. Current Opinion in Microbiology 7 (3): 262–266.
Camilios‐Neto, D., Bugay, C., de Santana‐Filho, A.P. et al. (2011). Production of rhamnolipids
in solid‐state cultivation using a mixture of sugarcane bagasse and corn bran supplemented
with glycerol and soybean oil. Applied Microbiology and Biotechnology 89 (5): 1395–1403.
Campos, J.M., Stamford, T.L., Sarubbo, L.A. et al. (2013). Microbial biosurfactants as additives
for food industries. Biotechnology Progress 29 (5): 1097–1108.
Chaves, F.S. (2017). Bioprospecção de leveduras da Antártica para a produção de
biossurfactante a partir de hidrolisado hemicelulósico de palha de cana‐de‐açúcar. Masters
thesis. Lorena Engineering School, University of São Paulo. www.teses.usp.br.
Chen, C., Junzhang, L., Weidong, W. et al. (2019). Cost‐effective production of Surfactin from
xylose‐rich corncob Hydrolysate using Bacillus subtilis BS‐37. Waste and Biomass
Valorization 10 (2): 341–347.
Chen, W.C., Juang, R.S., and Wei, Y.H. (2015). Applications of a lipopeptide biosurfactant,
surfactin, produced by microorganisms. Biochemical Engineering Journal 103: 158–169.
Chenikher, S., Guez, J.S., Coutte, F. et al. (2010). Control of the specific growth rate of Bacillus
subtilis for the production of biosurfactant lipopeptides in bioreactors with foam overflow.
Process Biochemistry 45 (11): 1800–1807.
Choi, W.J., Choi, H.G., and Lee, W.H. (1996). Effects of ethanol and phosphate on emulsion
production by Acinetobacter calcoaceticus RAG‐1. Journal of Biotechnology 45 (3): 217–225.
Christofi, N. and Ivshina, I.B. (2002). Microbial surfactants and their use in field studies of soil
remediation: a review. Journal of Applied Microbiology 93: 915–929.
Chuo, S.C., Ahmad, A., Mohd‐Setapar, S.H. et al. (2019). Utilization of green sophorolipids
biosurfactant in reverse micelle extraction of antibiotics: kinetic and mass transfer studies.
Journal of Molecular Liquids 276: 225–232.
Cirigliano, M. and Carman, G. (1984). Isolation of a bioemulsifier from Candida lipolytica.
Applied and Environmental Microbiology 48 (4): 747–750.
Colak, A.K. and Kahraman, H. (2013). The use of raw cheese whey and olive oil mill
wastewater for rhamnolipid production by recombinant Pseudomonas aeruginosa.
Environmental and Experimental Biology 11: 125–130.
Coppe – Instituto Alberto Luiz Coimbra de Pós‐Graduação e Pesquisa de Engenharia (2009). Coppe
inaugura primeira unidade para producao de biossurfactante. www.coppe.ufrj.br/pt‐br/planeta‐
coppe‐noticias/noticias/coppe‐inaugura‐primeira‐unidade‐para‐producao‐de‐biossurfactante.
Cortés‐Sánchez, A.J., Hernández‐Sánchez, H., and Jaramillo‐Flores, M.E. (2013). Biological
activity of glycolipids produced by microorganisms: new trends and possible therapeutic
alternatives. Microbiological Research 168 (1): 22–32.
Das, K. and Mukherjee, A. (2007). Differential utilization of pyrene as the sole source of
carbon by Bacillus subtilis and Pseudomonas aeruginosa strains: role of biosurfactants in
enhancing bioavailability. Journal of Applied Microbiology 102: 195–203.
Das, P., Mukherjee, S., and Sen, R. (2008). Antimicrobial potential of a Lipopeptide
biosurfactant derived from a marine Bacillus circulans. Journal of Applied Microbiology 104:
1675–1684.
Delbeke, E.I.P., Roman, B.I., Marin, G.B. et al. (2015). A new class of antimicrobial
biosurfactants: quaternary ammonium sophorolipids. Green Chemistry 17 (6): 3373–3377.
Desai, J.D. and Banat, I.M. (1997). Microbial production of surfactants and their commercial
potential. Microbiology and Molecular Biology Reviews 61 (1): 47–64.
Desai, J.D. and Desai, A.J. (1993). Production of biosurfactants. In: Biosurfactants: Production,
Properties, Applications (ed. N. Kosaric). New York: CRC Press.
de Souza Monteiro, A., Domingues, V.S., Souza, M.V.D. et al. (2012). Bioconversion of biodiesel
refinery waste in the bioemulsifier by Trichosporon mycotoxinivorans CLA2. Biotechnology
for Biofuels 5 (29): 1–12.
Durand, A. (2003). Bioreactor designs for solid state fermentation. Biochemical Engineering
Journal 13 (2–3): 113–125.
Elsevier’s Scopus (2018). www.scopus.com.
Fahim, S., Dimitrov, K., Vauchel, P. et al. (2013). Oxygen transfer in three phase inverse
fluidized bed bioreactor during biosurfactant production by Bacillus subtilis. Biochemical
Felix, A.K.N., Martins, J.J.L., Almeida, J.G.L. et al. (2019). Purification and characterization
of a biosurfactant produced by Bacillus subtilis in cashew apple juice and its application
in the remediation of oil‐contaminated soil. Colloids and Surfaces B: Biointerfaces 175:
256–263.
Felse, P.A., Shah, V., Chan, J. et al. (2007). Sophorolipid biosynthesis by Candida bombicola
from industrial fatty acid residues. Enzyme and Microbial Technology 40 (2): 316–323.
Fernandes, P.A.V., Arruda, I.R., Santos, A.F.A.B. et al. (2007). Antimicrobial activity of
surfactants produced by Bacillus subtilis R14 against multidrug‐resistant bacteria. Brazilian
Journal of Microbiology 38 (4): 704–709.
Ferreira, A., Vecino, X., Ferreira, D. et al. (2017). Novel cosmetic formulations containing a
biosurfactant from lactobacillus paracasei. Colloids and Surfaces B: Biointerfaces 155: 522–529.
Fontes, G.C., Amaral, P.F.F., and Coelho, M.A.Z. (2008). Production of biosurfactant from
yeast. Química Nova 31 (8): 2091–2099. (original article in Portuguese).
Fooladi, T., Abdeshahian, P., Moazami, N. et al. (2018). Enhanced biosurfactant production by
Bacillus pumilus 2IR in fed‐ batch fermentation using 5‐L bioreactor. Iranian Journal of
Science and Technology, Transactions A: Science 42 (3): 1111–1123.
Fox, S.L. and Bala, G.A. (2000). Production of surfactant from Bacillus subtilis ATCC 21332
using potato substrates. Bioresource Technology 75 (3): 235–240.
Gao, Y.Z., Li, Q.L., Ling, W.T., and Zhu, X.Z. (2011). Arbuscular mycorrhizal phytoremediation
of soils contaminated with phenanthrene and pyrene. Journal of Hazardous Materials 185
(2–3): 703–709.
Garg, M. and Chatterjee, M. (2018). Isolation, characterization and antibacterial effect of
biosurfactant from Candida parapsilosis. Biotechnology Reports 18: 1–6.
Gharaei‐Fathabad, E. (2011). Biosurfactants in pharmaceutical industry (a mini‐review).
American Journal of Drug Discovery and Development 1 (1): 58–69.
Ghribi, D., Abdelkefi‐Mesrati, L., Mnif, I. et al. (2012a). Investigation of antimicrobial activity
and statistical optimization of Bacillus subtilis SPB1 biosurfactant production in solid‐state
fermentation. Journal of Biomedicine and Biotechnology 2012: 1–12.
Ghribi, D., Elleuch, M., Abdelkefi, L., and Ellouze‐Chaabouni, S. (2012b). Evaluation of
larvicidal potency of Bacillus subtilis SPB1 biosurfactant against Ephestia kuehniella
(Lepidoptera: Pyralidae) larvae and influence of abiotic factors on its insecticidal activity.
Journal of Stored Products Research 48: 68–72.
Gobbert, U., Lang, S., and Wagner, E. (1984). Sophorose lipid formation by resting cells of
Torulopsis bombicola. Biotechnology Letters 6 (4): 225–230.
Grand View Research (2015). Global biosurfactants market by product (rhamnolipids,
sophorolipids, MES, APG, sorbitan esters, sucrose esters) expected to reach USD 2,308.8
million by 2020. www.grandviewresearch.com/press‐release/global‐biosurfactants‐market.
Gudiña, E.J., Rodrigues, A.I., Alves, E. et al. (2015). Bioconversion of agro‐industrial by‐
products in rhamnolipids toward applications in enhanced oil recovery and bioremediation.
Guerra‐Santos, L., Käppeli, O., and Fiechter, A. (1984). Pseudomonas aeruginosa biosurfactant
production in continuous culture with glucose as carbon source. Applied and Environmental
Microbiology 48 (2): 301–305.
Guerra‐Santos, L.H., Kappeli, O., and Fiechter, A. (1986). Dependence of Pseudomonas
aeruginosa continuous cultures biosurfactants production on nutritional and environmental
factor. Applied Microbiology and Biotechnology 24 (6): 443–448.
Hommel, R.K., Weber, L., Weiss, A. et al. (1994). Production of sophorose lipid by Candida
(Torulopsis) apicola grown on glucose. Journal of Biotechnology 33: 147–155.
Jadhav, M., Kalme, S., Tamboli, D., and Govindwar, S. (2011). Rhamnolipid from Pseudomonas
desmolyticum NCIM‐2112 and its role in the degradation of Brown 3REL. Journal of Basic
Microbiology 51: 1–12.
Jiménez‐Peñalver, P., Castillejos, M., Koh, A. et al. (2018). Production and characterization of
sophorolipids from stearic acid by solid‐state fermentation, a cleaner alternative to chemical
surfactants. Journal of Cleaner Production 172: 2735–2747.
Joshi, S., Bharucha, C., Jha, S. et al. (2008). Biosurfactant production using molasses and whey
under thermophilic conditions. Bioresource Technology 99: 195–199.
Joshi, S.J., Geetha, S.J., Yadav, S., and Desai, A.J. (2013). Optimization of bench‐scale
production of biosurfactant by Bacillus Licheniformis R2. APCBEE Procedia 5: 232–236.
Kappeli, O. and Finnerty, W.R. (1979). Partition of alkane by an extracellular vesicle derived
from hexadecane‐grown Acinetobacter. Journal of Bacteriology 140 (2): 707–712.
Karlapudi, A.P., Venkateswarulu, T.C., Srirama, K. et al. (2018). Evaluation of anti‐cancer,
anti‐microbial and anti‐biofilm potential of biosurfactant extracted from an Acinetobacter
M6 strain. Journal of King Saud University – Science 8 April: 1–5.
Kim, H.S., Kim, Y.B., Lee, B.S., and Kim, E.K. (2005). Sophorolipid production by Candida
bombicola ATCC 22214 from a corn oil processing by product. Journal of Microbiology and
Kim, S.K., Kim, Y.C., Lee, S. et al. (2011). Insecticidal activity of rhamnolipid isolated from
Pseudomonas sp. EP‐3 against green peach aphid (Myzus persicae). Journal of Agricultural
and Food Chemistry 59 (3): 934–938.
Kiran, G.S., Ninawe, A.S., Lipton, A.N. et al. (2016). Rhamnolipid biosurfactants: evolutionary
implications, applications and future prospects from untapped marine resource. Critical
Reviews in Biotechnology 36: 399–415.
Kitamoto, D., Fuzishiro, T., Yanagishita, H. et al. (1992). Production of mannosylerythriol lipids
as biosurfactants by resting cells of Candida antarctica. Biotechnology Letters 14 (4): 305–310.
Kosaric, N. and Vardar‐Sukan, F. (2015). Biosurfactants: Production and Utilization–Processes,

Technologies, and Economics. Boca Raton, FL: CRC Press.
Krishna, C. (2005). Solid‐state fermentation systems – an overview. Critical Reviews in
Biotechnology 25: 1–30.
Kulakovskaya, E. and Kulakovskaya, T. (2014). Extracellular Glycolipids of Yeasts: Biodiversity,
Biochemistry, and Prospects. Oxford: Academic Press.
Kulakovskaya, T.V., Golubev, W.I., Tomashevskaya, M.A. et al. (2010). Production of antifungal
cellobiose lipids by Trichosporon porosum. Mycopathologia 169 (2): 117–123.
Kurtzman, C.P., Price, N.P., Ray, K.J., and Kuo, T.M. (2010). Production of sophorolipid
biosurfactants by multiple species of the Starmerella (Candida) bombicola yeast clade. FEMS
Microbiology Letters 311 (2): 140–146.
Lai, C.C., Huang, Y.C., Wei, Y.H., and Chang, J.S. (2009). Biosurfactant‐enhanced removal of
total petroleum hydrocarbons from contaminated soil. Journal of Hazardous Materials 167
(3): 609–614.
Lang, S. and Wagner, F. (1987). Structures and properties of biosurfactants. In: Biosurfactants
and Biotechnology (eds. N. Kosaric, W.L. Cairns and N.C.C. Gray). New York: Marcel Dekker.
Lin, W., Brauers, G., Ebel, R. et al. (2003). Novel chromone derivatives from the fungus
Aspergillus versicolor isolated from the marine sponge Xestospongia exigua. Journal of
Natural Products 66: 57–61.
Liu, G., Zhong, S., Yang, X. et al. (2018a). Advances in applications of rhamnolipids biosurfactant
in environmental remediation: a review. Biotechnology and Bioengineering 115 (4): 796–814.
Liu, Z., Tiana, X., Chena, Y. et al. (2018b). Efficient sophorolipids production via a novel in situ
separation technology by Starmerella bombicola. Process Biochemistry 81: 1–46.
Lourith, N. and Kanlayavattanakul, M. (2009). Natural surfactants used in cosmetics:
glycolipids. International Journal of Cosmetic Science 31 (4): 255–261.
Luna, J.M., Rufino, R.D., Jara, A.M.A.T. et al. (2014). Environmental applications of the
biosurfactant produced by Candida sphaerica cultivated in low‐cost substrates. Colloids and
Surfaces A: Physicochemical and Engineering Aspects 480: 413–418.
Ma, X., Li, H., Shao, L.J. et al. (2011). Effects of nitrogen sources on production and
composition of sophorolipids by Wickerhamiella domercqiae var. sophorolipid CGMCC 1576.
Applied Microbiology and Biotechnology 91 (6): 1623–1632.
Maddikeri, G.L., Gogate, P.R., and Pandit, A.B. (2015). Improved synthesis of sophorolipids
from waste cooking oil using fed batch approach in the presence of ultrasound. Chemical
Makkar, R.S., Cameotra, S.S., and Banat, I.M. (2011). Advances in utilization of renewable
substrates for biosurfactant production. AMB Express 1: 1–5.
Marcelino, P.R.F., da Silva, V.L., Rodrigues Philippini, R. et al. (2017). Biosurfactants produced
by Scheffersomyces stipitis cultured in sugarcane bagasse hydrolysate as new green larvicides
for the control of Aedes aegypti, a vector of neglected tropical diseases. PLoS One 12 (11):
1–16.
Marcelino, P.R.F., Peres, G.F.D., Teran‐Hilares, R. et al. (2019). Biosurfactants production by
yeasts using sugarcane bagasse hemicellulosic hydrolysate as new sustainable alternative for
lignocellulosic biorefineries. Industrial Crops and Products 129: 212–223.
Marchant, R. and Banat, I.M. (2012). Microbial biosurfactants: challenges and opportunities
for future exploitation. Trends in Biotechnology 30 (11): 558–565.
Martins, P.C. and Martins, V.G. (2018). Biosurfactant production from industrial wastes with
potential remove of insoluble paint. International Biodeterioration & Biodegradation 127:
10–16.
Mercadé, M.E., Manresa, M.A., Robert, M. et al. (1993). Olive oil mill effluent (OOME) new
substrate for biosurfactant production. Bioresource Technology 43 (1): 1–6.
Mnif, I., Elleuch, M., Chaabouni, S.E., and Ghribi, D. (2013). Bacillus subtilis SPB1
biosurfactant: production optimization and insecticidal activity against the carob moth
Ectomyelois ceratoniae. Crop Protection 50: 66–72.
Moldes, A.B., Paradelo, R., and Vecino, X. (2013). Partial characterization of biosurfactant from
Lactobacillus pentosus and comparison with sodium dodecyl sulphate for the bioremediation
of hydrocarbon contaminated soil. BioMed Research International 2013: 1–6.
Moldes, A.B., Torrado, A.M., Barral, M.T., and Dominguez, J.M. (2007). Evaluation of
biosurfactant production from various agricultural residues by Lactobacillus pentosus.
Journal of Agricultural and Food Chemistry 55 (11): 4481–4486.
Monteiro, A.S., Coutinho, J.O.P.A., Júnior, A.C. et al. (2009). Characterization of new
biosurfactant produced by Trichosporon montevideense CLOA 72 isolated from dairy
industry effluents. Jounal of Basic Microbiology 49: 553–563.
Monteiro, S.A., Sassaki, G.L., de Souza, L.M. et al. (2007). Molecular and structural
characterization of the bio‐ surfactant produced by Pseudomonas aeruginosa DAUPE 614.
Chemistry and Physics of Lipids 147 (1): 1–13.
Morita, T., Fukuoka, T., Imura, T., and Kitamoto, D. (2015). Mannosylerythritol lipids:
production and applications. Journal of Oleo Science 64: 133–141.
Müller, M.M., Hörmann, B., Syldatk, C., and Hausmann, R. (2010). Pseudomonas aeruginosa
PAO1 as a model for Rhamnolipid production in bioreactor systems. Applied Micriobiology
Mulligan, C.N., Yong, R.N., and Gibbs, B.F. (2001). Surfactant‐enhanced remediation of
contaminated soil: a review. Engineering Geology 60: 371–380.
Muso‐Cachumba, J.J. (2017). Produção de L‐asparaginase extracelular por fermentação em
estado sólido. Masters thesis. Escola de Engenharia de Lorena, Universidade de São Paulo.
Muthusamy, K., Gopalakrishnan, S., Ravi, T.K., and Sivachidambaram, P. (2008).
Biosurfactants: properties, commercial production and application. Current Science 94 (6):
736–747.
Nitschke, M. and Costa, S. (2007). Biosurfactants in food industry. Trends in Food Science &
Technology 18 (5): 252–259.
Nitschke, M., Ferraz, C., and Pastore, G.M. (2004). Selection of microrganisms for biosurfactant
production using agroindustrial wastes. Brazilian Journal of Microbiology 35: 81–85.
Nitschke, M. and Pastore, G.M. (2002). Biossurfactantes: propriedades e aplicações. Química
Nova 25 (5): 772–776.
Nitschke, M. and Pastore, G.M. (2003). Biosurfatantes a partir de resíduos agroindustriais:
Avaliação de resíduos agroindustriais como substratos para a produção de biosurfatantes por
Bacillus. Biotecnologia Ciência & Desenvolvimento 31: 63–67.
Niu, Y., Fan, L., Gu, D. et al. (2017). Characterization, enhancement and modelling of
mannosylerythritol lipid production by fungal endophyte Ceriporia lacerate CHZJU. Food
Chemistry 228: 610–617.
Pandey, A. (2003). Solid‐state fermentation. Biochemical Engineering Journal 13: 81–84.
Pandey, A., Soccol, C.R., Nigam, P., and Soccol, V.T. (2000). Biotechnological potential of
agro‐industrial residues. I: sugarcane bagasse. Bioresource Technology 74 (1): 69–80.
Pereira, A.G., Pacheco, G.J., Tavares, L.F. et al. (2013). Optimization of biosurfactant
production using waste from biodiesel industry in a new membrane assisted bioreactor.
Process Biochemistry 48 (9): 1271–1278.
Perfumo, A., Banat, I.M., and Marchant, R. (2018). Going green and cold: biosurfactants from
low‐temperature environments to biotechnology applications. Trends in Biotechnology 36
(3): 277–289.
Pinto, G.A.S., Andrade, A.M.R., Fraga, S.L.P., and Teixeira, R.B. (2005). Solid state
fermentation: an alternative to reuse and valorization of tropical agroindustrial residues.
Embrapa Comunicado Técnico 102: 1–5.
Płociniczak, M.P., Płaza, G.A., Piotrowska‐Seget, Z., and Cameotra, S.S. (2011). Environmental
applications of biosurfactants: recent advances. International Journal of Molecular Sciences
12: 633–654.
Portilla‐Rivera, O.M., Moldes, M.A.B., Torrado Agrasar, A.M., and Domíguez Gonzalez, J.M. (2007).
Biosurfactants from grape marc: stability study. Journal of Biotechnology 131S: S133–S187.
Ramana, K.V. and Karanth, N.G. (1989). Factors affecting biosurfactant productionusing
Pseudomonas aeruginosa CFTR‐6 under submerged conditions. Journal of Chemical
Technology and Biotechnology 45: 249–257.
Rau, U., Nguyen, L.A., Roeper, H. et al. (2005). Fed‐batch bioreactor production of
mannosylerythritol lipids secreted by Pseudozyma aphidis. Applied Microbiology and
Recke, V.K., Gerlitzki, M., Hausmann, R. et al. (2013). Enzymatic production of modified 2‐
dodecyl sophorosides (biosurfactants) and their characterization. European Journal of Lipid
Science and Technology 115 (4): 452–463.
Rincón‐Fontán, M., Rodríguez‐López, L., Vecino, X. et al. (2018). Design and characterization
of greener sunscreen formulations based on mica powder and a biosurfactant extract.
Powder Technology 327: 442–448.
Robert, M., Mercadé, E., Bosh, M.P. et al. (1989). Effect of the carbon source on biosurfactant
production by Pseudomonas aeruginosa 44T1. Biotechnology Letters 1 (2): 871–874.
Rodrigues, L., Banat, I.M., Teixeira, J., and Oliveira, R. (2006). Biosurfactants: potential
applications in medicine. Journal of Antimicrobial Chemotherapy 57: 609–618.
Rodríguez Couto, S. and Sanromán, M.A. (2005). Application of solid‐state fermentation to
ligninolytic enzyme production. Biochemical Engineering Journal 22 (3): 211–219.
Roy, S., Chandni, S., Das, I. et al. (2015). Aquatic model for engine oil degradation by
rhamnolipid producing Nocardiopsis VITSISB. 3 Biotech 5 (2): 153–164.
Sachdev, D.P. and Cameotra, S.S. (2013). Biosurfactants in agriculture. Applied Microbiology
and Biotechnology 97 (3): 1005–1016.
Samad, A., Zhang, J., Chen, D., and Liang, Y. (2014). Sophorolipid production from biomass
Hydrolysates. Applied Biochemistry and Biotechnology 175: 2246–2257.
Santos, A.P.P., Silva, M.D.S., and Costa, E.V.L. (2016a). Biossurfactantes: Uma alternativa para
o mercado industrial. Fronteiras: Journal of Social, Technological and Environmental Science
5 (1): 88–103.
Santos, D.K.F., Meira, H.M., Rufino, R.D. et al. (2016b). Biosurfactant production from
Candida lipolytica in bioreactor and evaluation of its toxicity for application as a
bioremediation agent. Process Biochemistry 54: 20–27.
Sarwar, A., Brader, G., Corretto, E. et al. (2018). Qualitative analysis of biosurfactants from
Bacillus species exhibiting antifungal activity. PLoS One 3 (6): 1–15.
Satpute, S.K., Plaza, G.A., and Banpurkar, A.G. (2017). Biosurfactants’ production from
renewable natural resources: example of innovativeand smart technology in circular
bioeconomy. Management Systems in Production Engineering 25 (1): 46–54.
Sekhon Randhawa, K.K. and Rahman, P.K. (2014). Rhamnolipid biosurfactants‐past, present,
and future scenario of global market. Frontiers in Microbiology 5 (454): 1–7.
Sharma, D. and Saharan, B.S. (2016). Functional characterization of biomedical potential of
biosurfactant produced by Lactobacillus helveticus. Biotechnology Reports 11: 27–35.
Shekhar, S., Sundaramanickam, A., and Balasubramanian, T. (2014). Biosurfactant producing
microbes and their potential applications: a review. Critical Reviews in Environmental
Science and Technology 45: 1522–1554.
Shepherd, R., Rockey, J., Sutherland, I.W., and Roller, S. (1995). Novel bioemulsifiers from
microorganisms for use in foods. Journal of Biotechnology 40: 207–217.
Shete, A.M., Wadhawa, G., Banat, I.M., and Chopade, B.A. (2006). Mapping of patents on
bioemulsifier and biosurfactant: a review. Journal of Scientific & Industrial Research 65:
91–115.
Silva, E.J., Correa, P.F., Almeida, D.G. et al. (2018). Recovery of contaminated marine
environments by biosurfactant‐enhanced bioremediation. Colloids and Surfaces B:
Biointerfaces 172: 127–135.
Silva, V.L., Lovaglio, R.B., von Zuben, C.J., and Contiero, J. (2015). Rhamnolipids: solution
against Aedes aegypti? Frontiers in Microbiology 6 (88): 1–5.
Siñeriz, F., Hommel, R.K., and Kleber, R.K. (2009). Production of biosurfactants. www.eolss.
net/sample‐chapters/C17/E6‐58‐05‐06.pdf.
Slivinski, C.T., Mallmann, E., de Araújo, J.M. et al. (2012). Production of surfactin by
Bacillus pumilus UFPEDA 448 in solid‐state fermentation using a medium based on
okara with sugarcane bagasse as a bulking agent. Process Biochemistry 47 (12):
1848–1855.
Smyth, T.J.P., Perfumo, A., Marchant, R., and Banat, I.M. (2010). Isolation and analysis of
lipopeptides and high molecular weight biosurfactants. In: Handbook of Hydrocarbon and
Lipid Microbiology (ed. K.N. Timmis). Heidelberg: Springer‐Verlag.
Soccol, C.R., da Costa, E.S.F., Letti, L.A.J. et al. (2017). Recent developments and innovations
in solid state fermentation. Biotechnology Research and Innovation 1 (1): 52–71.
Sousa, M., Dantas, I.T., Felix, A.K. et al. (2014). Crude glycerol from biodiesel industry as
substrate for biosurfactant production by Bacillus subtilis ATCC 6633. Brazilian Archives of
Biology and Technology 57 (2): 295–301.
Stewart, P.S. (2002). Mechanisms of antibiotic resistance in bacterial biofilms. International
Journal of Medical Microbiology 292 (2): 107–113.
Stocks, S.M., Cooke, M., and Heggs, P.J. (2005). Inverted hollow spinning cone as a device for
controlling foam and hold‐up in pilot scale gassed agitated fermentation vessels. Chemical
Engineering Science 60 (8–9): 2231–2238.
Sudhanshu, S., Arumugam, S., and Tangavel, B. (2015). Biosurfactant producing microbes and
their potential applications: a review. Critical Reviews in Environmental Science and
Technology 45 (14): 1522–1554.
Szuhaj, B.F.H. (1989). Lecithins: Sources, Manufacture & Uses. Champaign, IL: American Oil
Chemists’ Society.
Thavasi, R., Jayalaksmi, S., Balasubramanian, T., and Banat, I.M. (2007). Biosurfactant
production by Corynebacterium kutscheri from waste motor lubricant oil and peanut oil
cake. Letters in Applied Microbiology 45 (6): 696–691.
Thavasi, R., Subramanyam Nambaru, V.R.M., Jaya‐lakshmi, S. et al. (2009). Biosurfactant
production by Azotobacter chroococ‐ cum isolated from the marine environment. Marine
Thompson, D.N., Fox, S.L., and Bala, G.A. (2000). Biosurfactants from potato process effluents.
Applied Biochemistry and Biotechnology 84/86: 917–929.
Thompson, D.N., Fox, S.L., and Bala, G.A. (2001). The effects of pretreatments on surfactin
production from potato process effluent by Bacillus subtilis. Applied Biochemistry and
Biotechnology 91–93: 487–502.
Trummler, K., Effenberger, F., and Syldatk, C. (2003). An integrated microbial/enzymatic
process for production of rhamnolipids and L‐(+)‐rhamnose from rapeseed oil with
Pseudomonas sp DSM 2874. European Journal of Lipid Science and Technology 105 (10):
563–571.
Van Bogaert, I., Zhang, J., and Soetaert, W. (2011). Microbial synthesis of sophorolipids. Process
Biochemistry 46 (4): 821–833.
Van Dyke, M.I., Lee, H., and Trevors, J.T. (1991). Applications of microbial surfactants.
Biotechnology Advances 9 (2): 241–252.
Vecino, X., Cruz, J.M., Moldes, A.B., and Rodrigues, L.R. (2017). Biosurfactants in cosmetic
formulations: trends and challenges. Critical Reviews in Biotechnology 37 (7): 1–14.
Vedaraman, N. and Venkatesh, N. (2011). Production of surfactin by Bacillus subtilis mtcc 2423
from waste frying oils. Brazilian Journal of Chemical Engineering 28: 175–180.
Velioglu, Z. and Urek, R.O. (2016). Physicochemical and structural characterization of
biosurfactant produced by Pleurotus djamor in solid‐state fermentation. Biotechnology and
Bioprocess Engineering 21 (3): 430–438.
Walaa, A.E., Aymen, S.Y., Amal, E.A., and Magdy, A.A. (2016). Utilization of crude glycerol as
a substrate for the production of Rhamnolipid by Pseudomonas aeruginosa. Biotechnology
Research International 2016: 1–9. www.hindawi.com/journals/btri/2016/3464509/.
Weschayanwiwat, P., Scamehorn, J.F., and Reilly, P.J. (2005). Surfactant properties of low
molecular weight phospholipids. Journal of Surfactants and Detergents 8 (1): 65–72.
Winterburn, J.B. and Martin, P.J. (2008). Mechanisms of ultrasound foam interactions.
Asia‐Pacific Journal of Chemical Engineering 4: 184–190.
Winterburn, J.B. and Martin, P.J. (2012). Foam mitigation and exploitation in biosurfactant
production. Biotechnology Letters 34 (2): 187–195.
Wu, Y.S., Ngai, S.C., GLoh, B.H. et al. (2017). Anticancer activities of Surfactin and potential
application of nanotechnology assisted Surfactin delivery. Frontiers in Pharmacology 8 (761):
1–22.
Xiaohong, P., Xinhua, Z., and Lixiang, Z. (2009). Effect of biosurfactant on the sorption of
phenanthrene onto original and H2O2‐treated soils. Journal of Environmental Sciences 28:
1378–1385.
Xue, C., Hu, Y., Saito, H. et al. (2002). Molecular species composition of glycolipids from
Spirulina platensis. Food Chemistry 77 (1): 9–13.
Yeh, M.S., Wei, Y.H., and Chang, J.S. (2006). Bioreactor design foe enhanced carrier‐assisted
surfactin production with Bacillus subtilis. Process Biochemistry 41 (8): 1799–1805.
Zhang, Y., Jiang, J., Zhao, Q. et al. (2017). Accelerating anodic biofilms formation and electron
transfer in microbial fuel cells: role of anionic biosurfactants and mechanism.
Bioelectrochemistry 117: 48–56.
Zheng, T., Xu, Y.S., Yong, X.Y. et al. (2015). Endogenously enhanced biosurfactant production
promotes electricity generation from microbial fuel cells. Bioresource Technology 197:
416–421.
Zhu, L., Yang, X., Xue, C. et al. (2012). Enhanced rhamnolipids production by Pseudomonas
aeruginosa based on a pH stage‐controlled fed‐batch fermentation process. Bioresource
Zinjarde, S.S., Chinnathambi, S., Lachke, A.H., and Pant, A. (1997). Isolation of an emulsifier
from Yarrowia lipolytica NCIM 3589 using a modified mini isoeletric focusing unit. Letters in
Applied Microbiology 24: 117–121.
Zinjarde, S.S. and Ghosh, M. (2010). Production of surface active compounds by biocatalyst
technology. In: Biosurfactants (ed. S. Ramakrishna). New York: Springer.
185
Lignocellulose as a Renewable Carbon Source

for Microbial Synthesis of Different Enzymes
Peyman Abdeshahian1,2, Abudukeremu Kadier 3,4, Pankaj Kumar Rai5, and
Silvio Silvério da Silva1
1
2
Department of Microbiology, Masjed Soleiman Branch, Islamic Azad University, Masjed Soleiman, Iran
3
Department of Chemical and Process Engineering, Faculty of Engineering and Built Environment, National University of
Malaysia, Bangi, Selangor, Malaysia
4
Research Centre for Sustainable Process Technology (CESPRO), Faculty of Engineering and Built Environment, National
University of Malaysia, Bangi, Selangor, Malaysia
5
Department of Biotechnology, Invertis University, Bareilly, Uttar Pradesh (UP), India
9.1 Introduction
Biomass might be regarded as the organic material from a biological source. A wide variety
of biomass resources are available which could be successfully converted into bioproducts.
Biomass may include whole plants or plant materials like seeds and stalks or plant compo-
nents such as starch, lipids, proteins, and fiber. Furthermore, processing by‐products (dis-
tiller’s grains, corn soluble), materials from the marine ecosystem, animal‐origin waste,
and waste obtained from municipal and industrial sectors are considered as biomass
(Howard et al. 2003).
Plant‐derived biomass is composed of carbohydrate and lignin and is known as lignocel-
lulose (Naik et al. 2010). Lignocellulose is the most abundant biomass produced during
photosynthesis and contributes more than 60% of plant biomass. The global annual pro-
duction of lignocellulose has been estimated to be 1 × 1010 MT (Tengerdy and Szakacs 2003;
Li et al. 2010). Lignocellulose is the most widely utilized feedstock on Earth and is the main
structural component of woody and nonwoody plants. Lignocellulosic biomass may be
obtained from the forestry and agricultural sectors, paper and pulp industries, timber
industries, and various agro‐industrial sectors (Howard et al. 2003; Martins et al. 2011). In
this context, the huge amounts of lignocellulosic substances are considered as waste and
are often burned.
Ever increasing human industrial activity had led to the generation of an enormous
amount of lignocellulosic residue from woody trees, herbaceous plants, agricultural crops,
forestry, and municipal sectors (Sánchez 2009). Although these residues are considered as

waste, they could be effectively converted into a wide range of value‐added products such as
biofuels, chemical feedstocks, biofertilizers, and animal feeds, and are utilized in the pro-
duction of organic acids, enzymes, biodegradable plastics, paper, and animal and human
nutrients (Howard et al. 2003; Tengerdy and Szakacs 2003; Ravindran and Jaiswal 2016).
Enzymes are the biological compounds that catalyze different biochemical reactions,
and thus they have a vital role in many biochemical processes. They are biodegradable and
therefore have an eco‐friendly impact. Enzymes enhance the rate of reaction via reducing
activation energy, thus helping to lessen the cost of production. All these characteristic
features add to their advantage compared to chemical processes (Panesar et al. 2016).
Enzyme production is an important field of biotechnology (Couto and Sanroman 2005) and
enzymes have many applications and biotechnologic potential for various industries such
as chemicals, fuels, food, brewery and wine, animal feed, textile and laundry, pulp and
paper, and agriculture (Howard et al. 2003).
Researchers have studied the further utilization of lignocellulose‐derived products such
as bioethanol, xylitol, and microbial enzymes (Chandel et al. 2010). Lignocellulosic bio-
mass, as a low‐cost raw material, could be utilized for cost‐effective production of enzymes
due to its low commercial value. The utilization of lignocellulosic materials in enzyme
production has an ecologically beneficial role in decreasing environmental pollution cre-
ated by the detrimental production of enormous amounts of lignocellulosic material
(Soccol et al. 2014). In natural ecosystems, microorganisms such as bacteria and fungi
thrive on natural lignocellulose. White‐rot fungi and other mushrooms are among the most
efficient decomposers of wood and other natural lignocelluloses. Lignocellulose‐degrading
fungi are now used at industrial scale, mostly for the production of enzymes (Tengerdy and
Szakacs 2003). The industrial demand for novel enzymes with different catalytic properties
has augmented the isolation and selection of new microbial strains capable of using vari-
ous lignocellulosic feedstocks. This chapter provides an overview of lignocellulose and its
utilization in biotechnologic enzyme production.
9.2 Lignocellulose
In general, lignocellulosic biomass is a by‐product of industries or agricultural processes.

Therefore, lignocellulosic biomass (except energy crops) is produced without consideration
of land, water, and energy. Meanwhile, biomass handling, transportation, and storage can
dramatically add to lignocellulosic raw material costs. In this context, lignocellulose‐
containing wastes include wood, agricultural residues, forest residues, industrial and
municipal wastes, energy crops, and paper waste (Banerjee et al. 2010).
Lignocellulose is a complex biopolymer which consists primarily of cellulose, followed
by hemicellulose and lignin (Sánchez 2009). In general, lignocellulose is composed of up to
75% of cellulose and hemicellulose (Liu et al. 2006). Polymeric content of cellulose, hemi-
cellulose and lignin is linked together by covalent and noncovalent bonds in the cell wall of
plants (Abdeshahian et al. 2010a). Table 9.1 represents a variety of lignocellulosic sub-
stances with a range of cellulose, hemicelluloses, and lignin content.
Plant cell walls contain cellulose microfibrils which are surrounded by lignin, hemicel-
lulose, and pectin. The middle lamella, a thin layer lying between individual cells, sticks
Lignocellulose as a Renewable Carbon Source for Microbial Synthesis of Different Enzymes 187
Table 9.1 Different lignocellulose-containing materials showing the cellulose, hemicellulose,

and lignin content.
Lignocellulosic materials Cellulose (%) Hemicellulose (%) Lignin (%)
Hardwood stems 40–55 24–40 18–25

Softwood stems 45–50 25–35 25–35
Nut shells 25–30 25–30 30–40
Corn cobs 45 35 15
Rice straw 32.1 24 18
Waste paper from chemical pulps 60–70 10–20 5–10
Switchgrass 45 31.4 12
Sugarcane bagasse 19–24 27–32 19–24
Wheat straw 29–35 26–32 16–21
Barley straw 31–34 24–29 14–15
Oat straw 31–37 27–38 16–19
Rye straw 33–35 27–30 16–19
Bamboo 26–43 15–26 21–31
Coffee pulp 35 46.3 18.8
Banana waste 13.2 14.8 14
Corn stalks 61.2 19.3 6.9
Sugarbeet waste 26.3 18.5 2.5
Soya stalks 34.5 24.8 19.8
Sunflower stalks 42.1 29.7 13.4
Source: Sánchez (2009), Kumar et al. (2014), and Sadh et al. (2018).
the cells together to create tissue. In the primary cell wall, the microfibrils make up an
interwoven network. The development of a cell wall begins in the primary wall, and then
proceeds in the secondary wall, which is usually composed of cellulose fibrils arranged in
a crossing or parallel pattern (Mitchell et al. 1992).
Cellulose and hemicelluloses are macromolecules composed of various sugars, while
lignin is an aromatic polymer made from phenylpropanoid precursors (Sánchez 2009). In
the following sections, various important fractions of lignocellulose structure are described
in detail.
9.2.1 Cellulose
Cellulose is the most common skeletal polysaccharide, constituting about 50% of the cell
wall content of plants. In addition to hemicellulose and lignin, cellulose represents the
main component of agricultural waste and municipal residues (Petre et al. 1999). It is one
of the most essential sources of carbon on the planet and its yearly production by both
plants and marine algae is estimated to be 0.85 × 1011 tons (Niranjane et al. 2007). Cellulose
is a linear polymer containing d‐glucose subunits linked by β‐1,4 glycosidic bonds which
constitute the dimeric cellobiose. These kinds of long chains (or elemental fibrils) are
linked together by hydrogen bonds and van der Waals forces. Cellulose usually is found as
a crystalline structure and a small quantity of nonorganized cellulose chains make up
amorphous cellulose (Malherbe and Cloete 2002; Sánchez 2009).
Cellulose molecules contain long slender bundles of long chains of β‐d‐glucopyranose
residues linked by 1‐4 glucosidic bonds which are referred as elementary fibrils. Each ele-
mentary fibril has cellulose molecules that are laterally bound, while the adjacent mole-
cules are parallel, placed in opposite directions with different degrees of orientation (Petre
et al. 1999).
9.2.2 Hemicellulose
Hemicellulose accounts for 35% of lignocellulosic biomass (Chandel et al. 2010). It is a
polysaccharide with a lower molecular weight than cellulose. It is composed of a number
of simple sugars (pentoses and hexoses) and glucouronic acid, including D‐xylose,
D‐mannose, D‐galactose, D‐glucose, L‐arabinose, 4‐O‐methylglucuronic, D‐galacturonic,
and D‐glucuronic acids. The major difference between cellulose and hemicellulose is
attributed to branches with short lateral chains of different sugars in hemicelluloses,
whereas cellulose contains oligomers that are easily hydrolyzed. It is noteworthy that
hemicellulose is more soluble than cellulose (Malherbe and Cloete 2002; Sánchez 2009).
9.2.2.1 Xylan
Xylan is a main hemicellulosic constituent of plant cell walls which represents the second
most abundant polysaccharide in Nature after cellulose. In woody tissues, xylan is present
mainly in the secondary cell wall. Xylan along with lignin constitutes an amorphous matrix
that surrounds cellulose microfibrils (Javier et al. 2007).
Xylan is the major hemicellulose in hardwoods and is found in smaller amounts in soft-
woods, accounting for approximately 20–25% and 7–12% of their total dry weights, respec-
tively (Senthilkumar et al. 2008). Xylan comprises a β‐1,4‐linked D‐xylosyl residue backbone
branched with a low portion of arabinose, glucuronic, and arabinoglucuronic acids linked
to the D‐xylose backbone (Katapodis et al. 2007; Lakshmi et al. 2009; Abdeshahian et al.
2010b). In this regard, arabino glucuronoxylans include a β‐1,4‐linked xylopyranose back-
bone substituted by α‐L‐arabinofuranosyl and glucuronic acid or its methyl ether (Gomes
et al. 2000).
Xylans are available as by‐products from the forestry, agriculture, wood, some seaweed,
pulp and paper industries in large quantities (Ebringerova et al. 2005). Xylan is normally
found in hardwoods and softwoods as glucuronoxylan and glucuronoarabinoxylan, respec-
tively (Javier et al. 2007).
9.2.2.2 Mannan
Mannan is an essential part of the hemicellulosic content of plants and is found primarily
in softwood, with four forms: linear mannan, glucomannan, galactomannan, and galacto-
glucomannan (Moreira and Filho 2008). The chemical structures of these polysaccharides
are arranged according to a backbone of (1,4)‐linked β‐D‐mannosyl residues (Kote et al.
2009). Linear mannans are homopolysaccharides consisting of linear main chains of
1,4‐linked β‐D‐mannopyranosyl residues together with less than 5% galactose.

Glucomannans are polysaccharides found in the hemicellulosic fraction of softwood in
large quantities. They comprise erratically arranged β‐(1,4)‐linked D‐mannose and β‐
(1,4)‐linked D‐glucose residues (Moreira and Filho 2008). When α‐1,6‐linked galactosyl
residues replace the backbone of both mannan and glucomannan, these polysaccharides
are named galactomannan and galactoglucomannan respectively (Gomesm et al. 2007).
In this regard, plant galactomannans contain water‐soluble 1,4‐linked residues of β‐D‐
mannopyranosyl in which side‐chains of single 1,6‐linked α‐D‐galactopyranosyl groups
are attached to the chain.
Biochemical studies reveal that galactoglucomannans are composed of residues of D‐
galactose linked to both D‐glucosyl and D‐mannosyl units as terminal branches α‐1,6
(Ebringerova et al. 2005). β‐mannan‐based polysaccharides are the main constituent units
in softwood and to a lesser extent in hardwoods. It has been found that mannans form the
polysaccharide content of leguminous seeds, such as guar (Cyanopsis tetragonoloba), locust
bean or carob (Ceratonia siliqua), and tara gum (Caesalpinia spinosa). Ivory nuts, green
coffee beans, coconut kernel, konjac tubers, and palm seeds are typically composed of
mannan‐based polysaccharide (Araujo and Ward 1990; Lin and Chen 2004; Ebringerova
et al. 2005; Moreira and Filho 2008).
9.2.3 Lignin
Lignin accounts for up to 30% of lignocellulose biomass. Lignin is joined to both hemicel-
lulose and cellulose, making an impenetrable barrier in the plant cell wall of. The lignin
present in the cell wall provides structural support, impermeability, and resistance to oxi-
dative reactions and microbial attacks. As far as its biochemical properties are concerned,
lignin has an amorphous structure which is watersoluble and an optically inactive heter-
opolymer. Lignin is a resinous substance that is composed of phenylpropane units attached
together by nonhydrolyzable linkages. Moreover, lignin is a noncrystalline structure which
has been seen as analogous to a gel or foam and thus is capable of binding cellulose fibers
(Sánchez 2009; Irmer 2017).
9.3 Lignocellulose as a Cost-Effective Substrate

for Enzyme Production
Numerous biotechnologic processes have been developed to use lignocellulosic biomass as

raw material for the production of value‐added chemicals such as organic acids, amino
acids, ethanol, single‐cell proteins, and biofuels. Hence, lignocellulosic substances are seen
as valuable alternative bio‐based substrates with the potential for utilization in the synthe-
sis of enzymes which can also reduce pollution problems caused by their disposal
(Niranjane et al. 2007; Lakshmi et al. 2009).
Lignocellulose represents a huge carbohydrate source which can provide soluble carbons
for use in biotechnologic processes to synthesize value‐added products. From the economic
point of view, carbohydrate polymers of lignocellulosic materials can be utilized as a low‐
cost carbon source for enzyme production by microorganisms. Generally, microorganisms
consume carbohydrate polymers of lignocellulose for their growth and metabolic activities.
In order to use the polymeric structure of carbons in cellulosic and hemicellulosic frac-
tions, extracellular enzymes are secreted to hydrolyze polysaccharides for release of sugar
monomers as a simple nutrient source. In this way, microorganisms use the carbon content
of lignocelluloses for growth and synthesize of cellular metabolites (Pandey et al. 2001).
9.4 Microorganisms as Enzyme Producers

Many microorganisms, including bacteria, yeast, actinomycetes and fungi, can produce
enzymes using various lignocellulosic substances. Filamentous fungi can produce enzymes
to a large extent in comparison with other microorganisms. Therefore, fungi are important
for the commercial production of enzymes. Various strains of fungi have been used for the
production of enzymes, in particulare the genera Aspergillus and Trichoderma. Table 9.2
lists various microorganisms used in enzyme production from a variety of lignocellulosic
substrates.
Table 9.2 Enzymes produced by different microorganisms using lignocellulosic feedstocks
Enzyme Microorganism Lignocellulosic substrate References
Cellulase, Aspergillus niger Wheat bran Pandey et al. (2000,

xylanase 2001)
Cellulase Penicillium citrinum Rice husk Pandey et al. (2000,
2001)
Cellulase Trichoderma reesei, Bagasse Pandey et al. (2000,
Aspergillus spp. 2001)
Cellulase, Polyporus spp. Bagasse Pandey et al. (2000,
ligninase 2001)
Cellulase Neurospora crassa Wheat straw Pandey et al. (2000,
2001)
Cellulase T. reesei, Sporotrichum Wheat bran Pandey et al. (2000,
pulverulentum 2001)
Cellulase, A. niger Palm oil mill waste Pandey et al. (2000,
xylanase 2001)
Cellulase, Pleurotus sajor‐caju Sago hampas Pandey et al. (2001)
xylanase,
ligninase
Cellulase T. reesei Willow Pandey et al. (2000,
2001)
Cellulase, Gliocladium sp. Sweet sorghum Pandey et al. (2000,
xylanase 2001)
Cellulase T. reesei Paddy straw Pandey et al. (2000,
2001)
Cellulase Lentinus edodes Wheat straw Pandey et al. (2001)
Cellulase, T. reesei, A. niger Sweet sorghum silage Pandey et al. (2001)

xylanase
Xylanase Bacillus licheniformis Wheat bran Pandey et al. (2001)
Cellulase, L. edodes Rice straw, cotton stalks, Pandey et al. (2001)
xylanase, Kenaf
ligninase
Xylanase A. niger Wheat bran, bagasse Dobrev et al. (2007)
Xylanase, Phanerochaete Wheat bran Pandey et al. (2000,
xylosidase chrysosporium 2001)
Xylanase Chaetomium globosum, Wheat bran, apple Pandey et al. (2000,
A. niger pomace, sugarbeet pulp, 2001)
wheat straw
Xylanase, A. fumigatus Rice straw, corn hull, corn Pandey et al. (2000,
xylosidase cobs, bagasse, wheat bran 2001)
Ligninase P. chrysosporium Wood chips Pandey et al. (2001)
Ligninase P. chrysosporium, Cotton stalks Abdel‐Hamid et al.
Phanerochaete ostreatus (2013)
Protease Bacillus amyloliquefaciens Soybean meal Pandey et al. (2000,
2001)
Protease A. niger Wheat bran Pandey et al. (2001)
Protease Rhizopus oligosporus Rice bran Pandey et al. (2001)
Protease A. oryzae Rice hull, corn bran, Pandey et al. (2000);
wheat bran, rice bran Panesar et al. (2016)
Protease P. chrysosporium Aspen wood Pandey et al. (2000);
Panesar et al. (2016)
Protease Actinomycetes strains Wheat bran Pandey et al. (2000,
2001)
Lipase Candida rugosa Coconut oil cake, rice Pandey et al. (2001)
bran
Lipase Neurospora sitophila, Peanut press cake Pandey et al. (2000);
R. oligosporus Soccol et al. (2014)
Lipase Penicillium candidum Wheat bran Pandey et al. (2001);
Soccol et al. (2014)
Glutaminase Vibrio costicola Wheat bran, rice husk, Pandey et al. (2000,
coconut oil cake 2001)
Amylase B. licheniformis Wheat bran Pandey et al. (2001)
Amylase Pycnoporus sanguineus Wheat bran Pandey et al. (2001)
Amylase Aeromonas caviae Banana waste Pandey et al. (2000)
Amylase A. oryzae Coconut oil cake Ramachandran et al.
(2004)
(Continued)
Amylase A. awamori Wheat bran Pandey et al. (2001)

Amylase A. niger Copra waste Pandey et al. (2001)
Inulinase Staphylococcus spp., Chicory roots, wheat bran Pandey et al. (2001)
Kluyveromyces marxianus
Phytase Aspergillus carbonarius, Canola meal Pandey et al. (2001)
A. ficuum
Phytase Bacillus subtilis Wheat bran Pandey et al. (2001)
Pectinases A. niger Deseeded sunflower head Patil and Dayanand
(2006)
Pectinases A. oryzae Sugarcane bagasse, citrus Biz et al. (2016)
pulp
Pectinases Penicillium viridicatum Wheat bran, orange Silva et al. (2005)
bagasse
Invertase Saccharomyces cerevisiae Red carrot jam processing Panesar et al. (2016)
residue
Invertase A. niger Carrot peels Ashraf and Bilal
(2015)
Mannanase, A. niger Palm kernel cake Abdeshahian et al.
β‐mannocidase (2010)
Mannanase A. flavus Defatted copra meal Kote et al. (2009)
Mannanase Ceriporiopsis Pinus taeda wood chips Magalhães and
subvermispora Milagres (2009)
Mannanase, A. awamori Coffee waste and wheat Kurakake and
β‐mannocidase bran Komaki (2001)
Mannanase B. subtilis Konjac powder Jiang et al. (2006)
Microorganisms can degrade lignocellulosic materials by various fermentation pro-

cesses. In this context, submerged fermentation (SmF) refers to the growth of microorgan-
isms in a liquid medium which contains nutrients. Another type of fermentation process
has been developed for the growth of microorganisms on solid‐based carbon sources in
which culture contains a very low amount of free water (Kim and Han 2014). In this con-
text, if the substrate itself acts as the carbon/energy source for the growth of microorgan-
isms, the process is called solid substrate fermentation. If the substrate acts as a carbon/
energy source or is used as an inert material in which microorganisms are placed for micro-
bial growth, the fermentation process is called solid‐state fermentation (Pandey et al. 2000).
Generally, solid substrate/state fermentation (SSF) presents economic and industrial
advantages over SmF, including lower waste water output, lower energy and fermentation
media requirement, less downstream processing cost, simplicity of process, higher produc-
tivity, and lower amount of investment capital (Pandey et al. 2001).
Although most enzyme manufacturers produce enzymes using SmF technology, there is
increasing interest in using SSF to produce enzymes (Pandey et al. 2000; Couto and
Sanroman 2005), since SSF has been reported to give higher yields and is economically
more suitable due to low input, higher output, and decreased streaming costs and capital
investment (Rajoka et al. 2006; Gao et al. 2008).
9.5 Enzymes Produced Using Different Lignocellulosic

Biomass
9.5.1 Cellulase
Cellulase is mainly composed of three enzymes: exocellobiohydrolase (1,4‐β‐D‐glucan cel-
lobiohydrolase, EC 3.2.1.91), endo‐1,4‐β‐D‐glucanase (1,4‐β‐D‐glucan glucanohydrolase,
EC 3.2.1.4), and β‐glucosidase (β‐D‐glucoside glucohydrolase, EC 3.2.1.21; cellobiase).
Exocellobiohydrolases (cellobiohydrolases) help to release glucose or cellobiose from the
non reducing end of cellulose chains. Endo‐1,4‐β‐D‐glucanases randomly cleave cellulose
chains to produce mainly glucose and cellobiose. Moreover, β‐glucosidases (cellobiases)
hydrolyze cellobiose and short‐chain cellooligosaccharides to produce glucose
(Abdeshahian et al. 2010d). In the classic action of cellulase, exoglucohydrolase (1,4‐β‐D‐
glucan glucohydrolase, EC 3.2.1.74) acts on soluble oligosaccharides.
Numerous microorganisms, such as Clostridium, Cellulomonas, Thermomonospora,
Trichoderma and Aspergillus genera, have been used as the main agents for cellulase pro-
duction. Many lignocellulosic substances have been applied as a substrate for cellulase pro-
duction by microbes. Among them, agricultural waste has received a great deal of interest
as a cheap source of lignocellulose. For cellulase production, agro‐industrial residues, such
as rice bran, wheat straw, barley straw, corn cobs, sugarcane bagasse, palm oil waste, sweet
sorghum pulp, coconut oil cake, and sugar beet pulp, could be used as shown in Table 9.2.
Cellulases are commonly used in large volumes in various industrial applications, includ-
ing starch processing, animal feed production, fermentation of grain alcohol, malting and
brewing, fruit and vegetable juice extraction, pulp and paper, and textile industry (Latifian
et al. 2007).
9.5.2 Xylan-Degrading Enzymes

As mentioned earlier, the backbone of xylan chains is composed of β‐1,4‐linked xylopyra-
nose residues with the side chains of arabinosyl, glucuronosyl, methylglucuronosyl, acetyl,
feruloyl and ρ‐coumaroyl residues. Xylan backbone is hydrolyzed by xylan degrading
enzymes, namely xylanase (endo‐β‐1,4‐xylanase, EC 3.2.1.8) and β‐xylosidases (EC 3.2.1.37)
in particular. Xylanase cleaves internal linkages inside xylan to release oligosaccharides,
and β‐xylosidases release xylose units by hydrolysis of xylobiose and xylooligomers. The
side‐chains of xylan are catalyzed by α‐L‐arabinofuranosidases (EC 3.2.1.55), α‐D‐glucuro-
nidases (EC 3.2.1.139), acetylxylan esterases (EC 3.1.1.72), ferulic acid esterases (EC
3.1.1.73) and ρ‐coumaric acid esterases (EC 3.1.1.1) (Dobrev et al. 2007; Liu et al. 2008;
Abdeshahian et al. 2009).
Different types of microbes can produce xylanolytic enzymes, including bacteria, fungi,
yeasts, and protozoa. Compared to yeast and bacteria, filamentous fungi like Aspergillus spp.
and Trichoderma spp. have received more attention from industry due to their high xylanase
production. In recent years, the production of xylanases by the genus Penicillium has gained
much interest due to the particular properties and applications of these kinds of xylanase
(Fang et al. 2007; Katapodis et al. 2007; Li et al. 2007; Lakshmi et al. 2009).
Potentially abundant and inexpensive lignocellulosic wastes, especially agricultural resi-
dues such as wheat bran, corn cobs, rice bran, wheat straw, rice husks and other similar
substrates, are widely used by microorganisms to produce xylanases (Motta et al. 2013).
Xylanases have been applied in various industries but are mainly used to clarify fruit juice,
in the wine, malting, and beverage industries, and in the production of baked goods, wheat
bread, and biscuits. Xylanases could be used to extract vegetable oil and raise the nutri-
tional value of coffee, starch, and feed for animals. Use of xylanases for the biobleaching of
paper pulp has been demonstrated (Park et al. 2002; Sá‐Pereira et al. 2003; Milagres et al.
2004; Azin et al. 2007; Li et al. 2007). In recent years, xylanases have gained much interest
due to their application in biofuel and bioenergy production from a wide range of lignocel-
lulosic feedstocks (Sá‐Pereira et al. 2003; Xin and He 2013; Thomas et al. 2016; Prasertsan
et al. 2017).
9.5.3 Mannan-Degrading Enzymes

The structure of mannan‐containing polysaccharides is based on 1,4‐β‐D‐mannosidic link-
ages. Hence, mannan‐degrading enzymes affect the β‐1,4‐linked internal connection
within mannan‐based polysaccharides. The complete hydrolysis of mannan is obtained by
the combined action of several enzymes. Endo‐acting β‐mannanases (EC 3.2.1.78) catalyze
the random hydrolysis of β‐1,4‐linkages within the main chain of β‐1,4‐mannans to short
linear or branched β‐1,4‐mannooligomers, which are further cleaved by the exo‐acting
β‐mannosidase (EC 3.2.1.25) to mannose. Enzymes such as α‐galactosidase (EC 3.2.1.22),
β‐glucosidase (EC 3.2.1.21), and esterase (EC 3.1.1.72) are essential to cleave side residues
including galactose, glucose, and acetyl groups, respectively (Gomes et al. 2007; Lin et al.
2007; Moreira and Filho 2008; Abdeshahian et al. 2010c).
Different types of fungi, bacteria and yeast are being used to produce mannose degrading
enzymes. For example, the genera Aspergillus, Trichoderma, Penicillium, Bacillus,
Clostridium, Pichia and Saccharomyces have been utilized as mannose degrading enzyme
producer (Moreira and Filho 2008).
Lignocellulosic substances including palm kernel cake, defatted copra meal and locust
bean gum have been found to be a source of microbial mannanase production. Mannan‐
degrading enzymes have been commonly used for pulp biobleaching in the paper industry,
coffee beans treatment, Konjac production, animal feedstuffs improvement, printing and
dyeing, textile industry, oil production and in the food industry as valuable food sweeteners
or additives (Lin and Chen 2004; Benech et al. 2007; Gomes et al. 2007; Lin et al. 2007).
9.5.4 Lignin-Degrading Enzymes

Three main ligninolytic enzymes, laccase (E.C.1.10.3.2), lignin peroxidase (E. 1.11.1.14),
and manganese peroxidase (E.C. 1.11.1.13), can degrade lignin. Laccase acts as phenol oxi-
dase and degrades the phenolic compounds. Lignin peroxidases are capable of oxidizing
nonphenolic aromatic compounds. Manganese peroxidase oxidizes manganese ions (Mn2+)

in concert with other lignin‐degrading enzymes to depolymerize lignin.
White‐rot fungi, classified as basidiomycetes, produce ligninolytic enzymes using ligno-
cellulosic materials to degrade the lignin fraction (Hofrichter 2002; Howard et al. 2003;
Martins et al. 2011). Numerous white‐rot fungi capable of producing lignolytic enzymes
have been studied such as Phanerochaete chrysosporium, Pleurotus ostreatus, Coriolus versi-
color, Cyathus stercoreus, and Ceriporiopsis subvermispora (Abdel‐Hamid et al. 2013).
Many lignocellulosic materials from the forestry and wood industries could be used by
microorganisms to produce ligninolytic enzymes. Lignin‐degrading enzymes have industrial
application in bleaching, paper deinking, polishing and preparation of textiles, wastewater
treatment, and herbicide degradation (Ravindran and Jaiswal 2016; Abdel‐Hamid et al. 2013).
9.5.5 Lipase
Triacylglycerol‐acylhydrolases or lipases (E.C. 3.1.1.3) are multifunctional enzymes which
are capable of catalyzing many reactions including esterification, transesterification, and
interesterification of lipids. Lipase can also catalyze complete or partial hydrolysis of tria-
cylglycerols (Panesar et al. 2016).
A group of microorganisms such as bacteria, yeast, fungi, actinomycetes, and archaea
have been exploited for lipase synthesis. Various strains of Candida spp., Aspergillus spp.,
Rhizopus spp., Neurospora sitophila, Penicillium candidum, Mucor spp., Bacillus spp.,
Yarrowia lipolytica, Pseudomonas aeruginosa, Acinetobacter calcoaceticus have been uti-
lized in lipase production (Pandey et al. 2000; Soccol et al. 2014).
A wide range of lignocellulosic wastes from agriculture has been used for the production of
lipase, including wheat bran, peanut press cake, coconut oil cake, and rice bran. Furthermore,
lignocellulosic substances obtained from agro‐industrial waste water such as the effluent
from palm oil mills are potential substrates for lipase production (Pandey et al. 2001; Panesar
et al. 2016). Lipases possess great potential in the food industry, especially for the manufac-
ture of mono‐ and diglycerides, the production of lipids with high levels of polyunsaturated
fatty acids, flavor development, and for cheese maturation (Panesar et al. 2016).
9.5.6 Protease
Protease is a hydrolytic enzyme which acts on the hydrolysis of peptide bonds between
amino acids in the polypeptide chain. There are six families of proteases known: serine
proteases (EC 3.4.21), serine carboxyl proteases (EC 3.4.16), cysteine proteases (EC 3.4.22),
aspartic proteases (EC 3.4.23), metalloproteases I (EC 3.4.24), and metallocarboxy pro-
teases (EC 3.4.17). Alkaline proteases, which typically have either a serine center or a metal
part, have been extensively used in industrial sectors and account for 89% of total protease
business (Soccol et al. 2014; Panesar et al. 2016; Sharma et al. 2017).
Numerous microorganisms have been investigated for the production of proteases,
including Aspergillus spp., Penicillium spp., Rhizopus spp., Bacillus spp., Album engyodon-
tium, Synergistes spp., and P. aeruginosa (Pandey et al. 2000; Soccol et al. 2014). Proteases
can be produced from a wide range of lignocellulosic precursors including wheat bran,
sunflower flour, coffee husk, soybean meal, rice bran, corn bran, rice hull, aspen wood,
c otton cake, mustard cake, maize bran, and cane bagasse (Pandey et al. 2000; Panesar et al.
2016). Proteases are used in the detergent, leather, pharmaceutical and chemical industries
in various ways. They have great potential in the food industry, for meat tenderization,
cheese production, and bakery products (Pandey et al. 2001; Panesar et al. 2016).
9.5.7 Amylase
Amylases are categorized as hydrolytic enzymes that split starch into glucose units.
Amylases are divided into three types: α‐amylase, β‐amylase, and glucoamylases (amylo-
glucosidase) which are responsible for complete starch hydrolysis (Pandey et al. 2001;
Panesar et al. 2016). Different strains of Aspergillus spp., Rhizopus spp., Mucor spp., Bacillus
spp., and Saccharomyces spp. have been reported to utilize lignocellulosic biomass for
amylase production (Pandey et al. 2000).
A variety of agro‐industrial lignocellulosic waste, such as wheat bran, rice bran, rice
husk, coconut cake, tea waste, cassava, cassava bagasse, sugarcane bagasse, banana waste,
corn flour, sawdust, soy meal, sweet potatoes, potatoes, rice hull, and sugarbeet pulp, has
been used as substrate for the production of amylases (Pandey et al. 2000). Amylase
enzymes possess great potential for the baking, brewing, fruit, textile, and paper industries
(Pandey et al. 2001; Panesar et al. 2016).
9.5.8 Phytase
Phytases (myoinositol hexakisphosphate phosphohydrolase, EC 3.1.3.8 and EC 3.1.3.26)
are in the subclass of histidine acid phosphatases which affect the hydrolysis of myoinosi-
tol hexakisphosphate (phytic acid) to release inorganic phosphate, myoinositol phosphate
esters and free myoinositol (Pandey et al. 2001; Soccol et al. 2014). Numerous studies have
reported that some microbes are capable of producing phytases from lignocellulosic bio-
mass, including Aspergillus niger, Bacillus subtilis, Thermoascus aurantiacus, Klebsiella
spp., Aspergillus ficuum, Mucor indicus, Nocardia spp., Schizophyllum commune, and
Aspergillus carbonarius (Pandey et al. 2000; Soccol et al. 2014).
Agricultural residues such as wheat bran, rice bran, and rice mill waste have been used
as lignocellulosic substances for the production of phytases (Pandey et al. 2000; Soccol
et al. 2014). Phytases have been extensively applied in animal feed to enhance access to
phosphorus and minerals (Soccol et al. 2014; Panesar et al. 2016).
9.5.9 Pectinase
Pectinase (EC 3.3.1.15) is a hydrolytic enzyme that can catalyze hydrolysis of pectic bonds,
particularly in the plant cell wall. Several microbial strains such as T. aurantiacus, A. niger,
Saccharomyces cerevisiae, and A. oryzae have been used for pectinase synthesis (Soccol
et al. 2014; Panesar et al. 2016; Ravindran and Jaiswal 2016). Lignocellulosic substances
like deseeded sunflower head, wheat bran, and coconut bran have been reported to be good
substrates for pectinase production by microorganisms (Pandey et al. 2001; Soccol et al.
2014; Panesar et al. 2016; Ravindran and Jaiswal 2016). Pectinases have been utilized in the
processing of starch, wine, and juice (Ravindran and Jaiswal 2016).
9.5.10 Glutaminase
The enzyme glutaminase (L‐glutamine amidohydrolase, E.C. 3.5.1.2) acts as deamidating
L‐glutamine and plays a pivotal role in cellular metabolism. Various bacteria, actinomy-
cetes, yeast, and fungi are capable of producing glutaminase. Bacteria Pseudomonas fluo-
rescens, Vibrio costicola, V. cholerae, Bacillus spp., and fungi A. oryzae, Actinomucor
elegans, and A. taiwanenesis have been studied for glutaminase production (Pandey et al.
2000, 2001).
Lignocellulosic biomass including wheat bran, rice husk, sawdust, and coconut oil cake
has been utilized for microbial production of glutaminase. Glutaminase has been used in
the food industry as a flavor enhancer particularly for fermented foods such as soy sauce,
miso, and sufu. It has been used in the pharmaceutical industry for cancer therapy (Pandey
et al. 2000, 2001; Binod et al. 2017).
9.5.11 Inulinase
Inulinase (2,1‐β‐D fructan fructanohydrolase, EC 3.2.1.7) is an inulin hydrolyzing enzyme
that acts by cleaving glycosidic linkages for cutting terminal fructosyl units (D‐fructose).
Microbial strains of Staphylococcus spp., Kluyveromyces marxianus, Aspergillus kawachii,
Pencillium rugulosum, Saccharomyces spp., and Staphylococcus spp. have been used for
inulinase production (Pandey et al. 2001; Ravindran and Jaiswal 2016).
Lignocellulosic substances such as wheat bran, coconut oil cake, rice bran, chicory roots,
and bagasse have been utilized as a substrate for inulinase production by microorganisms
(Pandey et al. 2001; Ravindran and Jaiswal 2016). Inulinase enzyme is used in the food
industry for production of pure fructose syrup from inulin‐rich substances (Fernandes and
Jiang 2013).
9.5.12 Invertase
Invertase (β‐fructofuranosidase, EC 3.2.1.26) catalyzes the hydrolysis of sucrose into
D‐glucose and D‐fructose. Microorganisms reported to produce invertase are Aspergillus,
Bacillus, Saccharomyces, and Fusarium. The most commonly used microorganisms for
inulinase synthesis in sucrose fermentation process are S. cerevisiae, S. carlsbergensis, A.
niger, and Cladosporium cladosporioides (Panesar et al. 2016; Ravindran and Jaiswal 2016).
Sugarcane bagasse is a suitable lignocellulosic biomass for inulinase production.
Moreover, the invertase enzyme has many potential applications in the food industry,
including candy, syrups, condensed milk, infant food, and artificial honey production. In
addition, it is used in cosmetics as well as in the analytical laboratory for making sucrose
biosensors (Panesar et al. 2016; Ravindran and Jaiswal 2016).
9.6 Conclusion
Lignocellulose represents a renewable feedstock, comprising cellulose, hemicellulose, and

lignin. A large number of lignocellulosic substances are considered as waste which could be
employed as raw materials for valorization and synthesis of value‐added products.

Lignocellulosic biomass is rich in organic carbons which could be utilized as a nutrient source
for microorganisms to produce various enzymes. Commercial enzyme synthesis entails high
cost because of expensive raw substrates. Lignocellulosic materials provide a low‐cost sub-
strate for economically viable production of enzymes in large‐scale bioprocesses.
Acknowledgment
Peyman Abdeshahian is grateful to the Research Council for the State of Sao Paulo
(FAPESP), Brazil, for providing financial assistance (Process No. 2018/14095‐7) in the form
of a postdoctoral fellowship, and Silvio Silverio da Silva is grateful to the Brazilian National
Council for Scientific and Technological Development (CNPq) (Process No. 303943/2017‐3)
and FAPESP (Process No. 2016/10636-8) for providing financial support for research.
References
Abdel‐Hamid, A.M., Solbiati, J.O., and Cann, I.K. (2013). Insights into lignin degradation and
its potential industrial applications. Advances in Applied Microbiology 82: 1–28.
Abdeshahian, P., Dashti, M.G., Kalil, M.S., and Yusoff, W.M.W. (2010a). Production of biofuel
using biomass as a sustainable biological resource. Biotechnology 9: 274–282.
Abdeshahian, P., Samat, N., Hamid, A.A., and Yusoff, W.M.W. (2010b). Utilization of palm
kernel cake for production of β‐mannanase by Aspergillus niger FTCC 5003 in solid substrate
fermentation using an aerated column bioreactor. Journal of Industrial Microbiology and
Abdeshahian, P., Samat, N., and Yusoff, W.M.W. (2009). Xylanase production by Aspergillus
niger FTCC 5003 using palm kernel cake in fermentative bioprocess. Pakistan Journal of
Biological Sciences 12: 1049–1055.
Abdeshahian, P., Samat, N., and Yusoff, W.M.W. (2010c). Production of β‐xylosidase by
Aspergillus niger FTCC 5003 using palm kernel cake in a packed‐bed bioreactor. Journal of
Applied Sciences 10: 419–424.
Abdeshahian, P., Samat, N., and Yusoff, W.M.W. (2010d). Utilization of palm kernel cake for
production of β‐glucosidase by Aspergillus niger FTCC 5003 in solid substrate fermentation
using an aerated column bioreactor. Biotechnology 9: 17–24.
Araujo, A. and Ward, O.P. (1990). Extracellular mannanases and galactanases from selected
fungi. Journal of Industrial Microbiology 6: 171–178.
Ashraf, H. and Bilal, Z.H. (2015). Biosynthesis, partial purification and characterization of
invertase through carrot (Daucus carota L.) peels. Journal of Biochemical Technology 6 (1):
867–874.
Azin, M., Moravej, R., and Zareh, D. (2007). Production of xylanase by Trichoderma
longibrachiatum on a mixture of wheat bran and wheat straw: optimization of culture
condition by Taguchi method. Enzyme and Microbial Technology 40: 801–805.
Banerjee, S., Mudliar, S., Sen, R. et al. (2010). Commercializing lignocellulosic bioethanol:
technology bottlenecks and possible remedies. Biofuels, Bioproducts and Biorefining 4: 77–93.
Benech, R.O., Li, X., Patton, D. et al. (2007). Recombinant expression, characterization, and
pulp pre‐bleaching property of a Phanerochaete chrysosporium endo‐β‐1,4‐mannanase.
Enzyme and Microbial Technology 41: 740–747.
Binod, P., Sindhu, R., Madhavan, A. et al. (2017). Recent developments in l‐glutaminase
production and applications – an overview. Bioresource Technology 245: 1766–1774.
Biz, A., Finkler, A.T.J., Pitol, L.O. et al. (2016). Production of pectinases by solidstate
fermentation of a mixture of citrus waste and sugarcane bagasse in a pilot‐scale packed‐bed
bioreactor. Biochemical Engineering Journal 111: 54–62.
Chandel, A.K., Singh, O.V., and Rao, L.V. (2010). Biotechnological applications of
hemicellulosic derived sugars: state‐of‐the‐art. In: Sustainable Biotechnology (eds. S. OV and
H. SP), 63–81. Germany: Springer Science+Business Media.
Couto, S.R. and Sanroman, M.A. (2005). Application of solid‐state fermentation to ligninolytic
enzyme production. Biochemical Engineering Journal 22: 211–219.
Dobrev, G.T., Pishtiyski, I.G., Stanchev, V.S., and Mirchev, R. (2007). Optimization of nutrient
medium containing agricultural wastes for xylanase production by Aspergillus niger B03
using optimal composite experimental design. Bioresource Technology 98: 2671–2678.
Ebringerova, A., Hromadkova, Z., and Heinze, T. (2005). Hemicellulose. Advances in Polymer
Science 186: 1–67.
Fang, H.Y., Chang, S.M., Hsieh, M.C., and Fang, T.J. (2007). Production, optimization growth
conditions and properties of the xylanase from Aspergillus carneus M34. Journal of
Molecular Catalysis B: Enzymatic 49: 36–42.
Fernandes, M.R.V.S. and Jiang, B. (2013). Fungal inulinases as potential enzymes for application
in the food industry. Advance Journal of Food Science and Technology 5: 1031–1042.
Gao, J., Weng, H., Zhu, D. et al. (2008). Production and characterization of cellulolytic enzymes
from the thermoacidophilic fungal Aspergillus terreus M11 under solid‐state cultivation of
corn Stover. Bioresource Technology 99: 7623–7629.
Gomes, J., Gomes, I., Terler, K. et al. (2000). Optimization of culture medium and conditions
for a‐L‐Arabinofuranosidase production by the extreme thermophilic eubacterium
Rhodothermus marinus. Enzyme and Microbial Technology 27: 414–422.
Gomes, J., Terler, K., Kratzer, R. et al. (2007). Production of thermostable β‐mannosidase by a
strain of Thermoascus aurantiacus: isolation, partial purification and characterization of the
enzyme. Enzyme and Microbial Technology 40: 969–975.
Hofrichter, M. (2002). Review: lignin conversion by manganese peroxidase (MnP). Enzyme and
Microbial Technology 30: 454–466.
Howard, R.L., Abotsi, E., van‐Rensburg, E.L.J., and Howard, S. (2003). Lignocellulose
biotechnology: issues of bioconversion and enzyme production. African Journal of
Irmer, J. (2017). Lignin – a natural resource with huge potential. www.biooekonomie‐bw.de/
en/articles/dossiers/lignin‐a‐natural‐resource‐with‐huge‐potential.
Javier, P.F.I., Gallardo, O., Sanz‐Aparicio, J., and Díaz, P. (2007). Xylanases: molecular
properties and applications. In: Industrial Enzymes (eds. J. Polaina and A.P. MacCabe),
65–82. Germany: Springer.
Jiang, Z., Wei, Y., Li, D. et al. (2006). High‐level production, purification and characterization
of a thermostable β‐mannanase from the newly isolated Bacillus subtilis WY34.
Carbohydrate Polymers 66: 88–96.
Katapodis, P., Christakopoulou, V., Kekos, D., and Christakopoulos, P. (2007). Optimization of
xylanase production by Chaetomium thermophilum in wheat straw using response surface
methodology. Biochemical Engineering Journal 35: 136–141.
Kim, D. and Han, G.D. (2014). Fermented rice bran attenuates oxidative stress. In: Wheat and
Rice in Disease Prevention and Health: Benefits, Risks and Mechanisms of Whole Grains in
Health Promotion (eds. R.R. Watson, V.R. Preedy and S. Zibadi), 467–480. UK: Elsevier.
Kote, N.V., Patil, A.G.G., and Mulimani, V.H. (2009). Optimization of the production of
thermostable endo‐β‐1,4 mannanases from a newly isolated Aspergillus niger gr. and
Aspergillus flavus gr. Applied Biochemistry and Biotechnology 152: 213–223.
Kurakake, M. and Komaki, T. (2001). Production of β‐mannanase and β‐mannosidase from
Aspergillus awamori K4 and their properties. Current Microbiology 42: 377–380.
Lakshmi, G.S., Rao, C.S., Rao, R.S. et al. (2009). Enhanced production of xylanase by a newly
isolated Aspergillus terreus under solid state fermentation using palm industrial waste: a
statistical optimization. Biochemical Engineering Journal 48: 51–57.
Latifian, M., Hamidi‐Esfahani, Z., and Barzegar, M. (2007). Evaluation of culture conditions
for cellulase production by two Trichoderma reesei mutants under solid‐state fermentation
conditions. Bioresource Technology 98: 3634–3637.
Li, H., Kim, N.J., Jiang, M. et al. (2010). Simultaneous saccharification and fermentation of
lignocellulosic residues pretreated with phosphoric acid–acetone for bioethanol production.
Li, Y., Cui, F., Liu, Z. et al. (2007). Improvement of xylanase production by Penicillium oxalicum
ZH‐30 using response surface methodology. Enzyme and Microbial Technology 40: 1381–1388.
Lin, S.S., Dou, W.F., Xu, H.Y. et al. (2007). Optimization of medium composition for the
production of alkaline β‐mannanase by alkaliphilic Bacillus sp. N16‐5 using response
surface methodology. Applied Microbiology and Biotechnology 75: 1015–1022.
Lin, T.C. and Chen, C. (2004). Enhanced mannanase production by submerged culture of
Aspergillus niger NCH‐189 using defatted copra based media. Process Biochemistry 39:
1103–1109.
Liu, C., Sun, Z.T., Du, J.H., and Wang, J. (2008). Response surface optimization of fermentation
conditions for producing xylanase by Aspergillus niger SL‐05. Journal of Industrial
Microbiology and Biotechnology 35: 703–711.
Liu, J., Yuan, X., Zeng, G. et al. (2006). Effect of biosurfactant on cellulase and xylanase
production by Trichoderma viride in solid substrate fermentation. Process Biochemistry 41:
2347–2351.
Magalhães, P.O. and Milagres, A.M.F. (2009). Biochemical properties of a β‐mannanase and
β‐xylanase produced by Ceriporiopsis subvermispora during biopulping conditions.
International Biodeterioration and Biodegradation 63: 191–195.
Malherbe, S. and Cloete, T.E. (2002). Lignocellulose biodegradation: fundamentals and
applications. Reviews in Environmental Science and Biotechnology 1: 105–114.
Martins, D.A.B., do Prado, H.F.A., Leite, R.S.R. et al. (2011). Agricultural wastes as substrates
for microbial enzymes production. In: Integrated Waste Management, vol. 2 (ed. S. Kumar),
319–360. London: InTech.
Milagres, A.M.F., Santos, E., Piovan, T., and Roberto, I.C. (2004). Production of xylanase by
Thermoascus aurantiacus from sugar cane bagasse in an aerated growth fermentor. Process
Biochemistry 39: 1387–1391.
Mitchell, D.A., Targonski, Z., Rogalski, J., and Leonowicz, A. (1992). Substrate for processes.
In: Solid Substrate Cultivation (eds. H.W. Doelle, D.A. Mitchell and C.E. Rolz), 29–52. UK:
Elsevier.
Moreira, L.R.S. and Filho, E.X.F. (2008). An overview of mannan structure and mannan‐
degrading enzyme systems. Applied Microbiology and Biotechnology 79: 165–178.
Motta, F.L., Andrade, C.C.P., and Santana, M.H.A. (2013). A review of xylanase production by
the fermentation of xylan: classification, characterization and applications. In: Sustainable
Degradation of Lignocellulosic Biomass (eds. A. Chandel and d.S. SS), 251–275. London:
Intech Open.
Naik, S.N., Goud, V.V., Rout, P.K., and Dalai, A.K. (2010). Production of first and second
generation biofuels: a comprehensive review. Renewable and Sustainable Energy Reviews 14:
578–597.
Niranjane, A.P., Madhou, P., and Stevenson, T.W. (2007). The effect of carbohydrate carbon
sources on the production of cellulase by Phlebia gigantean. Enzyme and Microbial
Pandey, A., Soccol, C.R., and Mitchell, D. (2000). New developments in solid state
fermentation: I‐bioprocesses and products. Process Biochemistry 35: 1153–1169.
Pandey, A., Soccol, C.R., Rodriguez‐Leon, J.A., and Nigam, P. (2001). Solid State Fermentation
in Biotechnology. New Delhi: Asiatech Publishers.
Panesar, P.S., Kaur, R., Singla, G., and Sangwan, R.S. (2016). Bio‐processing of agro‐industrial
wastes for production of food‐grade enzymes: progress and prospects. Applied Food
Park, Y.S., Kang, S.W., Lee, J.S. et al. (2002). Xylanase production in solid state fermentation by
Aspergillus niger mutant using statistical experimental designs. Applied Microbiology and
Patil, S.R. and Dayanand, A. (2006). Production of pectinase from deseeded sunfower head by
Aspergillus niger in submerged and solid‐state conditions. Bioresource Technology 97:
2054–2058.
Petre, M., Zarnea, G., Adrian, P., and Gheorghiu, E. (1999). Biodegradation and bioconversion
of cellulose wastes using bacterial and fungal cells immobilized in radiopolymerized
hydrogels. Resources, Conservation and Recycling 27: 309–332.
Prasertsan, P., Khangkhachit, W., Duangsuwan, W. et al. (2017). Direct hydrolysis of palm oil
mill effluent by xylanase enzyme to enhance biogas production using two‐steps
thermophilic fermentation under non‐sterile condition. International Journal of Hydrogen
Energy 42: 27759–27766.
Rajoka, M.I., Akhtar, M.W., Hanif, A., and Khalid, A.M. (2006). Production and
characterization of a highly active cellobiose from Aspergillus niger grown in solid state
fermentation. World Journal of Microbiology and Biotechnology 22: 991–998.
Ramachandran, S., Patel, A.K., Nampoothiri, K.M. et al. (2004). Coconut oil cake‐a potential
raw material for the production of a‐amylase. Bioresource Technology 93: 169–174.
Ravindran, R. and Jaiswal, A.K. (2016). Microbial enzyme production using lignocellulosic
food industry wastes as feedstock: a review. Bioengineering 3 (30): 1–22.
Sadh, P.K., Duhan, S., and Duhan, J.S. (2018). Agro‐industrial wastes and their utilization
using solid state fermentation: a review. Bioresources and Bioprocessing 5 (1) https://doi.
org/10.1186/s40643‐017‐0187‐z.
Sánchez, C. (2009). Lignocellulosic residues: biodegradation and bioconversion by fungi.

Biotechnology Advances 27: 185–194.
Sá‐Pereira, P., Paveia, H., Costa‐Ferreira, M., and Aires‐Barros, M.R. (2003). A new look at
xylanases: an overview of purification strategies. Molecular Biotechnology 24: 257–281.
Senthilkumar, S.R., Dempsey, M., Krishnan, C., and Gunasekaran, P. (2008). Optimization of
biobleaching of paper pulp in an expanded bed bioreactor with immobilized alkali stable
xylanase by using response surface methodology. Bioresource Technology 99: 7781–7787.
Sharma, K.M., Kumar, R., Panwar, S., and Kumar, A. (2017). Microbial alkaline proteases:
optimization of production parameters and their properties. Journal of Genetic Engineering
& Biotechnology 15: 115–126.
Silva, D., Tokuioshi, K., da Silva Martins, E. et al. (2005). Production of pectinase by solid‐state
fermentation with Penicillium viridicatum RFC3. Process Biochemistry 40: 2885–2889.
Soccol, C.R., Vandenberghe, L.P.S., Spier, M.R. et al. (2014). Enzymes production during
value‐addition of agro‐industrial wastes. In: Enzymes in Value‐Addition of Wastes (eds. S.K.
Brar and M. Verma), 1–38. New York: Nova Science Publishers.
Tengerdy, R.P. and Szakacs, G. (2003). Bioconversion of lignocellulose in solid substrate
fermentation. Biochemical Engineering Journal 13: 169–179.
Thomas, L., Parameswaran, B., and Pandey, A. (2016). Hydrolysis of pretreated rice straw by an
enzyme cocktail comprising acidic xylanase from Aspergillus sp. for bioethanol production.
Xin, F. and He, J. (2013). Characterization of a thermostable xylanase from a newly isolated
Kluyvera species and its application for biobutanol production. Bioresource Technology 135:
309–315.
203
10
Production of Organic Acids Via Fermentation of Sugars

Generated from Lignocellulosic Biomass
Lourdes Zumalacárregui de Cárdenas and Beatriz Zumalacárregui de
Cárdenas
Chemical Engineering Department, Chemical Engineering Faculty, Technological University of Havana, Havana, Cuba
10.1 Introduction
The generation of biochemical products from lignocelluloses is receiving attention from

researchers and producers because of its wide global availability (Perez et al. 2013; Shen
et al. 2014; Ganigué et al. 2016). The abundance and low cost of these materials make them
particularly attractive feedstocks to replace finite fossil resources and reduce their negative
impact on the environment (Saini et al. 2014). High capital and operating expenditures,
irregularities in biomass supply chain, technical process immaturity, and scale‐up chal-
lenges limit the commercialization of renewable products at the present time (Chandel
et al. 2018).
Three types of biopolymers – cellulose, hemicellulose, and lignin – appear in lignocel-
lulose composition. A typical composition (dry basis) is 20–50% cellulose, 15–35% hemicel-
lulose, and 10–30% lignin. In addition, lignocellulose contains smaller amounts of other
minor components: proteins (3–10%), lipids (1.5%), soluble sugars, and minerals (10.5%),
which in chemical analyses are estimated as ash (Rinaldi and Schuth 2009; Chundawat
et al. 2011). To be used as a renewable raw material for production of various bioproducts,
it is necessary to carry out hydrolysis of the cellulose and hemicellulose chains to obtain
glucose and xylose, respectively, which will serve as substrates for a fermentation process.
Different pretreatment strategies have been proposed for biomass processing. Knowing
the structural characteristics of lignocellulosic biomass and changes during pretreatment
can allow cost reductions to be made (Wi et al. 2011). The specific pretreatment type
depends on the lignocellulosic material to be employed. Literature reports diverse pretreat-
ment methods grouped into three categories: physicochemical, hydrothermal, and bio-
chemical (Galbe and Zacchi 2002; Saska and Martin 2006; Martín and Carrillo 2009;
Carrillo‐Nieves et al. 2014).
Biological pretreatment with enzymes is being studied. Biological removal of some
specific components, for example antimicrobial substances, has been determined. The

enzymes are molecules of protein, able to catalyze chemical reactions; enzymes can
locate a chemical specific group and act selectively in a single enantiomer or diastere-
omer, without secondary reactions. The activity can be regulated by inhibitors or activa-
tors (Gil et al. 2010).
Many enzymes have been studied, as well as genetically modified fungus and bacteria
that ferment all the biomass into sugars (Taherzadeh and Karimi 2008; Matano et al. 2012).
The high specificity of enzymatic reactions is sensitive to changes in the structural proper-
ties of the cellulosic substrates. Research demonstrates that differences in the structural
properties such as porosity (superficial accessible area), pore dimension, cellulose crystal-
linity, humidity, polymerization grade, and lignin and hemicellulose content (Gil et al.
2010) influence the enzymatic degradation process, hindering the accessibility of the
enzymes and reducing the efficiency of the hydrolysis (Alvira et al. 2010; Eisentraut 2010).
Biological methods have some advantages such as selectivity, higher yield without the
inconvenience of the formation of collateral products, low energy demands, low chemical
reagent consumption, and eco‐friendliness. For these reasons, the use of enzymes has
proliferated for the conversion of cellulose to units of glucose. The main limitations of
enzymatic hydrolysis are the high cost of the enzymes and the need for big reactors due to
the slowness of the reaction. Biorefinery processes based on microbial fermentation need
to be economically competitive with petroleum refinery processes so it is important to
maximize the performance of microorganisms with respect to titer, productivity, and yield
of desired product.
Different strategies have been investigated for enzymatic hydrolysis to overcome these
limitations. Genetic engineering, enzyme recycling, high solid loadings, pretreatment tech-
nologies, bioreactor designs, and nanomaterials to improve the thermal and pH stability of
cellulases are being studied. In order to improve commercial scale‐up, bioreactor configu-
rations are being studied, considering efficient mixing, sufficient mass transfer, low shear
stress, low foaming problems, and low consumption of water and energy. Bioreactor types
including stirred tank, rotating tubular, airlift, membrane, reciprocating plate, and solid‐
state fermentation have been reviewed for cellulase production (Shokrkar et al. 2018).
Currently, economical commercial production of biochemicals and biofuels with is
still insufficient. Efforts to produce chemicals are under way by companies like DSM
(succinic acid, cellulosic ethanol), Dow‐DuPont (1,3‐propanediol, 1,4‐butanediol),
Clariant‐Global Bioenergies‐INEOS (bio‐isobutene), Braskem (ethylene, polypropylene),
Raizen, Gran‐Bio and POET‐DSM (cellulosic ethanol), Amyris (farnesene), and others
(Chandel et al. 2018).
Production of organic acids by fermentation is a fast‐moving field. Acids such as glyceric,
glucaric, succinic, butyric, xylonic, fumaric, malic, itaconic, lactobionic, propionic, and
adipic are being studied through innovative fermentation strategies, mainly at laboratory
scale (Alonso et al. 2015). Industrial organic acids, as citric acid and lactic acid, are being
produced by fermentation of lignocellulosic materials (Liu et al. 2014; Chandel et al. 2018).
Metabolic engineering strategies could produce new strains with high performance, such
as those developed for succinic acid and 3‐hydroxypropionic acid production (Chandel
et al. 2018). More research is necessary at laboratory and commercial levels, in order to
produce these emerging bioproducts via large‐scale fermentation and fulfill the petroleum
refinery role.
Production of Organic Acids Via Fermentation of Sugars Generated from Lignocellulosic Biomass 205
10.2 Material Products of a Biorefinery
Material products of a biorefinery are those that are not used for energy purposes, but that
could be converted into valuable materials to reduce dependence on the production of
chemicals from fossil fuels. The different biomass transformation technologies are inte-
grated into platforms. The platform of sugars and lignocellulose interacts with the other
three platforms (thermochemistry platform, lipid platform (oils and fats) and other plat-
forms (proteins, biogas, etc.), showing alternative routes and synergy between them.
Several platforms can coexist in the same biorefinery.
In the sugars and lignocellulose platform, there are several stages that can be integrated
(de la Rosa 2015).
●● Hydrolysis of polysaccharides (cellulose and hemicellulose) to monosaccharides of five
and six atoms, such as xylose or glucose.
●● Conversion of glucose to intermediate chemical compounds such as ethanol, butanol,
organic acids, using fermentation and other conventional chemical transformations.
●● Conversion of xylose to products such as ethanol, xylitol, and furfural, using fermenta-
tion and other transformations.
●● Valorization of lignin and other waste.
As seen in Figure 10.1, the hydrolysis of cellulose and hemicellulose generates mono-
meric sugars, which are the main actors in biorefineries, obtaining value‐added products
by chemical, thermochemical or biological processes. However, hydrolysis of the biomass
is a reaction that, depending on the conditions and type of pretreatment, could degrade the
monosaccharides, forming organic acids such as formic acid, levulinic acid, furfural, and
5‐hydroxymethylfurfural (Ramos 2003).
The use of biocatalyst technologies can broaden the catalog of chemical products that
can be produced in a biorefinery, starting from either the sugars obtained from the biomass
Lignocellulosic material
Cellulose Hemicellulose Lignin
Glucose Mannose Galactose Xylose Arabinose
Hydroxymethylfurfural Furfural Phenols
Levulinic acid Formic acid
Figure 10.1 Biomass hydrolysis and formation of inhibitor compounds.

or the primary fermentation products. Biocatalysis is the application of biocatalysts or

enzymes for the synthesis, interconversion or degradation of chemical products. Biocatalysts
are characterized by their high specificity and activity, which implies that the processes
catalyzed by them generally require lower consumption of raw materials and generate
lower amounts of by‐products and residues than the corresponding chemical processes. In
addition, biocatalytic processes are usually carried out at much lower temperatures and
pressures than chemical processes, so they need less energy.
When enzymes are used in the transformation process, it must be borne in mind that
conventionally, yeasts are not capable of transforming the pentoses (xylose and arabinose),
which reduces the efficiency and profitability of the process. Metabolic engineering has
allowed the creation of new strains of recombinant microorganisms, such as the bacterium
Zymomonas mobilis that can ferment glucose and pentoses.
In order to integrate the process, a consolidated bioprocess has been studied that inte-
grates the stages of saccharification, fermentation, and in situ production of hydrolytic
enzymes, so that all the processes are carried out simultaneously in a single step and in a
single reactor. This avoids the problem of inhibition by products associated with the enzy-
matic activity that occurs when, during the enzymatic saccharification process, the accu-
mulation of glucose causes inhibition of the enzyme β‐glucosidase, which ceases to
hydrolyze the cellobiose, which is thus accumulated in the culture medium. This increase
in cellobiose concentration inhibits the hydrolysis of cellulose, affecting the efficiency of
the saccharification process. Integration prevents the accumulation of fermentable sugars,
increasing the yield (Lynd et al. 2002).
A team from Pacific Northwest National Laboratory (PNNL) and National Renewable
Energy Laboratory (NREL) proposed a list of 12 potential biobased chemicals, starting
from more than 300 candidates. The list includes 1,4‐diacids (succinic, fumaric, and malic),
2,5‐furan dicarboxylic acid, 3‐hydroxypropionic acid, aspartic acid, glucaric acid, glutamic
acid, itaconic acid, levulinic acid, 3‐hydroxybutyrolactone, glycerol, sorbitol, and xylitol/
arabinitol (Werpy et al. 2004; Kamm and Kamm 2007; FEDIT 2008).
10.3 Organic Acid Production in Biorefineries
Carboxylic acids are high‐value compounds used in large volumes mainly as precursors for
the chemical industry (Liew et al. 2016). The production of these chemicals from microor-
ganisms has been normal practice, and they are the third largest class of bulk chemicals in
the global market obtained via biological means, with a projected growing demand (Daniell
et al. 2012; Liew et al. 2016).
Short‐chain carboxylic compounds such as acetic, propionic and butyric acids can be
synthesized naturally during the biodegradation of organic substrates (Kim et al. 2011).
Similarly, these compounds can be produced by metabolism mainly as intermediate prod-
ucts or occasionally as the major products by microorganisms using inorganic substrates
such as syngas (Henstra et al. 2007; Daniell et al. 2012). Syngas is a term used to describe a
gaseous mixture that consists mainly of CO and H2 (Bain and Broer 2011). It is generated
during a thermal process known as gasification that transforms any carbon‐based material
into gaseous products without burning it. Depending on the process conditions, the final
product of gasification may also contain CO2, CH4, N2, traces of other low molecular weight
hydrocarbons, tars, and ash (Munasinghe and Khanal 2010; Bain and Broer 2011).
Production of acetic acid from a gaseous substrate was first reported in 1939. A mixture
of H2 and CO2 was converted biologically into acetic acid by Clostridium aceticum. The
optimum pH range for this process was reported as 8–9, which is higher than the range
recommended for the fermentation of organic materials (Visioli et al. 2014). In another
report, Sim et al. (2007) found that C. aceticum can also consume CO to produce acetic acid.
This study concluded that the fermentation time significantly influenced the amount of
biomass and the total acetic acid produced.
The 10 most promising organic acids from that list that can be obtained from biomass
are:
●● succinic acid
●● levulinic acid
●● fumaric acid
●● L‐aspartic acid
●● L‐malic acid
●● itaconic acid
●● glucaric acid
●● 3‐hydroxypropionic acid
●● 2,5‐furan‐dicarboxylic acid
●● L‐glutamic acid
Other organic acids including gluconic and adipic have also been studied. A more exten-
sive report of this analysis can be found in Chandel and Silveira (2018). This chapter will
focus on the processes that are carried out enzymatically to obtain the above‐mentioned
organic acids. The chemical structures of each of these acids are presented in Table 10.1.
10.3.1 Succinic Acid

Succinic acid (SA) or butanodioic acid (C4H6O4) is a C4‐dicarboxylic acid which is part of
the Krebs cycle, and can be obtained from microbiological fermentation and used to syn-
thesize several polymers of commercial importance. Methylene succinic acid was origi-
nally known as a product of pyrolytic distillation of citric acid. In the 1940s, it was found
that this acid could be produced by fermentation using Aspergillus terreus. Its current
industrial production is by a fermentation process, using fermentable sugars from corn or
sugarcane as substrates. Nowadays, nonfood raw materials are being studied. Natural suc-
cinate producer strains such as Actinobacillus succinogenes, Basfia succiniciproducens, and
Mannheimia succiniciproducens and engineered strains such as Escherichia coli and
Corynebacterium glutamicum have been specifically used as microbial platforms (Alonso
et al. 2015; Choi et al. 2015). The common characteristics of the native strains are that they
grow in anaerobic conditions and produce succinic acid as the final product.
Sugarcane bagasse (SCB) hemicellulosic acid hydrolyzate was used as a carbon source
for SA production by A. succinogenes (Xi et al. 2013). The maximal succinic acid produc-
tion (19.7 g/L) and yield (65.7%) were achieved by using 30 g/L of total sugar concentra-
tion in detoxified hydrolyzate. When fermentation was carried out using nondetoxified
Table 10.1 Chemical structures of the 10 most promising organic acids.
Succinic acid O
HO
OH
O
Levulinic acid O
OH
Fumaric acid O
HO
OH
O
L‐aspartic acid O
HO
OH
O NH2
L‐malic acid O
HO
OH
O OH
Itaconic acid O
HO
OH
O
Glucaric acid OH OH O
HO
OH
O OH OH
3‐Hydroxypropionic acid O
HO OH
2,5‐Furan‐dicarboxylic acid HO OH
O
O O
L‐glutamic acid O O
HO OH
NH2
Gluconic acid OH OH O
HO
OH
OH OH
ydrolyzate containing 30 g/L total sugar, 23.7 g/L SA concentration was achieved with a
h
79% SA yield.
The use of SCB hemicellulosic hydrolyzate for SA production by A. succinogenes was also
reported by Borges and Pereira (2011). In their study, hemicellulosic acid hydrolyzate contain-
ing 52 g/L xylose was fermented with a continuous supply of carbon dioxide into the medium
at 0.05 vvm for 24 hours at 37 °C, pH 7.0, and 150 rpm. In that condition, the SA concentration
and product yield in relation to substrate were 22.5 g/L and 0.43 g/g, respectively.
An engineered M. succiniciproducens strain LPK7 was developed, producing 52.4 g/L suc-
cinic acid with yield 0.76 g/g glucose and productivity 1.8 g/L/h in a complex medium. A
higher yield (0.86 g/g glucose) was obtained in a defined medium later with the same strain.
Further metabolic engineering of M. succiniciproducens strain allowed homosuccinic acid
production with titer, yield, and productivity about 90–100 g/L, 0.9–1.0 g/g glucose or
sucrose, and 5–30 g/L/h, respectively. For B. succiniciproducens, the highest yield reported
was 0.76 g/g glucose obtained using an encoding lactate dehydrogenase (ldhA) and encod-
ing pyruvate formate lyase (pflB) deleted mutant (Choi et al. 2015).
Under large‐scale conditions, recombinant E. coli has been chosen as the preferred strain
for succinate production due to its high substrate tolerance and ease of cultivation (Thakker
et al. 2012). Sugarcane molasses (Chan et al. 2012) or soybean meal hydrolyzate (Thakker
et al. 2013) have been used as carbon sources by an engineered strain of E. coli.
Succinate titer of 56 g/L with volumetric productivity of 0.78 g/L/h under anaerobic con-
ditions was reported by Chan et al. (2012). In addition to these approaches, a wheat‐based
biorefining strategy has been proposed as an innovative approach for the fermentative pro-
duction of SA (Lin et al. 2011). However, the fact that the saccharification process has yet
to be optimized may constitute a drawback for high yield of the proposed strategy.
An engineered E. coli strain was employed for SA production in a one‐step fermentation
process, deleting ldhA, encoding alcohol dehydrogenase (adhE), pflB, encoding formate
transporter (focA), pta‐ackA, methylglyoxal synthase (mgsA), encoding pyruvate oxidase
(poxB), encoding threonine decarboxylase (tdcD), encoding 2‐ketobutyrate formate‐lyase
(tdcE), encoding citrate lyase (citF), encoding aspartate aminotransferase (aspC) and
encoding malic enzyme (sfcA) genes in order to eliminate by‐products. The engineering
strain produces 71.6 g/L succinic acid with yield and productivity of 1.00 g/g glucose and
0.75 g/L/h (Jantama et al. 2008).
Saccharomyces cerevisiae was engineered by the deletion of encoding pyruvate decar-
boxylase (pdc), encoding fumarate hydratase (fum) and GPD (encoding glycerol 3‐phos-
phate dehydrogenase (gpd) genes with overexpression of encoding malate dehydrogenase
(mdh) and encoding pyruvate carboxylase (pyc) genes. 13 g/L of succinic acid with 0.14 g/g
glucose and 0.108 g/Lh at pH 3.8 was obtained (Yan et al. 2014).
Patent EP 2297297 (Jansen and Verwaal 2017) shows the process for the preparation of
SA at low pH, which comprises yeast fermentation with a carbohydrate substrate and low
amounts of oxygen at a pH value (2–4) which is below the lowest pKa of SA, wherein the
oxygen is supplied at a specific oxygen absorption rate ranging from 8 to 0.2 mmol oxygen/g
dry weight biomass/h. The acid production is preceded preferably by a biomass production
phase at pH 4–5. The yeast used in the process is genetically modified.
Microbial production of SA has been successfully transferred to the industrial scale.
Some industrial facilities are shown in Table 10.2.
Table 10.2 Industrial plants for succinic acid production.
Location Owner Raw material Technology
Cassano Reverdia Starch and sugars Hydrolysis and fermentation

Biorefinery with yeasts at low pH
Lake Myriant Renewable raw materials such One‐step anaerobic
Providence (now GC as sorghum, sugarcane bagasse fermentation process
Biorefinery Innovation (SCB), and other cellulosic development by Myriant
America) materials
Montmeló Succinity Renewable substrates (sugars, Fermentation process
Biorefinery GmbH glycerin) (patented) using Basfia
succiniciproducens
Sarnia BioAmber and Glucose Fermentation technology
Biorefinery Mitsui and Co using a patented yeast
Source: Choi et al. (2015) and Bio Refineries Blog (2018).
From a chemical point of view, the compound of renewable origin is identical to the one
manufactured in a conventional manner, so it has the same properties and can be used in
the same applications. Succinic acid has a multitude of potential uses.
●● Production of polybutylene succinate (PBS), a modern biopolymer whose applications
(compostable films, disposable cups, plastic bags, etc.) are still being studied.
●● Production of plasticizers for the manufacture of PVC. It can also be used to cover the
growing demand for plasticizers for bioplastics.
●● Replacement of the petrochemical products derived from adipic acid used in the produc-
tion of polyester polyols for polyurethanes (adhesives, coatings, sealants, shoe soles, flex-
ible and rigid foams).
●● Production of 1,4‐butanediol (BDO) for subsequent production of tetrahydrofuran (elas-
tane fibers) and polybutylene terephthalate (electrical equipment, wheel covers, etc.).
●● Production of dimethyl‐succinate (DMS), a green solvent, miscible with alcohols,
ketones, and most of the hydrocarbons. It is used as a coalescing agent for emulsion of
paints with low contents of volatile organic compounds
Succinic acid can also be used as a chemical platform to be transformed by simple chemi-
cal processes into 1,4‐butanediol (1,4‐BDO), γ‐butyrolactone (GBL), tetrahydrofuran
(THF), and N‐methylpyrrolidone (NMP) (Choi et al. 2015; Tsuge et al. 2016).
10.3.2 Levulinic Acid

Levulinic acid (LA) or 4‐oxopentanoic acid is an organic compound with the formula
CH3C(O)CH2CH2CO2H or C5H8O3. LA is a short‐chain fatty acid with a ketone carbonyl
and an acidic carboxyl group, which has wide applications in industry and agriculture to
produce a variety of products such as plasticizers, oil additives, and fragrances. Up to now,
production of LA has been broadly investigated from a variety of feedstocks such as sugars,
cellulose, chitin, agricultural waste, and raw lignocellulosic biomasses by homogeneous or
heterogeneous catalysts. Heterogeneous catalysts are still not satisfactory in LA production,
especially from cellulose and raw biomasses feedstocks, due to the solid–solid mass trans-
fer limitation (Jeong 2014). Homogeneous catalysts with mineral acids have the disadvan-
tages of equipment corrosion and difficulty in recycling.
The controlled degradation of C6 sugars by acids is the most commonly used process to
prepare LA from lignocellulosic biomass. Other methods have been studied, including the
hydrolysis of acetylsuccinate esters, acid hydrolysis of furfuryl alcohol, and oxidation of
ketones. However, these methods require high‐cost raw materials and give rise to relatively
high quantities of by‐products.
When cellulose or raw lignocellulosic biomasses are used as starting materials, LA is usu-
ally produced by the acid‐catalyzed conversion of cellulose, following a reaction pathway
in which cellulose is hydrolyzed into glucose via enzymatic or acid‐catalysis methods, and
the generated glucose is dehydrated to 5‐hydroxymethylfurfural (HMF), followed by a
rehydration step converting HMF to LA.
The following companies have developed technologies following this model: Biofine,
DSM, GFBiochemicals, and Segetis (acquired by GFBiochemicals). The Biofine process
involves the use of dilute sulfuric acid as a catalyst but it differs from other dilute‐acid lig-
nocellulosic‐fractionating technologies in that free monomeric sugars are not the product.
Instead, the 6‐carbon and 5‐carbon monosaccharides undergo multiple acid‐catalyzed
reactions to give the platform chemicals LA and furfural as the final products.
Hydroxymethylfurfural is an intermediate in the production of levulinic from 6‐carbon
sugars in the Biofine process. The process then consists of two distinct acid‐catalyzed stages
that give optimal yields with a minimum of degradation products and tar formation. The
maximum theoretical yield of LA from a hexose is 71.6% w/w and formic acid makes up the
remainder. The Biofine process, due to its efficient reactor system and the use of polymeri-
zation inhibitors that reduce excessive char formation, achieves from cellulose LA yields of
70–80% of the theoretical maximum. These claims have been supported by process data
from a pilot plant located in Glens Falls, New York State. This processes one dry ton of
feedstock per day and has been operational for several test run periods since 1996 (Hayes
et al. 2006).
Cow dung has been used as feedstock for the production of a high value‐added chemical
LA in dilute HCl acid aqueous solutions. Crude cow dung could afford a LA yield of 135 g/
kg at 180 °C in a 210 minute reaction time, and the LA yield could be improved to 338.9 g/
kg after the cow dung was pretreated with KOH aqueous solution, ascribed to enhance-
ment of the accessibility of cow dung to the acid sites in the catalytic reaction, due to the
removal of the lignin fraction from the lignocellulose structure of the dung by the KOH
solution in the pretreatment step. In addition, a formic acid yield of c.160 g/kg could be
achieved in the process, indicating that a total yield of c.500 g/kg could be obtained for LA
and formic acid from pretreated cow dung in the proposed reaction system. For LA, the
value is 80.5% based on the theoretical maximum yield and for formic acid this value is
95.6% of the theoretical maximum yield. Compared with the conventional utilization
method for cow dung, this work provides a promising strategy for the value‐increment
utilization of cow dung (Su et al. 2017).
Recently, a United States patent has been published showing a fermentation route for the
production of LA, levulinate esters and valerolactone and derivatives thereof. The method
comprises multiple enzymatic steps integrated into a single metabolic pathway in a
f ermentation host such as Sacharomyces spp., Pichia spp., Pseudomonas spp., Bacillus spp.,
Chrysosporium spp., and E. coli. The invention converts two molecules of pyruvate into one
C5 molecule such as LA and 4‐valerolactone. For C6 sugars, the carbon yield can be 80%.
For C5 sugars, the carbon yield can be theoretically 100%. The direct fermentation of sugars
to LA, 4‐valerolactone or any other C5 derivatives could be produced, if the microbial strain
could transform C5 and C6 such as recombinant S. cerevisiae and Pichia stipitis. The mole-
cule yield is considerable higher than for the chemical transformation (40% molar yield or
less) (Zanghellini 2017).
Levulinic acid is a very versatile platform for obtaining chemical products and materials
derived directly from biomass. It is used as a precursor in the following processes.
●● Fuel additives: levulinate esters are additives for diesel and gasoline. Methyltetrahydrofuran
(MeTHF), a derivative of LA, can be mixed with up to 50% gasoline to increase vehicle
performance and reduce emissions of polluting gases.
●● Solvents: the esters of LA, γ‐valerolactone (GVL), and MeTHF are suitable solvents for
many applications. GVL can replace ethyl acetate and MeTHF can be used as a substitute
for tetrahydrofuran (THF) in fine chemistry and in the pharmaceutical industry.
●● Resins and coatings: LA can be used in polyester resins and polyester polyols to increase
the scratch resistance of coatings for exteriors and interiors. Diphenolic acid (DPA) is
used in finishes for protection and decoration.
●● Agrochemicals: δ‐aminolevulinic acid (DALA) is used as a herbicide for pastures and
certain crops.
●● Polymers and plasticizers: esters of ketals derived from LA can replace most phthalate‐
based plasticizers. Methylbutanediol (MeBDO) has potential as a monomer for polyure-
thanes. GVL can be a monomer for polyesters and a precursor of the isomers of
pyrrolidone.
●● Pharmaceutical products: LA is used in antiinflammatory medications, antiallergenic
agents, mineral supplements, and transdermal patches. DALA is used in the diagnosis
and treatment of cancer.
●● Cosmetic products: LA and its derivatives are used in organic and natural cosmetics for
perfumes, skin conditioners, and pH regulators.
10.3.3 Fumaric Acid

Fumaric acid is an organic dicarboxylic compound, with molecular mass 116.07 and
density20/4°C 1.635 g/cm3. It is also known as 2‐butenedioic acid, trans‐1,2‐ethylenedicarbo-
xylic acid, algalelic acid, boleic acid, lichenic acid or tumaric acid. It was isolated for the
first time from the plant Fumaria officinalis, from which it derives its name (Roa Engel
et al. 2008).
Fumaric acid is usually produced by chemical synthesis and is mainly used as an acidu-
lant in the food industry (35% of world consumption) due to its nontoxic character and
nonhygroscopic properties. It is used to adjust pH, enhance flavor, and extend shelf‐life by
controlling the growth of microorganisms. Other significant end‐uses for fumaric acid
include unsaturated polyester resins (19% of world consumption), rosin paper sizes (18%),
alkyd resins (11%), and animal feed (5%) (HIS 2017).
The beginning of fumaric acid production by biotechnological routes occurred in 1911

after the discovery of fumaric fermentation with Rhizopus nigricans by Felix Ehrlich. In
1938, Foster and Waksman studied 41 strains of eight different genera to identify the strain
with the highest productivity of fumaric acid. The identified fumarate production genera
were Rhizopus, Mucor, Cunninghamella, and Circinella. Of these strains, Rhizopus species
(nigricans, stolonifer, arrhizus, oryzae, and formosa) have the best results, producing fuma-
ric acid under aerobic and anaerobic conditions (Giraldo 2013) while fumaric acid as a
minor fermentation product can be excreted by other microorganisms, such as S. cerevisiae
and some Aspergillus niger and A. flavus strains. Genetically engineered bacteria have been
used for the production of fumaric acid with yields up to 85% w/w from glucose.
Fumaric acid is primarily an intermediary of the Krebs cycle and is also involved in other
metabolic pathways. In 1969, Overman and Romano discovered a mechanism for the pro-
duction of fumaric acid that involved the condensation of a three‐carbon acid (C3) plus one
carbon (C1), and involved the catalytic fixation of CO2 with pyruvate carboxylase under
aerobic conditions, explaining the high molar yields in fumarate production. The fixation
of CO2 leads to the formation of oxalic acid so that the organic acids of four carbons (C4),
intermediates of the Krebs cycle, can be removed by biosynthesis during the growth phase
of the microorganism under aerobic conditions. When nitrogen is limited and the microor-
ganism ends the growth phase, glucose metabolism and CO2 fixation can continue and lead
to the accumulation of C4 acids. However, if reductive pyruvate carboxylation was the only
route for production, no ATP would be produced for maintenance and transport purposes,
so the Krebs cycle is also active during the production of fumaric acid. Carbon labeling
studies show that the two routes, the Krebs cycle and reductive pyruvate carboxylation,
under aerobic conditions are used simultaneously in the production of fumaric acid (Roa
Engel et al. 2008).
Main carbon flows during fumaric acid production can be seen in Figure 10.2. The inner
block represents the citrate cycle reactions inside the mitochondria. The outer block repre-
sents the cytoplasmic membranes. Carbon‐rich substrates are required as a carbon source
for the fermentation of carbohydrates. Commonly, glucose has been used as a substrate but
it has not been the only carbon source used in fermentation. The use of different raw materi-
als such as xylose, sucrose, starch and agro‐industrial residues such as molasses, cassava
bagasse, and dairy manure has been studied to evaluate their feasibility for fumaric acid
production (Kautola and Linko 1989; Carta et al. 1999; Zhang et al. 2007; Liao et al. 2008; Xu
et al. 2010). Fumaric acid production by fermentation must be carried out at an optimum
temperature and pH, according to the type of microorganism, an adequate concentration of
the substrate and nutrient salts based on nitrogen and essential metals (Giraldo 2013).
A US patent describes a submerged culture process for fumaric acid production by fer-
mentation of a carbohydrate‐containing substance by means of Rhizopus oryzae in the
presence of a nitrogen‐containing nutrient within the range of one part of nitrogen per 25
to 300 parts of the carbohydrate (as carbon). An improvement comprises initiating the
fermentation in the presence of substantially all the carbohydrate‐containing substance
but only from about 10–75% by weight of the total required amount of the nitrogen‐
containing nutrient, and adding the remainder of the nitrogen‐containing nutrient during
the course of fermentation. Fumaric acid yields of about 50% were obtained (La Roe 1959).
Figure 10.2 shows the flowchart for the production of fumaric acid.
CO2 Ethanol
Glucose
Glucose Ethanol
CO2 CO2 CO2
Oxaloacetate Pyruvate Acetaldehyde
Pyruvate CO2 CO2

Malate
Acetyl CoA
Oxaloacetate Citrate Iso- CO2

citrate
Fumarate Malate Oxoglutarate
Fumarate Succinate
CO2
Fumarate
Figure 10.2 Main carbon flows during fumaric acid production.
Alternative substrates such as corn straw hydrolyzates, composed mainly of glucose and
xylose, have also been used for fumaric acid production by R. oryzae, obtaining a fumaric
acid titer of 27.8 g/L, yield of 0.35 g/g glucose, with a volumetric productivity of 0.33 g/L/h
(Zhang et al. 2007). For enhancement of the bioconversion ability of R. oryzae, both endog-
enous pyruvate carboxylase and exogenous phosphoenolpyruvate carboxylase were overex-
pressed in order to increase yields by 26% (24 g/L) (Zhang 2012). This strong overexpression
has specifically enabled an increase in the carbon flux toward fumaric acid production,
suggesting that further metabolic engineering improvements may lead to industrially sig-
nificant fumaric acid titers from renewable sources (Xi et al. 2013).
Rhizopus oryzae ATCC 20344 can use dairy manure hydrolyzate (lignocellulosic materi-
als) supplemented with glucose to produce 31 g/L fumaric acid with a yield of 31% (La Roe
1959). In this process, enzymatic hydrolysis of pretreated lignocellulosic material was per-
formed before fermentation by R. oryzae in what was described as separated hydrolysis and
fermentation (SHF). However, SHF may have several problems, such as end‐product inhi-
bition of enzymatic hydrolysis, slow hydrolysis rate, and low yield and product concentra-
tion. A simultaneous saccharification and fermentation (SSF) process could afford
enzymatic hydrolysis of lignocellulose and fermentation of released sugars in the same
vessel and hence overcome the problems associated with SHF.
Production of fumaric acid from alkali‐pretreated corncob (APC) at high solids loading
was investigated using a combination of SHF and fed‐batch SSF by R. oryzae, the most effi-
cient fumarate producer strain. Four different fermentation modes were tested to maxi-
mize fumaric acid concentration at high solids loading. The highest concentration of
41.32 g/L fumaric acid was obtained from 20% (w/v) APC at 38 °C in the combined SHF and
fed‐batch SSF process, compared with 19.13 g/L fumaric acid in batch SSF alone. The
results indicated that a combination of SHF and fed‐batch SSF significantly improved pro-
duction of fumaric acid from lignocellulose by R. oryzae than that achieved with batch SSF
at high solids loading (Li et al. 2016).
Fumaric acid can be used in corn tortillas, wheat, sourdough, fruit juice, rye breads, refrig-
erated biscuit dough, nutraceutical drinks, gelatin desserts, pie fillings gelling aids, wine, and
as a supplement in animal feed. Fumaric acid increases value and decreases costs of most
food items and beverage products. It also increases feed efficiency for poultry and pigs. A very
different application of fumaric acids is in a medicine to treat psoriasis (Khan et al. 2017).
10.3.4 Aspartic Acid

Aspartic acid is a 4‐carbon amino acid that is an essential part of human protein produc-
tion. Several configurations of aspartic acid exist but L‐aspartic is the most common. L‐
aspartic acid is primarily used to produce aspartame, a synthetic sweetener. Aspartic acid
was discovered in 1827 by Auguste‐Arthur Plisson and Etienne Ossian Henry, as a deriva-
tive of asparagine, isolated from asparagus juice in 1806, by boiling with a base. Aspartic
acid or its ionized form, aspartate (symbols Asp and D), is one of the 20 amino acids with
which cells form proteins. In RNA, it is encoded by the GAU or GAC codons. It has a car-
boxyl group (─COOH) at the end of the side‐chain. Its developed chemical formula is
HO2CCH (NH2) CH2CO2H.
At physiologic pH, it has a negative charge (it is acid); it belongs to the group of amino
acids with charged polar side‐chains. Its biosynthesis takes place by transamination of oxa-
lacetic acid, an intermediate metabolite of the Krebs cycle. It is also synthesized from
phthalimidomalonate [C6H4 (CO) 2 NC (CO2Et)2]. Aspartate participates in the formation
of glutamate through cytosolic glutamate‐aspartate transaminase. Aspartate is also a
metabolite of the urea cycle and participates in gluconeogenesis.
There are four primary routes to produce L‐aspartic acid: chemical synthesis, protein
extraction, fermentation, and enzymatic conversion. The preferred method currently is the
enzymatic route, reacting ammonia with fumaric acid, catalyzed by a lyase enzyme. The
advantages of this pathway include high product concentration, high productivity, fewer
by‐products, and easy separation (crystallization).
The major technical obstacle for the development of aspartic acid as a building block is
related to the selection of a fermentation route (using sugar substrate) that is cost‐
competitive with the existing enzymatic conversion process. Direct fermentation routes are
not cost‐competitive yet, but the use of biotechnology could overcome this obstacle.
Moreover, the engineering pathway of biocatalytic organisms should overproduce oxaloac-
etate without compromising viability of organisms. A second strategy for reducing the cost
of aspartic acid is to make improvements to the current technology. The primary focus of
this effort would be reducing the cost of fumaric acid, which is currently used as the feed-
stock for producing aspartic acid.
High fermentation yields and product recovery are the two primary technical goals to
strive for. A direct fermentation using sugar substrate could potentially be cheaper than
fumaric acid and ammonia feedstocks, if the technical production performance can be
improved. Bayer AG also has a competing route using maleic anhydride that may be
otentially cheaper. Producing fumaric acid at lower cost could have a near term impact on
p
reducing the cost of aspartic acid. This strategy would have the advantage of using existing
capital and infrastructure. Productivity improvements are required to reduce the capital
and operating costs of the fermentation. The existing enzyme route through fumaric acid
achieves productivities which are satisfactory for specialty applications but for commer-
cial‐scale applications, further improvements in productivity will be required.
By enzymatic synthesis, a one‐phase process for the fermentative production of L‐aspartic
acid in a medium containing fumaric acid and ammonia was developed by Chibata in 1965.
The microorganisms were Pseudomonas trifolli B269‐PY‐5 and yield was higher than 35%
(Praveen et al. 2017). The best known biological method to obtain L‐aspartic acid is by an
enzymatic process using fumaric acid as the starting material in the presence of ammo-
nium and with the action of the enzyme aspartase or an aspartase‐producing microorgan-
ism. A long list of microorganisms have aspartase activity, including E. coli, Pseudomonas
fluorescens, Proteus vulgaris, Serratia marcescens, Salmonella spp., Vibrio spp.,
Staphylococcus spp., Aerobacter spp., Bacterium cadaveris, Lactobacillus helveticus, and
Alcaligenes metalcaligenes. Figure 10.3 shows fumaric acid metabolism. Fumaric acid can
be converted to aspartic acid (target product) by the activity of aspartase and to L‐malic
acid (by‐product) by the activity of fumarase (Tajima et al. 2015).
The aspartic acid production process by fermentation requires pH control by the addition
of calcium hydroxide to produce a precipitate of calcium fumarate. Subsequently, the
ammonium fumarate is produced by separating the calcium fumarate precipitate from the
fermentation broth and reacting it with a reagent selected from ammonia, ammonium car-
bonate, ammonia in combination with CO2 and mixtures thereof, to form ammonium
fumarate and a co‐product which is calcium carbonate and calcium hydroxide, depending
on the amination reagent used. The energy of the indirect neutralization of fumaric acid by
ammonia is a driving force for calcium fumarate conversion to ammonium fumarate prod-
uct and for regeneration of a calcium‐based reagent. Diammonium fumarate is converted
enzymatically into an ammonium aspartate and acidified to form aspartic acid.
In 1974, at the Lomonosov Moscow State University, researchers began to use immobi-
lized microorganism cells to obtain natural amino acids. Several bacterial strains with
aspartase activity were selected; subsequently, the activation and immobilization condi-
tions of E. coli cells were studied. A technique was developed by Chibata et al. (1986) using
κ‐carrageenan as the immobilizing matrix for E. coli (ATCC 11303) cells. The aspartase
activity was about seven times higher than that of the parent cells. Mutant bacterial strains
Aspartase
Aspartic acid
NH3
Fumaric acid
Fumarase
Malic acid
H2O
Figure 10.3 Fumaric acid metabolism.

optimized for efficient aspartic acid biosynthesis and low fermentation levels of by‐products
have been patented, such as E. coli TA5003, E. coli TA5004, and E. coli TA5005. As the car-
bon source, carbohydrates such as glucose, fructose, sucrose, molasses containing these,
starch and starch hydrolyzates, organic acids such as acetic acid, fumaric acid, lactic acid,
and propionic acid, and alcohols such as ethanol, glycerin, propanol, etc. can be used
(Takano and Kino 1999). In 2000 at the Autonomous University of Nuevo Leon, the isola-
tion of two bacterial strains was achieved – Bacillus cereus and Enterobacter cloacae – with
high activity to be used as biocatalysts in the biotransformation process of fumaric acid in
the amino acid L‐aspartic acid, in batch‐type reactors (Garza 2000).
In 2009, researchers from the microbiology laboratory of the Universidad Pontificia
Bolivariana produced amino acid L‐aspartic acid at 1.5 mmol/L using the native strain of
Pseudomonas fluorescens and fumaric acid as a source of carbon and energy. L‐aspartic acid
is formed in a submerged fermentation process. Under the laboratory conditions studied,
in a culture medium with 1% fumaric acid, the growth of P. fluorescens native strain of the
UPB is consistent with the Monod model, with a maximum specific growth rate (μmax) of
0.0097/h and half‐velocity constant (Ks) of 0.013 g/L (Oviedo et al. 2009). Free and immo-
bilized thermostable aspartase of Bacillus sp. YM55–1 expressed in E. coli could yield over
430 mmol/L aspartic acid after 24‐hour fermentation (Cárdenas‐Fernández et al. 2012).
Derbikov et al. (2017) report the construction of E. coli strain MG1655 derivatives with
deleted genes that encode fumarases (fumAC, fumB, and fumABC). When the E. coli
strains with deleted fumarase genes were used to catalyze L‐aspartic acid synthesis from
ammonium fumarate (1.5 mol/L), it was observed that only the simultaneous loss of both
the fumA and fumC genes led to an at least 20% increase in aspartic acid yields and a con-
current decrease in the content of the by‐product malic acid in the reaction mixture from
40 to 1.5–2 g/L. The results obtained in this work may be used to generate more efficient
novel biocatalysts of L‐aspartic acid synthesis.
The main uses of aspartic acid include the following.
●● Interest in the production of the amino acid L‐aspartic acid, from the point of view of its
application in pharmacology, increased significantly after 1958 related to its physiologic
and therapeutic action. Aspartic acid participates in transamination reactions that are of
vital importance for the cell, since it is transformed into oxalacetic acid, an important
intermediate in the tricarboxylic acid cycle, by transamination with α‐ketoglutaric acid.
Oxalacetic acid is one of the final stages of oxidative catabolism. In addition, it is an
ancestor of the pyrimidic nucleotides; in the organism, it serves as a basis for orotic acid
synthesis, an ancestor of uridine. In combination with glucose and sorbitol, it is used for
the cure of liver diseases. Its derivatives are used in the treatment of alcohol intoxication.
Aspartate potassium and magnesium are used to treat fatigue and heart failure and iron
aspartate is used to treat anemias.
●● Different derivatives of L‐aspartic acid are used in the food industry; for example, sodium
aspartate is a component of spices to improve the flavor of some food products. Aspartame
(L‐aspartyl‐L‐phenylalanine methyl ester) is a dipeptide formed by L‐aspartic acid and
the methyl ester of phenylalanine, which is marketed as a low‐calorie sweetener.
●● Aspartic acid is useful as an initial or intermediate material for the formation of sur-
factants, metal ion sequestrants, detergents, cosmetics, peptide synthesis, pharmaceutical
products and coatings and for the formation of the biodegradable polymer of polyaspar-
tic acid (PAA). The latter can be used as a co‐builder or as a sequestrant in detergents and
as a superabsorbent polymer in addition to other applications
●● In agriculture, aspartic acid in combination with lysine is used as a fungicide stabilizer.
10.3.5 Malic Acid

Malic acid (C4H6O5) is an organic dicarboxylic acid with a similar structure to maleic acid.
Its salt and esters are known as malates. There are three microbial groups able to produce
malic acid. The first uses a one‐step fermentation process from glucose; the second trans-
forms fumaric acid into malic acid by the enzyme furamase and the third one by synthesis
of poly‐b‐malic acid which can be hydrolyzed to L‐malic acid. Several wild and metaboli-
cally engineered strains are able to produce L‐malic acid, including A. niger, A. oryzae, A.
flavus, Pennicillium sp. K034, Aureobasidium pullulans ZX‐10, S. cerevisiae, E. coli XZ658,
and E. coli WGS‐10. Since wild‐type S. cerevisiae strains produce only low levels of malate,
metabolic engineering is required to achieve efficient malate production with this yeast.
Three pathways are recognized to produce L‐malic acid from glucose: a nonoxidative
pathway, an oxidative pathway, and from the glyoxylate cycle. The nonoxidative pathway
involves pyruvate carboxylation to oxaloacetate followed by its reduction to malate, while
S. cerevisiae lacks phosphoenolpyruvate carboxylase. The oxidative pathway involves the
condensation of acetyl‐coenzyme A (acetyl‐CoA) and oxaloacetate to citric acid, followed
by oxidation to malate in the tricarboxylic acid (TCA) cycle. The third pathway involves
the production of malates from two molecules of acetyl‐CoA via the glyoxylate cycle (Zou
et al. 2013).
Via carboxylation of pyruvate, followed by reduction of oxaloacetate to malate, an engi-
neered glucose‐tolerant S. cerevisiae strain was used. In glucose‐grown batch cultures, the
resulting engineered strain produced malate at titers of up to 59 g/L at a malate yield of
0.42 mol/mol glucose. The engineered strains still produced substantial amounts of pyru-
vate, indicating that the pathway efficiency can be further improved (Zelle et al. 2008).
A malic acid titer of 143 g/L with a productivity of 0.74 g/L/h was achieved by combining
a fermentation process carried out by A. pullulans with acid hydrolysis of the produced
polymalic acid (Zou et al. 2013). Also, a high malate production rate (0.94 g/L/h) along with
high titer (154 g/L) were achieved using an engineered A. oryzae strain after overexpressing
the enzymes involved in the reductive TCA pathway in conjunction with the malate trans-
porter (Brown et al. 2013).
A sequential fermentation system using syngas, as primary carbon source, for acetic acid
production, followed by acetic acid for the production of malic acid was demonstrated by
Oswald et al. (2016). In this technique, an anaerobic Clostridium ljungdahlii bacterial strain
was used in the first reactor, while an aerobic Aspergillus oryzae fungal strain was used in
the second reactor. According to the authors, the final amounts of malic acid represented
the highest yields of any C4 compound derived from syngas fermentation process. In addi-
tion, this bioprocess technique showed the potential of anaerobic syngas fermentation for
the synthesis of a wider range of bioproducts as well as providing a new feedstock for the
biosynthesis of malic acid other than glycerol and sugars (Oswald et al. 2016). Malic acid is
used as a flavor enhancer and an acidulant in carbonated beverages, foods, and candies. It
is also used as one component of an antimicrobial agent against Salmonella enteric ser.
Saintpaul and E. coli O157:H7 in sterile and fresh orange and mango juices.
10.3.6 Itaconic Acid

Itaconic acid (2‐methylenebutanedioic acid) is a dicarboxylic acid with five carbon atoms,
also known as methylene succinic acid. Its chemical formula is C5H6O4. Its density is
1.63 g/L. The use of itaconic acid (IA), despite its enormous potential as a basic chemical
compound, has been limited by its high production costs from petroleum. Its production
through fermentation, which is the industrially established process, has a lower cost, but
the volume of production is still not high enough to allow its use to expand. Since the
1940s, Aspergillus itaconicus and A. terreus have been well known as excellent producers for
the biotechnological production of IA in a batch submerged fermentation process. Several
microorganisms have been used to produce IA at laboratory scale which mainly include
Candida spp., Ustilago zeae, Rhodotorula sp., Candida mutant, A. terreus TN‐484‐M1, A.
terreus SKR10‐20, etc.
The biosynthesis of IA occurs by the Krebs cycle, in a process similar to that used for
citric acid from glucose, with two additional steps. Although both processes are similar, as
expected for a by‐product of the citric acid cycle (TCA), they present a significant differ-
ence: the tolerance of the producer microorganism toward the product is lower in the case
of IA. For this reason, neutralization with calcium hydroxide at the low pH required (2) is
necessary in order to obtain higher concentrations of IA. Also, IA can be oxidized to itatar-
taric acid by IA oxidase. This enzyme has to be inhibited by adding calcium hydroxide for
a maximum yield of IA. The yield is around 75% of the theoretical calculation when the
medium contains 15% sucrose (FEDIT 2008).
Sugar is metabolized through glycolysis to pyruvate, a compound that can follow two
paths. One part is carboxylated to oxaloacetate by the action of pyruvate carboxylase;
another part is converted to acetyl‐CoA, with release of a CO2 molecule. Both compounds,
that is, oxaloacetate and acetyl‐CoA, enter the TCA and, through a condensation reaction
catalyzed by citrate synthase, give rise to citrate. Finally, citrate is dehydrated by aconitase
to cis‐aconite and catalyzed decarboxylation to IA by the enzyme cis‐aconitic decarboxy-
lase. The overall balance is that CO2 is reused in the process since the one generated in one
reaction is used in the other (FEDIT 2008).
A simple way to represent the reaction is:
Glucose (C6H12O6) + 3 NAD + Itaconic acid (C5H6O4) + 3 NADH + 3 H+ + CO2
Very little research has been directed toward improving the production of IA. In fact, one
of the most efficient processes was patented in the last century, by fermenting the sugar
present in molasses which presented a yield of 70%. The maximum published concentra-
tions of the acid in the fermentation medium are slightly higher than 80 g/L. The sub-
merged fermentation is carried out at 40 °C. The fermentation is highly aerobic. Aspergillus
terreus has been chosen as the preferred itaconate‐producing strain (Okabe et al. 2009).
Kuenz et al. (2012) obtained IA with a self‐ isolated strain of A. terreus DSM 23081 at a
15 L scale. A concentration of 86.2 g/L was achieved within seven days with an overall pro-
ductivity of 0.51 g/(L/h), a maximum productivity of 1.2 g/(L/h), and a yield of 86% (mol).
Cultures with A. terreus strains with the same process showed no significant differences.
An engineered A. niger has been explored as an itaconate‐producing strain by localizing
and co‐overexpressing the key enzymes of the production pathway (cis‐aconitate decar-
boxylase and aconitase) in the mitochondria. Productivity doubled the value for native
A. niger (Blumhoff et al. 2013).
This acid is used in the manufacture of acrylic plastics, acrylate latex, detergents, coat-
ings, and rubber, and perhaps its greatest expansion is to produce superabsorbent polymers
such as poly(acrylamide‐co‐itaconic acid), with the great characteristic of absorbing large
amounts of water that could be used in feminine care products. The company Itaconix has
been successfully producing poly(itaconic acid) commercially. Due to its chemical struc-
ture, IA can be considered as an α‐substituted acrylic or methacrylic acid, so it can compete
with methyl methacrylate and other acrylates, as well as in the field of pressure‐sensitive
adhesives. In addition, its conversion into methyl methacrylate, also known as Plexiglass®,
represents an attractive alternative for the industry (Jang et al. 2012; Choi et al. 2015).
The basic chemistry of IA is similar to that of the petrochemical compound maleic acid
(and its anhydride), so that compounds such as 2‐methyl‐1,4‐butanediol,3‐methyl
tetrahydrofuran, 3‐ and 4‐γ‐butyrolactone, and 2‐methyl‐1,4‐butanediamine can be derived
from it by hydrogenation/reduction reactions. It can also be converted into pyrrolidone‐
type derivatives. Most IA has been produced by Chinese companies including Qingdao
Kehai Biochemistry, Zhejiang Guoguang Biochemistry, and Jinan Huaming Biochemistry.
10.3.7 Glucaric Acid

Glucaric acid (C6H10O8), also known as saccharic acid, is the product of sugar or polysac-
charide oxidation with nitric acid. German chemist Heinrich Kiliani first described the
reaction in 1925. Glucaric acid is a member of a much larger family of materials known as
oxidized sugars. These materials represent a significant market opportunity. For example,
oxidation of glucose to glucaric acid can be carried out using chemical or biochemical
catalysis. In contrast, production of glucaric acid as a building block is more difficult.
However, glucaric acid can be used as a starting point for a wide range of products with
applicability in high‐volume markets.
Nitric oxidation of D‐glucose in D‐glucaric acid is the main technology used currently.
Although low yields of D‐glucaric acid are obtained due to other collateral reactions that
generate several oxidation products, the method continues to be attractive for its relative
simplicity, with nitric acid acting simultaneously as a solvent and oxidizing agent. An effi-
cient and selective glucose oxidation technology should be developed, eliminating nitric
acid as the oxidant. Further technical barriers include development of selective methods
for sugar dehydration to transform glucaric acid into sugar lactones, particularly glucaric
dilactone. The biological production of D‐glucaric acid offers the potential of a cheaper and
environmentally friendly process. Some synthetic biological routes have already been
proposed.
Moon et al. (2009) reported production of D‐glucaric acid from glucose in E. coli using
enzymes from three different organisms: myo‐inositol‐1‐phosphate synthase (Ino1) from S.
cerevisiae, myo‐inositol oxygenase (MIOX) from mice, and uronate dehydrogenase (Udh)
from Pseudomonas syringae. 1.13 g/L glucaric acid and 13% (mol) yield were achieved in
strain BL21 (DE3) in LB medium supplemented with 10 g/L glucose. In this process,
co‐expression of the genes encoding Ino1 from S. cerevisiae and MIOX from mice led to
production of glucuronic acid through the intermediate myo‐inositol. Glucuronic acid con-
centrations up to 0.3 g/L were measured. The activity of MIOX was rate limiting, resulting
in the accumulation of both myo‐inositol and glucuronic acid. Inclusion of Udh from
P. syringae facilitated the conversion of glucuronic to glucaric acid. Thus, improving the
stability and activity of MIOX is crucial for optimizing glucaric acid production.
Saccharomyces cerevisiae was also investigated for glucaric acid production because of its
satisfactory acid tolerance. The same biosynthetic pathway was used with codon‐optimized
MIOX. Applying a fed‐batch fermentation strategy, production was increased to 1.6 g/L
from glucose supplemented with myo‐inositol (Kang and Gong 2016). MIOX activity and
myo‐inositol availability were rate limiting in glucaric acid production, not only in E. coli
but also in S. cerevisiae. Therefore, changes in MIOX are required in order to increase glu-
caric acid production. By expressing a more stable MIOX4 from A. thaliana and integrating
the target genes into the delta sequence of the genomes, the glucaric acid titer in S. cerevi-
siae was increased. Delta sequence‐based constitutive expression increased both the num-
ber of target gene copies and their stability and can be used for a wide range of metabolic
pathway engineering projects in S. cerevisiae. The final strain produced 6.0 g/L glucaric
acid, which is the highest titer reported in S. cerevisiae (Chen et al. 2018).
In a fed‐batch fermentation process with Komagataella phaffii (previously Pichia pasto-
ris) and after optimization of the expression of MIOX and Udh with a fusion expression
strategy, the titer of glucaric acid was significantly increased to 6.61 g/L from glucose and
myo‐inositol. The inefficient biosynthesis of myo‐inositol affects the production of glucaric
acid from glucose (Lui et al. 2016). More efforts are needed to construct a novel synthetic
pathway toward glucaric acid. Glucaric acid titers and yields could be improved by control-
ling the phosphofructokinase (Pfk) activity in the production of glucaric acid from glucose
in a semi‐defined medium under batch and fed‐batch conditions. Modified E. coli strain
IB1486‐GA was used and was compared with wild‐type MG1655. Timed knockdown of Pfk
activity produces a maximum improvement of up to 42% (Brockman et al. 2015). Kalion, a
biotechnology company, is using this type of low‐cost fermentation technology to obtain
D‐glucose in recombinant E. coli D‐glucaric acid.
The main uses of glucaric acid are as follows (Figure 10.4).
Concrete Adhesives/coating Corrosion inhibitor Adipic acid

admixture paints
1,6-hexanediol
D-saccharic acid 1,4-lactone Glucaric acid
Hexamethylenediamine
Potential cancer Biodegradable Detergent

prevention agents chelating builder
agents Fibers, polymers
Figure 10.4 Overview of glucaric acid uses in different sectors.

●● A wide range of other biodegradable polymers are also possible including methacrylate
hydroxylase, nylons, and other ester/amide polymers. Glucaric acid acetate (GAA),
which is acyclic, was synthesized in an acetic anhydride/sulfuric acid mixture. GAA
was converted to glucaric acid chloride acetate (GACA) and then polymerized with
various diols and diamines in dimethylacetamide solution or by interfacial polymeriza-
tion in water and chloroform solutions. The polyesters and polyamides were amphi-
philic and soluble in water and common organic solvents. Differential scanning
calorimetry showed that the polyamides were thermoplastic and melted at c.140 °C,
indicating crystallinity; the melting points increased with increasing number of
diamine alkyl carbons. Novel biobased crystalline amphiphilic polymers were synthe-
sized from glucaric acid.
●● Glucaric acid itself has a diverse market potential with uses ranging from detergents (as
it exhibits useful chelating properties for cations) to polymers, to food additives (due to
its health benefits).
●● Dicarboxylic acid functionalization also makes it an attractive bio‐derived precursor to
adipic acid, which could be used as a more sustainable alternative to fossil fuel‐derived
adipic acid in nylon production plants.
●● Corrosion inhibition properties have been recognized for glucaric acid.
●● Glucaric acid is a potential cancer prevention agent.
10.3.8 3-Hydroxypropionic Acid

3‐Hydroxypropionic acid (3HP) is a potential chemical building block for sustainable pro-
duction. It can be transformed into acrylic acid,1,3‐propandiol, malonic acid, super‐absor-
bent polymers, and acrylic plastics. It is a 3‐carbon, nonchiral organic molecule and a
structural isomer of lactic acid. The two functional groups, a carboxyl group and a β‐
hydroxyl group, give high reactivity for 3HP. Five pathways have been reported to convert
sugars (mainly glucose) to 3HP: malonyl‐CoA pathway, β‐alanine pathway, propionyl‐CoA
pathway, glycerate pathway, and lactate pathway (Matsakas et al. 2018).
The malonyl‐CoA pathway is one of the most investigated for 3HP production. Glucose
is transformed through glycolysis to acetyl‐CoA, which is then converted to malonyl‐CoA
by acetyl‐CoA carboxylase. Malonyl‐CoA reductase converts malonyl‐CoA to malonyl
semialdehyde, which is further converted to 3HP by malonyl‐CoA reductase. This route is
considered the most promising (Vidra and Németh 2018).
The β‐alanine pathway begins with fumarate, as part of the TCA cycle. This fumarate is
converted to aspartate by aspartase, then to β‐alanine by aspartate α‐carboxylase. β‐alanine
pyruvate aminotransferase converts β‐alanine to malonic semialdehyde and finally malonic
semialdehyde reductase converts it to 3HP. Theoretically, the yields achieved through the
β‐alanine intermediate pathway should be higher than those obtained through the malonic
intermediate pathway, as the latter is highly oxygendependent and requires high levels of
ATP for acetyl‐CoA synthesis (Song et al. 2016). The route by which β‐alanine is converted
into malonic semialdehyde by the action of either β‐alanine‐pyruvate aminotransferase
(BAPAT) or γ‐amino butyrate transaminase (GABT) has been reported in E. coli (Liao et al.
2007) and yeast (Jensen et al. 2014) respectively.
Succinic acid can be produced from glucose by the propionyl‐CoA pathway. It can be
transformed into propionic acid by the action of succinate decarboxylase, followed by the
conversion of propionate to propionyl‐CoA by propionyl‐CoA synthetase. Propionyl‐CoA
is changed to acryloyl‐CoA by the enzyme propionyl‐CoA dehydrogenase, which is con-
verted to 3‐hydroxypropionyl‐CoA by 3‐hydroxypropionyl‐CoA dehydratase and finally to
3HP by the enzyme propionate‐CoA transferase or the enzyme 3‐hydroxyisobutyryl‐CoA
hydrolase (Luo et al. 2016). Klebsiella pneumoniae, Lactobacillus reuteri, and L. collinoides
produce 3HP by oxidizing 3‐hydroxypropionaldehyde (3‐HPA).
The glycerate pathway has not yet been constructed in a host microorganism (Matsakas
et al. 2018). The lactate pathway is not recommended because conversion of lactate into
3HP is not so favorable (Borodina et al. 2015). Another option to convert glucose (or other
sugars) to 3HP is by initially producing glycerol from the central metabolism and then con-
verting the glycerol to 3HP with strategies that have been established for glycerol, instead
of directly converting the glucose (Matsakas et al. 201). Engineered E. coli and native and
engineered Klebsiella strains are frequently used to transform glycerol into 3HP via glycerol
dehydratase and aldehyde dehydrogenase. There are two main routes to the production of
3HP from glycerol that have been investigated during recent years: CoA‐dependent path-
way and CoA‐independent pathway (Kwak et al. 2013).
Kildegaard et al. (2015) engineered S. cerevisiae producing 3HP from glucose and
xylose. Researchers introduced the 3 HP biosynthetic pathways via malonyl‐CoA or β‐
alanine intermediates into xylose‐consuming yeast. Using controlled fed‐batch cultiva-
tion, 7.37 g 3HP/L in 120 hours with an overall yield of 71% mol/mol of xylose was
achieved. Borodina et al. (2015) optimized a synthetic pathway for de novo biosynthesis
of β‐alanine and its conversion into 3HP using a novel β‐alanine‐pyruvate aminotrans-
ferase discovered in B. cereus. 13.7 g/L of 3HP and 0.14 mol/mol (on glucose) yield was
achieved using a batch fermentation at pH 5 and 30 °C. Main results for 3HP production
at laboratory scale (less than 5 L), using glucose and xylose as substrates, are shown in
Table 10.3.
With glucose as a substrate, only a few studies have been reported, while there is a patent
on high‐level 3HP production from glucose, fructose, sucrose, lactose, and dextran as car-
bon sources and different recombinant strains, modified to increase enzymatic activity in
the malonyl‐CoA reductase (mcr) pathway (Lynch et al. 2011). Unfortunately, it is still far
from commercial application. 3HP production should be above 100 g/L with a yield higher
than 50% and productivity over 2 g/L/h, to be economically feasible (Vidra and Németh
2018). For these, more research is required. 3HP acid can be used as a precursor for many
compounds such as acrylamide, 1,3‐propanediol acrylic acid, and methyl acrylate. Also, it
can be used to synthesize chemical intermediates such as malonic acid, propiolactone, and
alcohol esters of 3HP. Novozymes and Cargill are working to develop microorganisms that
can convert renewable feedstock into 3HP in an economically feasible process.
10.3.9 2,5-Furan-Dicarboxylic Acid

2,5‐Furan‐dicarboxylic acid (FDCA) is aa oxidized hydroxymethyl furan derivative also
known as dehydromucic acid; it has the chemical structure C6H4O5. It is a white solid
Table 10.3 3-Hydroxypropionic acid fermentation results from glucose and xylose at laboratory

scale.
Titer Yield Productivity

Substrate Strain g/L g/g g/L/h Reference
Glucose Corynebacterium glutamicum 62.6 0.51 0.87 Chen et al. (2017)

MH15
Glucose Saccharomyces cerevisiae 13.7 0.55 0.17 Borodina et al.
SCE‐R2‐200 BcBAPAT EcYDFG (2015)
TcP and ScAAT2 ScPYC1/PYC2
Glucose Escherichia coli BL21_mcr_acc_ 0.2 0.03 0.01 Rathnasingh
pntAB et al. (2012)
Glucose S. cerevisiae SCIYC33/ pJC5 / 0.46 0.02 0.006 Chen et al. (2014)
pJC9
Glucose S. cerevisiae ST687 ALD6, 9.8 0.33 0.098 Kildegaard et al.
SEacsL641P, (2015)
PDC1, and TY4‐CaMCR‐ACC1
Glucose E. coli, upregulation of sdhC, 31.1 0.42 0.63 Song et al. (2016)
overexpression of ppc
Glucose and E. coli BL21_dhaB_ dhaR _ aldH 14.3 – 0.26 Kwak et al.
glycerol (2013)
Glucose and E. coli W3110.ackA‐pta.yqhD_ 71.9 – 1.8 Chu et al. (2015)
glycerol dhaB_gdrAB and_ mutant gabD4
Glucose and E. coli JHS01300/pELDRR + 29.4 0.36 0.54 Jung et al. (2015)
xylose pCPaGGRm
Glucose and C. glutamicum MH15 54.8 0.49 0.76 Chen et al. (2017)
xylose
Xylose S. cerevisiae ST2547 7.37 0.37 0.06 Kildegaard et al.
(2015)
i nsoluble in water and completely dissolves only in DMSO. It was first detected in human
urine; in fact, a healthy human produces 3–5 mg/day. It is a very stable compound with a
melting point of 342 °C and a boiling point of 420 °C. It has a density of 1.604 g/cm3. This
substance has two pKa values: 2 and 28.
2,5‐Furan‐dicarboxylic acid was first prepared from mucic acid by Fittig and Heinzelmann
in 1876 by reacting with fuming hydrobromic acid under pressure. FDCA is a monomer
that is not commonly found in Nature. It is synthetically produced by the dehydration of
hexoses, mainly fructose, to 5‐hydroxymethyl furfuraldehyde and its subsequent oxidation
through chemical processes (Shuguo et al. 2015). 5‐HMF is not stable and degrades upon
storage. It may undergo rehydration in aqueous phase, thereby originating by‐products
such as levulinic and formic acid, or even condense into polymers called humins. So, the
use of stable intermediates (for instance, alkoxy derivatives) or the direct conversion of
fructose into FDCA are preferred. Since FDCA has no polymerizing action, it cannot be
used as such as a polymer unless it is combined with ethane‐1,2‐diol (monoethylene glycol)
through esterification. This esterified or combined polymer, known as polyethylene
furanoate (PEF), can be used as an analog of petroleum‐based polymers, and polyethylene

terephthalate (PET) and polybutylene terephthalate (PBT) can reduce this nonrenewable
energy use by 45–55% (Gandini et al. 2009).
Polymers like polyesters, polyamides, and polyurethanes from FDCA can be used for bot-
tles, containers, nylons, coating resins, engineering plastics, and films. Apart from this, it
can be used as an ingredient of fire foams, a drug to treat kidney stone and artificial veins
for transplantation. The current FDCA manufacturers Avantium, Avalon Industries,
Eastman, Novamont, Carbone Scientific, Tokyo Chemical Industry, and Petrobras produce
it through chemical processes using either fructose or 5‐HMF as substrates. Avantium col-
laborated with BASF to develop the YXY process to produce biobased FDCA from plant‐
based sugars and other chemicals, with two catalytic steps. Dehydroxylation of the aldaric
acid into furan carboxylic acid (FCA) and FDCA or muconic acid depending on the reac-
tion conditions is a process developed by the VTT Technical Research Centre of Finland
(Bio Refineries Blog 2018).
To study the biological transformation of HMF into FDCA, a pathway route has been
found. HMF or 5‐hydroxymethyl furfuraldehyde (C6H4O3) is an aldehyde toxic to microor-
ganisms. The required electron transfer system is catalyzed by either peroxidases or inter-
nal and external dehydrogenases present in the microorganisms. This pathway was first
reported in the microorganism Cupravidus basilensis. Selective oxidation of HMF is con-
verted into furanic chemicals such as HMF alcohol, maleic anhydride (MA),
5‐hydroxymethyl‐2‐furancarboxylic acid (HFCA) (HMF acid), 2,5‐diformylfuran (DFF), 5‐
formyl furoic acid (FFA), and FDCA.
A few approaches have been evolved for the bioconversion of substrates like biomass, C6
sugars glucose or fructose and 5‐HMF to FDCA using nontoxic biocatalysts. Lack of suita-
ble enzymes for these substrates was considered as the major technical barrier for an effi-
cient technology (Werpy et al. 2004).
Production of FDCA with microorganisms required a substrate HMF in pure form. At
least three subsequent oxidations of HMF, in which two oxidations are on an aldehyde
medium, eventually lead to FDCA. Generally, aldehydes cause extensive damage to the
organism’s proteins, nucleic acids, and other cell organelles through the formation of reac-
tive oxygen species. HMF is a type of furan aldehyde which inhibits the activity of primary
metabolic enzymes of microorganisms and they slowly reach the death phase without fur-
ther multiplication. To overcome this, certain microorganisms have naturally evolved alde-
hyde dehydrogenases or oxidases which convert these furan aldehydes into their alcohol or
acid product which are normally less toxic. The first report on biological FDCA production
using a microorganism was with the organism Cupriavidus basilensis HMF14 isolated from
5‐HMF and furfural containing lignocellulosic hydrolyzate. The authors characterized the
HMF and furfural degradation pathways of this organism both at the biochemical and the
genetic level. The formation of FDCA from HMF required an FAD‐dependent oxidoreduc-
tase, encoded by an hmfH gene from the hmfFGH’H gene cluster. This HmfH oxidase is an
alternative to the nonspecific dehydrogenases (Koopman et al. 2010a).
Pseudomonas putida S12 strain is able to use HMF and furfural as carbon sources because
of nonspecific “furanic dehydrogenases.” HMF oxidase gene was isolated from C. basilensis
HMF14 and engineered into P. putida S12. Diacids were formed during the production of
FDCA when they used glucose as carbon source and many buffers should be used to adjust
the acid pH. When glycerol is used as carbon source, 276 ± 89 μmol/g cell dry weight
(CDW)/h of FDCA was obtained (Koopman et al. 2010a,b).
As a cheaper biomass, the marine macroalgae Gracilaria verrucosa and Gelidium aman-
sii were used as substrates for the production of HMF after thermal acid treatment and
successfully transformed this algal hydrolyzate containing HMF into FDCA. Burkholderia
cepacia H‐2 was inoculated in zero‐fold and twofold diluted acid hydrolyzate form and
989.5 and 1031 mg/L FDCA production was reported after 24 hours at pH of 7 and tempera-
ture 28 °C. This is the first report of the production of FDCA from a biomass containing
HMF (Yang and Huang 2016).
For a fed‐batch fermentation process, preculture of genetically engineered P. putida S12
(0.2 g/L CDW) is inoculated into a 1 l fermenter with modified media containing 4 mol/L
glycerol, 0.1 mol/L MgCl2, and 1 mol/L HMF. HMF was increased and in order to avoid
that, the rate of HMF feed was reduced to 0.09 mmol/g CDW/h after 72.8 hours and stopped
after 117.4 hours. To increase the rate of oxidation and product formation from HMF, the
feed rate of glycerol was increased to 12 mmol/L/h. Oxygen was supplied at 11 mL/min for
the first 25 hours after which it was changed ito 200 mL/min. pH was maintained at 7 by
adding NH4OH and NaOH. FDCA was obtained with 97% conversion and concentration of
30.1 ± 0.7 g/L (Koopman et al. 2010a,b).
Wierckx and group engineered HmfH and HmfT1 into P. putida S12 B‐38 and HmfH, and
HmfT1 and aldehyde dehydrogenase into P. putida S12 B‐51 for co‐expression of these
genes for the production of FDCA from HMF ina fed‐batch fermentation process, in a
medium supplemented with glycerol, producing 150 g/L FDCA with a CDW of 28 g/L after
90 hours (Wierckx et al. 2012, 2015).
Enzymatic oxidation of 5‐HMF and its oxidized derivatives was studied using three fun-
gal enzymes: wild‐type aryl alcohol oxidase (AAO) from three fungal species, wild‐type
peroxygenase from Agrocybe aegerita (AaeUPO), and recombinant galactose oxidase
(GAO). Experiments were done at laboratory scale and need optimization and further
research (Karich et al. 2018). An enzymatic cascade involving three fungal oxidoreductases
has been developed for the production of FDCA from 5‐methoxymethylfurfural (MMF).
Previously, it was obtained from HMF, but yields are higher with MMF. Aryl alcohol oxi-
dase and unspecific peroxygenase act on MMF and its partially oxidized derivatives, yield-
ing FDCA, as well as methanol as a by‐product. Methanol oxidase takes advantage of the
methanol released for in situ production of H2O2 that, along with that produced by aryl
alcohol oxidase, fuels the peroxygenase reactions. In this way, the enzymatic cascade pro-
ceeds independently, with the only input of atmospheric O2, to attain a 70% conversion of
initial MMF. The addition of exogenous methanol to the reaction further improves the
yield to attain an almost complete conversion of MMF into FDCA (Carro et al. 2018).
Several microorganisms used to produce any of the furfural or HMF derivatives after fer-
mentation include Amorphotheca resinae ZN1, C. basilensis HMF14, Arthrobacter nicotiana,
Telluriamixta, Burkholderiales, Rhizobiales, and Coniochaetales (Rajesh et al. 2018). Even
though microbial and enzyme‐assisted production of FDCA has been reported, there is no
report of the biological production of FDCA from industry. However, the newest plant planned
by Synvina (joint venture between Avantium and BASF) should produce by 2023–2024 50
ktons FDCA from sugars, using dehydration and oxidation (Bio Refineries Blog 2018). Corbion
is developing 100% biobased FDCA for the production of high‐performance PEF resin.
The main uses of FDCA are as follow.

●● Polyesters, polyamides and polyurethanes: the most important group of FDCA conver-
sions is undoubtedly polymerization. The FDCA monomer offers great opportunities to
create a wide range of polymers: polyesters (bottles, containers, and films), polyamides
(for new nylons), and polyurethanes.
●● Plasticizers: FDCA esters have recently been evaluated as replacements for phthalate
plasticizers for PVC.
●● Fire foams: FDCA, as for most polycarboxylic acids, is an ingredient of fire foams. Such
foams help to extinguish fires caused by polar and nonpolar solvents in a short time.
●● Precursor of levulinic and succinic acids.
●● Pharmacology: it has been demonstrated that FDCA diethyl ester has strong anesthetic
action similar to cocaine. Screening studies on some FDCA derivatives showed impor-
tant antibacterial properties. A diluted solution of FDCA in tetrahydrofuran is utilized
for preparing artificial veins for transplantation.
10.3.10 L-Glutamic Acid

Glutamic acid (GA) is a 5‐carbon α‐amino acid (C5H9O4N) that is used by almost all living
beings in the biosynthesis of proteins. It has the potential to be a novel building block for
5‐ carbon polymers. Its molecular structure could be idealized as HOOC‐CH(NH2)‐(CH2)2‐
COOH, with two carboxyl groups ‐COOH and one amino group ‐NH2. However, in the
solid state and mildly acid water solutions, the molecule assumes an electrically neutral
zwitterion structure ‐OOC‐CH(NH3+)‐(CH2)2‐COOH. It is encoded by the codons GAA or
GAG. The building block and its derivatives have the potential to build similar polymers
but with new functionality to derivatives of the petrochemicals derived from maleic anhy-
dride. These polymers could include polyesters and polyamides (glutaminol, 5‐amino‐1‐
butanol, 1,5‐pentanediol, norvoline). The acid can lose one proton from its second carboxyl
group to form the conjugate base, the singly negative anion glutamate ‐OOC‐CH(NH3+)‐
(CH2)2‐COO‐. This form of the compound is prevalent in neutral solutions. In highly alka-
line solutions, the doubly negative anion ‐OOC‐CH(NH2)‐(CH2)2‐COO‐ prevails. The
radical corresponding to glutamate is called glutamyl.
When glutamic acid is dissolved in water, the amino group (‐NH2) may gain a proton
(H+) and/or the carboxyl groups may lose protons, depending on the acidity of the
medium. In sufficiently acidic environments, the amino group gains a proton and the
molecule becomes a cation with a single positive charge, HOOC‐CH(NH3+)‐(CH2)2‐
COOH. The substance was identified in 1866 by the German chemist Karl Heinrich
Ritthausen who treated wheat gluten (for which it was named) with sulfuric acid. In 1908,
the Japanese researcher Kikunae Ikeda of the Tokyo Imperial University identified glu-
tamic acid and patented a method of mass‐producing a crystalline salt of glutamic acid,
monosodium glutamate (MSG). The breakthrough in MSG production was the isolation
of a specific soil‐inhabiting gram‐positive bacterium, Corynebacterium glutamicum, by
Dr S. Ukada and Dr S. Kinoshita in 1957. Aerobic fermentation of sugars and ammonia
with C. glutamicum (also known as Brevibacterium flavum) substituted the chemical
synthesis (Shyamkumar et al. 2014).
Glucose is one of the major carbon sources for production of GA. GA has been produced
from various kinds of raw materials, including submerged fermentation of palm waste
hydrolyzate, cassava starch, SCB, and date waste. GA is commercially one of the most
important amino acids produced mainly by fermentation. GA biosynthesis is an aerobic
process that requires oxygen throughout the fermentation. It was shown that the absence
of ammonium ions, but with sufficient oxygen supply, resulted in the accumulation of α‐
ketoglutaric acid instead of GA.
As for other organic acids, a very low‐cost fermentation route is necessary. There are cur-
rently several fermentation routes for the production of MSG which are all based on the
production of the sodium salt. One of the major challenges for the expansion of low‐cost
fermentation is to develop an organism that can produce GA as the free acid. This would
eliminate the need for neutralization and substantially reduce the costs of purification and
conversion of the sodium salt to the free acid. Additional improvements in the fermenta-
tion would include increasing the productivity of the organism and improving final fer-
mentation titer. A minimum productivity of 2.5 g/L/h needs to be achieved in order to be
economically competitive on a commodity scale.
Glutamic acid can be synthesized from different precursors and reactions. Yelamanchi
et al. (2016) reported the sequences of chemical reactions, enzymes involved, and thermo-
dynamic parameters for the production of L‐glutamate. The metabolism of pyruvic acid via
acetyl‐CoA leads to the formation of citrate by condensation with oxaloacetate in the Krebs
cycle. The most important step in this cycle for the present context is that the α‐ketoglutar-
ate (α‐KG) acid, one of the intermediates, can exit the cycle to form glutamate. The reaction
by which GA can be generated is transamination. The enzymes of the glutamate metabo-
lism pathway are tightly controlled by regulators. Each enzyme reported is regulated by
different activators and inhibitors. Enzymes such as ALDH4A1, GOT1, GOT2, GPT1,
GPT2, GLS, and CAD are subject to regulation via feedback inhibition by GA. In addition
to product inhibition, substrate inhibition at different concentrations were also docu-
mented in glutamate metabolic pathway; α‐ketoglutarate is the substrate inhibitor of GOT2,
which inhibits the enzyme at high concentration. It has been found that enzymes that
metabolize glutamate are also regulated by interactions with proteins. Interactions between
the GLS enzyme and protein phosphatase 2, catalytic subunits, and ATCAY inhibit the
catalytic activity of the enzyme (Yelamanchi et al. 2016).
Methods for induction of GA production such as biotin limitation, Tween 40 addition,
and penicillin addition were also developed. It is well known that biotin has a marked
effect on L‐glutamic acid fermentation. Oleic acid‐requiring mutants were obtained from a
strain of Brevibacterium thiogenitalis which is an auxotroph for biotin. Palmitoleic acid and
linoleic acid could be used (Kanzaki et al. 1967). Recently, metabolic regulation during GA
production has been investigated at the molecular level. Moreover, novel microorganisms
such as Corynebacterium efficiens, which can produce glutamic acid at high temperatures,
and Pantoea ananatis, which can produce glutamic acid under acidic conditions, were iso-
lated (Hirasawa and Shimizu 2016).
A study on immobilization and reusability of cells was presented by Yelamanchi et al.
(2016). The production medium was inoculated with C. glutamicum and mixed culture of
C. glutamicum and Pseudomonas reptilivora with appropriate inoculum size. First, GA
yield was calculated for 24–72 hours. The preliminary study results showed that the mixed
culture of C. glutamicum and P. reptilivora produced higher yield than C. glutamicum
alone. The higher yield was 5.42 g/L with C. glutamicum and 7.96 g/L with mixed culture.
The optimized medium (glucose 50 g/L, urea 10 g/L, salt solution 19.24%) was used along
with standard concentration of biotin. Moreover, the optimized medium was chosen for
immobilization studies.
Also, experimental investigations were carried out on continuous and direct production
of L‐glutamic acid in a hybrid reactor system that integrated a conventional fermentative
production step with downstream membrane‐based separation and purification in flat
sheet cross‐flow membrane modules. Overcoming the substrate–product inhibitions of tra-
ditional batch production systems, this new compact, flexible, and largely fouling‐free
design ensured steady and continuous production of L‐glutamic acid directly from a renew-
able carbon source at the rate of about 8.4 g/L/h. Provisions of continuous product with-
drawal, separation, and recycling of unconverted sugar and microbial cells ensured almost
inhibition‐free production under high cell concentrations. Well‐screened nanofiltration
membranes with high selectivity helped achieve over 97% product purity while ensuring
recovery and recycling of more than 95% unconverted carbon source, resulting in a high
yield of 0.95 g/g. The direct production scheme involves no phase change or use of harsh
chemicals (Vikramachakravarthi et al. 2014).
Industrial production of GA is predominantly by microbial processes, although it is also
produced chemically. The direct fermentation method uses different carbon sources (glu-
cose, fructose, molasses, starch hydrolysates, n‐alkanes, ethanol, glycerol, acetate, propion-
ate); nitrogen (urea, ammonia salts, macerated corn liquid or soybean meal); and inorganic
salts of calcium, iron, manganese, zinc, cobalt, and biotin. C. glutamicum gave a higher
production of GA. Other industrially important strains belong to the genera Corynebacterium,
Brevibacterium, Mycobacterium, and Arthrobacter. Essential in the fermentation process is
the abundant supply of an adequate nitrogen source such as ammoniacal salts, since NH3
is incorporated into the amino acid molecule. However, the concentration of ammonium
ions should remain stable in the medium since too high concentrations are detrimental to
cell growth and product formation. In addition to the ammonia salts, it can be used as a
source of ammonium nitrogen (gaseous or in aqueous solution). In industrial production,
the addition of ammonium allows the control of pH and eliminates the problem of its tox-
icity. Most of the GA‐producing bacteria possess urease activity, so urea is also frequently
used as a source of nitrogen.
The concentration of oxygen must be balanced; lactate and succinate secretion occurs in
O2 deficiency, while excess oxygen with low concentration of ammonium leads to the inhi-
bition of growth and production of α‐ketoglutarate. Temperature affects the microbial
growth; fermentations are generally carried out in the mesophilic range (15–35 °C) and the
appropriate temperature must be chosen to achieve maximum growth and optimal product
formation. Since microorganisms have an optimum pH in which they have a higher growth
rate and yield, that value should be adopted. For C. glutamicum, it has been found that at
pH values between 6 and 9, the microorganism reaches homeostatis and maintains an opti-
mum internal pH of 7.5 ± 0.5. The Ajinomoto Group from Japan produces L‐glutamic acid
by fermentation.
The main uses of L‐glutamic acid and its derived products are as follows.
●● GABA precursor: glutamate also serves as the precursor for synthesis of inhibitory
γ‐aminobutyric acid (GABA) in GABA‐ergic neurons. This reaction is catalyzed by gluta-
mate decarboxylase (GAD), which is most abundant in the cerebellum and pancreas.
Stiff person syndrome is a neurologic disorder caused by anti‐GAD antibodies, leading to
a decrease in GABA synthesis and, therefore, impaired motor function such as muscle
stiffness and spasm. Since the pancreas has abundant GAD, direct immunologic destruc-
tion occurs in the pancreas and the patient will have diabetes mellitus.
●● Flavor enhancer: as a constituent of protein, GA is present in foods that contain protein,
but it can only be tasted when it is present in an unbound form. Significant amounts of
free GA are present in a wide variety of foods, including cheese and soy sauce, and are
responsible for umami, one of the five basic tastes. Glutamic acid is often used as a food
additive and flavor enhancer in the form of its sodium salt, known as MSG.
●● Nutrient: all meats, poultry, fish, eggs, dairy products, and kombu are excellent sources
of GA. Some protein‐rich plant foods also serve as sources. Thirty to 35% of gluten (much
of the protein in wheat) is GA. Ninety‐five percent of dietary glutamate is metabolized by
intestinal cells in a first pass.
●● Plant growth: Auxigro is a plant growth preparation that contains 30% glutamic acid.
●● Health: GA may treat personality and childhood behavioral issues. It may also aid in
epilepsy, muscular dystrophy, and intellectual disorders.
10.3.11 Gluconic Acid

Gluconic acid (GA) (C6H12O7) is an acid sugar belonging to the aldonic acid family. In
aqueous solutions, the acid is in equilibrium with its lactones. GA is a weak, nonvolatile,
harmless (odorless, noncorrosive, nontoxic), easily biodegradable acid that is soluble in
water and insoluble in nonpolar solvents. GA is commercially available as a 50% aqueous
solution with a pH of 1.82 and 1.23 g/cm3 density (Cañete‐Rodríguez et al. 2016).
Theoretically, gluconic acid can be produced by a wide variety of prokaryotic and eukary-
otic microorganisms, such as bacterial species of the genera Gluconobacter, Acetobacter,
and Pseudomonas, and fungal species of the genera Aspergillus, Penicillium, and
Gliocladium. A. niger remains the most efficient gluconate producer strain, showing prod-
uct yields of 98% from carbohydrate‐rich feedstocks.
Gluconic acid production is carried out using a medium rich in glucose and with high
levels of aeration. The oxidation of glucose with molecular oxygen to obtain D‐glucono‐
lactone is catalyzed by the enzyme glucose oxidase. This enzyme, which is usually intracel-
lular, is partially excreted or maintained extracellularly associated with the membrane of
certain filamentous fungi, especially in Aspergillus and Penicillium. Then the D‐glucono‐
lactone is hydrolyzed spontaneously to gluconate, releasing hydrogen peroxide that is
decomposed by the action of the catalase enzyme, which is present in the aforementioned
fungi. The enzymatic reactions for the formation of gluconic acid in Gluconobacter subox-
idans (bacteria) and A. niger (fungus) are depicted in Figure 10.5.
In bacteria, intracellular glucose is converted to extracellular gluconic acid. A membrane‐
bound enzyme, glucose dehydrogenase, utilizes pyrroloquinoline quinone (PQQ) as
(a)
Glucose dehydrogenase
PQQ PQQH2
H2O
D-Glucose D-Gluconolactone Gluconic acid

Lactonase
(b)
FAD FADH2
Catalase
H2O2 O2
Figure 10.5 Metabolic routes for gluconic acid in (a) bacterium (Gluconobacter suboxidans) and
(b) fungus (Aspergillus niger).
co‐enzyme and converts glucose to 5‐D‐gluconolactone which undergoes hydrolysis (spon-

taneous or enzymatic) to form gluconic acid. However, in fungal production, glucose is oxi-
dized by the extracellular enzyme glucose oxidase to form 8‐D‐gluconolactone, which
subsequently is converted to gluconic acid by lactonase. Glucose oxidase can be induced by
high concentrations of glucose and at pH above 4. It is believed that H2O2 produced by glu-
cose oxidase acts as an antagonist against other microorganisms (antimicrobial activity).
Submerged processes are normally used to produce gluconic acid by fermentation. The
culture medium contains large amount of glucose (110–250 g/L) and small amounts of
nitrogen compounds (>20 mmol/L N/P). The fermentation is carried out at pH 4.5–6.5 and
at 28–30 °C for a period of about 24 hours. It is an aerobic fermentation so increasing the
supply of O2 enhances gluconic acid yield. Because fungal growth and GA formation are
unrelated, continuous operation is impossible. As a result, most industrial processes involv-
ing A. niger are performed in batches, adding a neutralizing agent as GA accumulates
(Cañete‐Rodríguez et al. 2016).
Anastassiadis and Rehm (2006) have reported several continuous GA production meth-
ods using A. pullulans strains with and without biomass retention capable of producing up
to 375 g GA/L with 78% selectivity. This strain is a strictly aerobic yeast with a high osmo-
tolerance that degrades hexoses preferentially through the pentose phosphate pathway.
Gluconic acid was obtained with grape must as the feedstock and with A. niger, in an anaer-
obic fermentation (96 g/L, yield 0.70 g/g, productivity 0.8 g/L/h) as reported by Singh and
Singh (2006). From lignocellulosic biomass, gluconic acid was obtained in a batch fermen-
tation process using A. niger SIIM M276 and hydrolysis as a biomass pretreatment, with a
GA yield of 94.83% (Zhang et al. 2016).
Singh (2008), working in repeated batches of solid‐state surface fermentation, obtained
GA production rates as high as 22.5 g/L/d (i.e., 2.5 times greater than those of a typical
fermentation cycle), with 95–98% yield, by using A. niger ORS‐4.410 cells immobilized on
polyurethane foam. In the same research, the behavior of Ca‐alginate immobilized cells of
A. niger in submerged fermentations was also studied; as for similar highly aerobic systems,
the rate of oxygen transfer in the fermentation medium governed the process. The oxygen
mass transfer limitations required vigorous agitation, leading to important shear stresses,
which sharply affected the rate of GA production.
Based on renewable raw materials like maize, Roquette’s production plant in Italy pro-
duces an extensive range of gluconic derivatives: gluconic acid, glucono‐δ‐lactone (GDL)
and sodium gluconate. Gluconic acid is used in the manufacture of metals, stainless steel
and leather, as it can remove calcareous and rust deposits. It is also used as an additive to
foods and beverages since it acts as an acidulant with a delayed effect. This acid is used in
the dairy industry to delay sedimentation in milk. It is utilized in the manufacture of highly
resistant (to frost and cracking) concrete. Moreover, gluconic acid has pharmaceutical
applications for calcium and iron therapy. Sodium gluconate is used as a sequestering agent
in many detergents.
10.3.12 Adipic Acid

Adipic acid (AA), or hexanedioic acid (C6H10O4), is one of the most produced chemicals
worldwide. Its density is 1.360 g/cm3. It rarely occurs in Nature. AA is known to be a versa-
tile building block for an array of processes in the chemical, pharmaceutical, and food
industries. Its primary use is as a precursor for the synthesis of the polyamide nylon‐6,6. It
was obtained by Dieterle by the oxidation of castor oil with nitric acid. It was synthesized
in 1902 from tetramethylene bromide. Adipic acid has traditionally been produced from
various petroleum‐based feedstocks (phenol, benzene, and cyclohexane). In recent years,
cyclohexane‐based processes have accounted for about 93% of global production. Two steps
are involved in AA production: oxidation of cyclohexane to produce KA oil (cyclohexanone
and cyclohexanol) and nitric acid oxidation of KA oil to produce adipic acid (Yu et al. 2006).
Instead of oxidizing one specific linear or cyclic C6 molecule, AA production could come
from a more abundant and versatile feedstock such as lignocellulose. The conversion of
biomass components, in particular cellulose, hemicellulose, and lignin, is a current topic of
intense scientific research. Since AA is not naturally produced by any known organism,
attempts to produce it through fermentation require heterologous expression of pathways
that can convert intracellular metabolites into AA. Three biotechnologic routes are known
to produce AA: the biosynthesis of cis,cis‐muconic acid from glucose by fermentation, fol-
lowed by its chemical catalytic hydrogenation to AA; the enzymatic conversion of benzene
or cyclohexanol to AA; and the enzymatic conversion of adiponitrile to ammonium adi-
pate. Of these three routes, the most interesting and promising is the first, since it is the
only one that involves the use of a renewable raw material.
Several pathways have been proposed for the conversion of 5‐HMF into AA. For exam-
ple, in 1981, Faber (Hydrocarbon Research Inc., Lawrenceville, NJ, US) patented a multi-
step process for AA synthesis based on lignocellulosic biomass‐derived 5‐HMF. The four
steps in this process are: (i) acid‐catalyzed hydrolysis/dehydration of lignocellulose to
5‐HMF in aqueous solutions of H2SO4; (ii) RANEYs‐Ni catalyzed hydrogenation of 5‐HMF
to 2,5‐dihydroxymethyl‐tetrahydrofuran (DHMTHF); (iii) hydrogenolysis of DHMTHF to
1,6‐hexanediol with a copper chromite catalyst in a fixed‐bed reactor; and (iv) biocatalytic
oxidation of 1,6‐hexanediol (1,6‐HD) to AA by Gluconobacter oxydans (van de Vyver and
Román‐Leshkov 2013). Rennovia uses a patented catalytic chemical process to produce
AA, using glucose as a raw material. The technology uses heterogeneous catalysis, which
first produces glucaric acid through aerobic oxidation of glucose followed by catalytic
hydrogenation to AA (van de Vyver and Román‐Leshkov 2013).
In a biochemical alternative route, strains of E. coli have been constructed to produce AA
from D‐glucose. The enzymatic method proceeds via an aerobic pathway in which
D‐glucose is transformed into the intermediates catechol and cis,cis‐muconic acid
(cis,cis‐2,4‐hexadienodioic acid), respectively. The conversion to AA is performed in a chem-
ocatalytic step by hydrogenating cis,cis‐muconic acid (MA) with Pt/C, bimetallic RuPt nan-
oparticles, or titania‐supported Re catalysts. The conversion of MA into AA provides a yield
of 97%. However, the production of MA from glucose has a low yield (24%), combined with
difficulties in its purification. In fermentations carried out in fed‐batch mode, cis,cis‐
muconic acid production close to 37 g/L has been described, corresponding to 22% yields (in
mol/mol of consumed glucose), which is approximately 50% of the theoretical maximum.
A recombinant microorganism has been created, specifically by introducing three exog-
enous genes: those that encode the enzymes dehydroshikimate dehydratase and proto-
catecuate decarboxylase of K. pneumoniae, and catechol 1,2‐dioxygenase of Acinetobacter
calcoaceticus. In the recombinant microorganism, the carbon flux directed toward the com-
mon route of synthesis of aromatic amino acids is sidetracked toward the synthesis of
cis,cis‐muconic acid. The MA obtained in the fermentation is finally hydrogenated to AA
at high pressure by means of a platinum catalyst, a process that has a yield of 97%.
More recently, the total biosynthesis (the conversion of substrate to final product in a
single process) of AA directly from biomass feedstock has been the focus of much attention
in the biotechnology sector. A total of eight unique pathways are proposed in four patents
(Picataggio et al. 1992; Baynes and Geremia 2010; Burgard et al. 2010; Raemakers‐Franken
et al. 2010) (Table 10.4).
The current industrial process for the production of AA relies on the catalytic oxida-
tion of a mixture of cyclohexanol and cyclohexanone, also referred to as KA oil. A
review by Cavani and Alini (2009) describes different methods to produce the KA oil,
Table 10.4 Yields reported for several processes to produce adipic acid from glucose.
Pathway Yield (mol adipate/

(starting from) Microorganism, condition mol glucose) References
Oxoglutarate Escherichia coli aerobic 0.83 Burgard et al. (2010)

pathway 1 E. coli anaerobic 0.36
Oxoglutarate E. coli aerobic 0.81 Burgard et al. (2010)
pathway 2 E. coli anaerobic 0.32
Oxoglutarate E. coli aerobic 0.68 Baynes and Geremia
pathway 3 E. coli anaerobic 0.20 (2010)
3‐Oxoadipate E. coli aerobic 0.92 Burgard et al. (2010)
Lysine E. coli aerobic 0.87 Burgard et al. (2010)
E. coli anaerobic 0.57
Malonyl‐CoA E. coli aerobic 0.37 Picataggio et al. (1992)
the most common being the cobalt‐catalyzed oxidation of benzene‐derived cyclohexane

with air. Unfortunately, conversion is very low (4–8%), so recycling of unconverted
cyclohexane is necessary. The second step of the process involves oxidation of the KA oil
with an excess of HNO3 in the presence of copper(II) and ammonium metavanadate
catalysts.
Cyclohexene could be oxidized to AA in a biphasic system using 30% H2O2 in the pres-
ence of Na2WO4 as a homogeneous catalyst and [CH3(n‐C8H17)3N]HSO4 as a phase‐transfer
catalyst. Because cyclohexene and water are almost immiscible, their mixture creates a
two‐phase system containing the tungstate and H2O2 in the aqueous phase, and the phase
transfer catalyst in the organic cyclohexene phase. At a temperature of 75–90 °C, Na2WO4
is oxidized by H2O2 into an anionic peroxo species that is extracted by the quaternary
ammonium cation into the organic phase. The reaction with cyclohexene restores the
reduced form of the catalyst and returns it into the aqueous phase to initiate a new catalytic
cycle. The solvent‐ and halide‐free process affords analytically pure AA crystals in yields of
90% after eight hours of reaction (Sato et al. 1998).
Sporadic reports over recent decades have proposed the direct oxidation of cyclohexane
to AA using Co and Mn catalysts with acetic acid as the solvent. Inspired by this work,
Bonnet et al. (2006) investigated the combination of lipophilic carboxylic acids and low
loadings of Co and Mn salts. The choice of lipophilic carboxylic acids was motivated by the
need to facilitate their recycling after partitioning of AA in the aqueous phase. A screening
of several combinations led to the selection of 4‐tertbutylbenzoic acid with ppm levels of
Co and Mn as the most effective system. The applicability of the method was further dem-
onstrated in semi‐batch and continuous‐flow experiments in a 1 L reactor under 2 MPa bar
air, resulting in a maximum selectivity of 71% AA and a productivity of 95 g/L/h (van de
Vyver and Román‐Leshkov 2013).
Alshammari et al. (2012) showed that the gold‐catalyzed oxidation of cyclohexane to AA
is possible to some extent by fine‐tuning the particle size and dispersion of Au nanoparti-
cles on TiO2. Using acetonitrile as solvent and t‐BHP as initiator, the highest AA yield was
8% after four hours of reaction at 150 °C and 1 MPa of O2. In another study, Yu et al. (2011)
approached the field from a new angle by pioneering the use of multiwalled carbon nano-
tubes (CNTs) as a metal‐free catalyst for the aerobic oxidation of cyclohexane. The perfor-
mance of nitrogen‐doped CNTs was demonstrated to exceed the activity of most Au‐based
catalysts. For instance, after reacting for over eight hours at 125 °C and 1.5 MPa of O2, N‐
doped CNTs gave 45% conversion with 60% AA selectivity. The inhibition of catalytic activ-
ity in the presence of p‐benzoquinone as a radical scavenger indicated that this CNT‐catalyzed
oxidation reaction proceeds through a similar radical chain mechanism as proposed for the
liquid‐phase autoxidation of cyclohexane.
Major global producers of AA include Invista, Ascend, Honeywell, BASF, Radici,
China Shenma, and PetroChina. High‐purity fiber‐grade AA is used to make nylon‐66,
while lower purity AA is used primarily to produce polyurethanes. Recently, start‐up
companies such as Rennovia, Verdezyne, BioAmber, Celexion, and Genomatica have
developed bio‐based routes to produce AA, aiming at creating 100% bio‐based nylon;
some have reached advanced pilot or demonstration scales (van de Vyver and Román‐
Leshkov 2013).
The main uses of AA include the following.

●● Most of the millions of kg of AA produced annually are used as a monomer for the pro-
duction of nylon by a polycondensation reaction forming nylon‐6.6 which is processed to
manufacture clothes, car tires, and carpets.
●● It is also used in the manufacture of plasticized products and lubricants, applications
that also involve polymers, with which the monomer is used in the production of polyu-
rethane and its esters, especially in PVC.
●● In the food industry, it is also applied as an assistant in gelatinization, as an acidulant,
buffer agent and leavening agent.
●● It is widely used in the paint and coatings industry.
10.4 Downstream Processes
Separation and purification of the target compound and the elimination of wastes from the
fermentation process and others are included in downstream processing. The first step
should be the removal of dead cells from fermentation, mainly by filtration or centrifuga-
tion. To upgrade purity, ion exchange columns, extraction with solvents or other processes
with high selectivity could be included. Finally, crystallization of the broth, together with a
centrifugation and drying process, make up the purification stage of the product.
Some classic technologies for purification processes include distillation, adsorption,
liquid–liquid extraction, extraction with supercritical fluids, pervaporation, ionic exchange,
crystallization, membrane separation technologies, and chromatography. Application for
the purification of acid organic products from feedstocks requires more research in order
to diminish process costs. Low‐cost process is necessary in a biorefinery, mainly because of
low concentration of the target product during fermentation, requiring several purification
steps that diminish the overall yield.
Especially in fermentative processes with high product concentration, volumetric pro-
duction can be limited by either the inhibition of the products themselves or by degrada-
tion. In these cases, rapid reduction of the compounds formed is required to prevent
interactions between the products and the cell medium. An example of an environmentally
friendly application is the separation of salts of organic acids produced by fermentation by
electrodialysis with bipolar membranes (electrohydrolysis) to obtain an aqueous stream of
organic acid that can be subsequently isolated by, for example, evaporation and crystalliza-
tion, and also, an aqueous stream of the alkaline agent that is recycled to the fermentation
process (Huang et al. 2006). Specific purification processes have been reported in patents
and papers.
For itaconic acid downstream process, a filtration process could be used to remove the
medium, then the clarified liquid could pass to an evaporation process. The last step could
be crystallization, or an alternative process such as ionic exchange or extraction with sol-
vents. Separation of itaconic acid relies on operations such as crystallization, precipitation,
extraction, electrodialysis, diafiltration, and adsorption.
Everything considered, there are two main directions for the recovery of IA from fer-
mented broths: the use of chemical‐demanding, reaction‐based separations, and the use of
energy‐demanding, physical separations. Some can be used in processes coupled to

fermentation. Chemical‐intensive operations such as precipitation, extraction, and
adsorption reduce the volume for further processing without using large amounts of
energy, and thus may continue to be used for IA production, despite the generation of by‐
products. Extraction and adsorption are the most promising operations in this direction.
Crystallization can be used as a recovery and a polishing operation. As recovery operation,
it is relatively simple and very efficient, but demands high energy and relatively costly
equipment. However, it gives a pure product. A single crystallization step is not enough to
recover all the products in a stream, and there must always be a recycling step. As a polish-
ing operation, crystallization is used to crystallize solutions previously concentrated and in
this case, the size, cost, and energy requirements are lower. Crystallization is very common
for polishing steps. Other methods such as membrane separation are promising as a recov-
ery step coupled to the fermentation, potentially enhancing the overall process yield.
Another approach is adsorption in fixed‐bed columns, which efficiently separates IA.
Despite recent advances in separation and recovery methods, there is still space for
improvement in IA recovery and purification (Magalhães et al. 2016).
For FDCA, a general description of the purification process establishes that after fermen-
tation of P. putida S12 B‐38 and P. putida S12 B‐51, the obtained FDCA was recovered from
the broth by acid precipitation (for low pH), cooling, crystallization, and separation in crys-
tallized form. The whole‐cell biocatalyst P. putida S12 is removed from the broth by centrifu-
gation at 9500 g for 10 minutes. Collected supernatant is boiled for three minutes to remove
unwanted proteins followed by acidification with 96% H2SO4 at 4 °C and lowering the pH to
0.5. Precipitate is centrifuged and the pellet contains FDCA. This pellet is washed with dis-
tilled water and centrifuged again. Then the pellet is dried in air and dissolved in tetrahydro-
furan (THF) and shaken for 30 minutes at room temperature. This THF is vacuum dried at
50 °C until a clear dry powder is obtained. After elemental analysis, the authors confirmed
that the off‐white powder consisted of 99.4 ± 0.28% of pure FDCA (Wierckx et al. 2012).
Another description shows that after fermentation, pH is adjusted to 3 and a precipitate
is obtained. This precipitate is resuspended in water and again pH adjusted to 3. The
obtained precipitate is kept in a refrigerator for two days and air dried, spun down, and
again air dried. About 15 mg of FDCA was obtained from this reaction and the purity was
confirmed by TLC (Hanke 2009). Other authors used an upstream process with an enzy-
matic cascade reaction with galactose oxidase M3–5, periplasmic aldehyde oxidase
(PaoABC), and a catalase for sequential oxidations of HMF. After the enzyme process, reac-
tion mixture was heated to 80 °C for five minutes and allowed to cool. After 17 hours, reac-
tion was stopped. For purification, its supernatant was cooled to 0 °C after centrifugation
and concentrated HCl was added drop‐wise until a precipitation formed. The solution was
then again centrifuged and the obtained pellet washed with 1 M HCl. The pellet was dis-
solved in acetone followed by concentration (in vacuum) three times and FDCA was
obtained as a pale yellow solid with 74% yield (McKenna et al. 2015).
For gluconic acid, a downstream process is proposed with the following steps: cell separa-
tion by centrifugation or ultrafiltration, separation, evaporation, and drying. Specifically, a
patent claims that gluconic acid obtained from crude fermentation solutions is purified by
multistep processes including biomass separation (filtration), carbon treatment (decoloriza-
tion), evaporation (concentration), and crystallization (purification) to provide a final product
with high purity (Anastassiadis and Rehm 2006). For glutamic acid, the produced crude glu-
tamic acid is filtered, purified, and neutralized to MSG. After further purification, crystalliza-
tion and drying, the glutamate is in the form of white crystals ready to be packaged.
Numerous separation techniques, such as precipitation, ion exchange adsorption, solvent
extraction and electrodialysis, have been successfully established for the downstream pro-
cessing of fermented organic acids such as succinic acid (Xu et al. 2010). The recovery tech-
niques for fumaric acid production have not been well developed. The most common
mechanism of fumaric acid recovery is precipitation, but the concentration of fumaric acid in
the broth needs to be limited to 50 g/L to prevent complications during the separation pro-
cess. This process has no special equipment requirements, and the apparatus used in citric
acid and lactic acid production can also be used for fumaric acid production (Xu et al. 2010).
Various methods may be practiced to remove biomass and/or separate 3‐HPA from the
culture broth and its components. Methods to separate and/or concentrate the 3‐HPA
include centrifugation, filtration, extraction, chemical conversion such as esterification,
distillation (which may result in chemical conversion, such as dehydration to acrylic acid,
under some reactive‐distillation conditions), crystallization, chromatography, and ion
exchange, in various forms. Additionally, cell rupture may be conducted as needed to
release 3‐HPA from the cell mass, such as by sonication, homogenization, pH adjustment
or heating. 3‐HPA may be further separated and/or purified by methods including any
combination of centrifugation, liquid–liquid separations, including extractions such as sol-
vent extraction, reactive extraction, two‐phase aqueous extraction and two‐phase solvent
extraction, membrane separation technologies, distillation, evaporation, ion exchange
chromatography, adsorption chromatography, reverse phase chromatography, and crystal-
lization (Lynch et al. 2011).
10.5 Conclusion
Several organic acids can be obtained from lignocellulosic biomass. Although many chemi-
cal conversion technologies are in use, biological technologies require more research. Only
a few acids can currently be produced by these biological technologies. Research necessary
to improve the processes are related to sugars fermentation, such as development of known
or new microorganisms with higher productivity, resistance to stress, and tolerance to high
concentrations of substrates, products and by‐products; development of microorganisms
capable of fermenting various C6 and C5 sugars; manipulation of microbial metabolism
(genetic and metabolic engineering) to direct it toward the synthesis of products of high
interest (natural or not); development of microorganisms capable of simultaneous sac-
charification of biomass and fermentation of products obtained in saccharification;
increase in the volumetric productivity of fermentations; design of more efficient ferment-
ers and improvement of the fermentation processes and their control; reduction of enzyme
cost; development of continuous fermentation; improvement of reaction yields through
the development of more specific and selective biocatalysts; optimization of processes;
improvements in the immobilization of enzyme technologies.
For the purification process, researchers need to study new concepts of separation tech-
nology different from those used in oil refineries and petrochemicals, such as extraction
with supercritical fluids; processes based on selective permeable membranes; combination

and integration of different separation principles, such as solvent extraction with mem-
brane permeation; separation and purification of products; integration of separation and
production phases; introduction and optimization of on‐site product recovery processes.
Although many organic acids will be produced from biomass in the future, and a few in
the near future, several challenges still remain to optimize commercialization. Academic
research and industrial development should be reinforced to reduce the gap between labo-
ratory and industrial process. Advances in synthetic biology and metabolic engineering
will allow the development of high‐performance strains. This is the greatest challenge that
requires much time, effort, and financial input.
References
Alonso, S., Rendueles, M., and Díaz, M. (2015). Microbial production of specialty organic acids
from renewable and waste materials. Critical Reviews in Biotechnology 35 (4): 497–513.
Alshammari, A., Koeckritz, A., Kalevaru, V.N. et al. (2012). Significant formation of adipic acid
by direct oxidation of cyclohexane using supported nano‐gold catalysts. ChemCatChem 4:
1330–1336.
Alvira, P., Tomas‐Pejo, E., Ballesteros, M., and Negro, M.J. (2010). Pretreatment technologies
for an efficient bioethanol production process based on enzymatic hydrolysis: a review.
Anastassiadis, S. and Rehm, H.J. (2006). Continuous gluconic acid production by
Aureobasidium pullulans with and without biomass retention. Electronic Journal of
Bain, R.L. and Broer, K. (2011). Gasification. In: Thermochemical Processing of Biomass (eds.
R.C. Brown and C. Stevens). Chichester, UK: Wiley.
Baynes, B. and Geremia, J. (2010). Biological synthesis of difunctional alkanes from alpha
ketoacids. US Patent WO2010068944, 17 June 2010.
Bio Refineries Blog (2018). https://biorrefineria.blogspot.com/2015/10/biorrefinerias‐de‐acido‐
succinico.html.
Blumhoff, M.L., Steiger, M.G., Mattanovich, D., and Sauer, M. (2013). Targeting enzymes to the
right compartment: metabolic engineering for itaconic acid production by Aspergillus niger.
Metabolic Engineering 19: 26–32.
Bonnet, D., Ireland, T., Fache, E., and Simonato, J.P. (2006). Innovative direct synthesis of
adipic acid by air oxidation of cyclohexane. Green Chemistry, vol. 8: 556–559.
Borges, E.R. and Pereira, N. Jr. (2011). Succinic acid production from sugarcane bagasse
hemicellulose hydrolysate by Actinobacillus succinogenes. Journal of Industrial Microbiology
and Biotechnology 38: 1001–1011.
Borodina, I., Kildegaard, K., Jensen, N. et al. (2015). Establishing a synthetic pathway for
high‐level production of 3‐hydroxypropionic acid in Saccharomyces cerevisiae via β‐alanine.
Brockman, I., Stenger, A., Reisch, C. et al. (2015). Improvement of glucaric acid production in
E. coli via dynamic control of metabolic fluxes. Metabolic Engineering Communications 2:
109–116.
Brown, S.H., Bashkirova, L., Berka, R. et al. (2013). Metabolic engineering of Aspergillus oryzae
NRRL 3488 for increased production of L‐malic acid. Applied Microbiology and
Burgard, P., Pharkya, P., and Osterhout, R. (2010). Microorganisms for the production of adipic
acid and other compounds. US Patent 7,799,545, B2, 21 September 2010.
Cañete‐Rodríguez, A.M., Santos‐Dueñas, I.M., Jimenez‐Hornero, J., and García‐García, I.
(2016). Gluconic acid: properties, production methods and applications – an excellent
opportunity for agro‐industrial by‐products and waste bio‐valorization. Process Biochemistry
51 (12): 1891–1903.
Cárdenas‐Fernández, M., López, C., Alvaro, G., and López‐Santín, J. (2012). Immobilized L‐
aspartate ammonia‐lyase from Bacillus sp. YM55‐1 as biocatalyst for highly concentrated
L‐aspartate synthesis. Bioprocess and Biosystems Engineering 35: 1437–1444.
Carrillo‐Nieves, D., Zumalacárregui, L., Sánchez, O. et al. (2014). Pretreatments employed in
lignocellulosic materials for bioethanol production: an overview. ICIDCA sobre los derivados
de la caña de azúcar 48 (1 (enero‐abril)): 71–79.
Carro, J., Fernández‐Fueyo, E., Fernández‐Alonso, C. et al. (2018). Self‐sustained enzymatic
cascade for the production of 2,5‐furandicarboxylic acid from 5‐methoxymethylfurfural.
Biotechnology for Biofuels 11: 86.
Carta, F., Soccol, C., Ramos, L., and Fontana, J. (1999). Production of fumaric acid by
fermentation of enzymatic hydrolysates derived from cassava bagasse. Bioresource
Cavani, F. and Alini, S. (2009). Synthesis of adipic acid: on the way to a more sustainable
production. In: Sustainable Industrial Chemistry (eds. F. Cavani, G. Centi, S. Perathoner and
F. Trifiró), 367–425. Weinheim, Germany: Wiley‐VCH.
Chan, S., Kanchanatawee, S., and Jantama, K. (2012). Production of succinic acid from sucrose
and sugarcane molasses by metabolically engineered Escherichia coli. Bioresource Technology
103: 329–336.
Chandel, A.K. and Silveira, M.H.L. (2018). Advances in Sugarcane Biorefinery: Technologies,
Commercialization, Policy Issues and Paradigm Shift for Bioethanol and by‐Products. Oxford,
UK: Elsevier.
Chen, N., Wang, J., Zhao, Y., and Deng, Y. (2018). Metabolic engineering of Saccharomyces
cerevisiae for efficient production of glucaric acid at high titer. Microbial Cell Factories 17: 67.
Chen, Y., Bao, J., Kim, I.K. et al. (2014). Coupled incremental precursor and co‐factor supply
improves 3‐hydroxypropionic acid production in Saccharomyces cerevisiae. Metabolic
Chen, Z., Huang, J., Wu, Y., and Liu, D. (2017). Metabolic engineering of Corynebacterium
glutamicum for the production of 3‐hydroxypropionic acid from glucose and xylose.
Chibata, I., Tosa, T., and Sato, T. (1986). Continuous production of L‐aspartic acid. Applied
Biochemistry and Biotechnology 13 (3): 231–240.
Choi, S., Song, C.W., Shin, J.H., and Lee, S.Y. (2015). Biorefineries for the production of top
building block chemicals and their derivatives. Metabolic Engineering 28: 223–239.
Chu, H.S., Kim, Y.S., Lee, C.M. et al. (2015). Metabolic engineering of 3‐hydroxypropionic acid
biosynthesis in Escherichia coli. Biotechnology and Bioengineering 112 (2): 356–364.
Chundawat, S.P.S., Beckham, G.T., Himmel, M.E., and Dale, B.E. (2011). Deconstruction of
lignocellulosic biomass to fuels and chemicals. Annual Review of Chemical and Biomolecular
Daniell, J., Köpke, M., and Simpson, S. (2012). Commercial biomass syngas fermentation.
Energies 5 (12): 5372–5417.
Derbikov, D.D., Novikov, A.D., Gubanova, T.A. et al. (2017). Aspartic acid synthesis by
Escherichia coli strains with deleted Fumarase genes as biocatalysts. Applied Biochemistry
and Microbiology 53 (9): 859–866.
de la Rosa, S.M. (2015). Acid hydrolysis of cellulose and lignocellulosic biomass assisted with
ionic liquids. Doctoral thesis in Chemical Sciences. Autonomous University of Madrid
(original in Spanish).
Eisentraut, A. (2010). Sustainable Production of Second‐Generation Biofuels. Paris: IEA
Renewable Energy Division.
FEDIT (2008). Trends in biotechnology uses in the chemical sector. Technological Centres of
Spain, p. 102.
Galbe, M. and Zacchi, G. (2002). A review of the production of ethanol from softwood. Applied
Gandini, A., Silvestre, A.J.D., Neto, C.P. et al. (2009). The furan counterpart of poly (ethylene
terephthalate): an alternative material based on renewable resources. Journal of Polymer
Science Part A: Polymer Chemistry 47 (1): 295–298.
Ganigué, R., Sánchez‐Paredes, P., Bañeras, L., and Colprim, J. (2016). Low fermentation pH is
a trigger to alcohol production, but a killer to chain elongation. Frontiers in Microbiology 7:
702–713.
Garza, Y. (2000). Development of aspartate ammonia lyase activity in microorganism’s cell for
the synthesis of L‐aspartic amino acid. PhD thesis. Autonomous University of Nuevo León.
Gil, N., Ferreira, S., Amaral, M. et al. (2010). The influence of dilute acid pretreatment
conditions on the enzymatic saccharification of Erica spp. for bioethanol production.
Giraldo, S. (2013). Ensayos para la producción biotecnológica de ácido fumárico empleando
residuos agrícolas. Tesis de Ingeniero Químico. Universidad Pontificia Bolivariana,
Medellín, Colombia.
Hanke, P.D. (2009). Patent WO2009/023174, 19 February 2009.
Hayes, D.J., Fitzpatrick, S., Hayes, M.H.B., and Ross, J.R.H. (2006). The biofine process–
production of levulinic acid, furfural, and formic acid from lignocellulosic feedstocks. In:
Biorefineries – Industrial Processes and Prodwucts (eds. B. Kamm, P.R. Gruber and M.
Kamm), 139–164. Weinheim, Germany: Wiley‐VCH.
Henstra, A.M., Sipma, J., Rinzema, A., and Stams, A.J. (2007). Microbiology of synthesis gas
fermentation for biofuel production. Current Opinion in Biotechnology 18 (3): 200–206.
Hirasawa, T. and Shimizu, H. (2016). Glutamic acid fermentation: discovery of glutamic acid‐
producing microorganisms, analysis of the production mechanism, metabolic engineering,
and industrial production process. In: Industrial Biotechnology: Products and Processes (eds.
C. Wittmann and J.C. Liao). Weinheim, Germany: Wiley‐VCH.
HIS (2017). Chemical Economics Handbook. https://ihsmarkit.com/about/index.html.
Huang, C., Xu, T., Zhang, Y. et al. (2006). Application of electrodialysis to the production of
organic acids: state‐of‐the‐art and recent developments. Journal of Membrane Science 288
(1–2): 1–12.
Jang, Y.S., Kim, B., Shin, J.H. et al. (2012). Bio‐based production of C2‐C6 platform chemicals.
Biotechnology and Bioengineering 10: 2437–2459.
Jansen, M.L.A. and Verwaal, R. (2017). Succinic acid production by fermentation at low pH.
European patent: EP 2297297, 8 February 2017.
Jantama, K., Zhang, X., Moore, J.C. et al. (2008). Eliminating side products and increasing
succinate yields in engineered strains of Escherichia coli C. Biotechnology & Bioengineering
101: 881–893.
Jensen, N., Borodina, I., Chen, Y. et al. (2014). Microbial production of 3‐hydroxypropionic
acid. WO Patent Application, 2014/198831A1.
Jeong, G.T. (2014). Production of levulinic acid from glucosamine by dilute‐acid catalyzed
hydrothermal process. Industrial Crops and Products 62: 77–83.
Jung, I.Y., Lee, J.W., Min, W.K. et al. (2015). Simultaneous conversion of glucose and xylose to
3‐hydroxypropionic acid in engineered Escherichia coli by modulation of sugar transport and
glycerol synthesis. Bioresource Technology 198: 709–716.
Kamm, B. and Kamm, M. (2007). International biorefinery systems. Pure and Applied
Chemistry 79 (11): 1983–1997.
Kang, Z. and Gong, X. (2016). Biosynthesis of glucaric acid with microbial cell factories.
Research & Reviews: Journal of Microbiology and Biotechnology 5 (4): 39–44.
Kanzaki, T., Iso, K., Okazaki, H. et al. (1967). L‐glutamic acid fermentation: part I. selection of
an oleic acid‐requiring mutant and its properties, part II: the production of l‐glutamic acid
by an oleic acid‐requiring mutant. Agricultural and Biological Chemistry 31 (11): 3107–3117.
Karich, A., Kleeberg, A., Ullrich, R., and Hofrichter, M. (2018). Enzymatic preparation of 2,5‐
furandicarboxylic acid (FDCA), a substitute of terephthalic acid by the joined action of three
fungal enzymes. Microorganisms 6 (5): ii.
Kautola, H. and Linko, Y. (1989). Fumaric acid production from xylose by immobilized
Rhizopus arrhizus cells. Applied Microbiology and Biotechnology 31: 448–452.
Khan, I., Qayyum, S., Maqbool, F. et al. (2017). Microbial organic acids production,
biosynthetic mechanism and applications – mini review. Indian Journal of Geo Marine
Sciences 46 (11): 2165–2174.
Kildegaard, K., Wang, Z., Chen, Y. et al. (2015). Production of 3‐hydroxypropionic acid from
glucose and xylose by metabolically engineered Saccharomyces cerevisiae. Metabolic
Engineering Communications 2: 132–136.
Kim, S.H., Huang, Y., Sawatdeenarunat, C. et al. (2011). Selective sequestration of carboxylic
acids from biomass fermentation by surface‐functionalized mesoporous silica nanoparticles.
Journal of Materials Chemistry 21 (32): 12103–12109.
Koopman, F., Wierckx, N., de Winde, J.H., and Ruijssenaars, H.J. (2010a). Identification and
characterization of the furfural and 5‐(hydroxymethyl) furfural degradation pathways of
cupriavidus basilensis HMF14. Proceedings of the National Academy of Science 107 (11):
4919–4924.
Koopman, F., Wierckx, N., de Winde, J.H., and Ruijssenaars, H.J. (2010b). Efficient whole‐cell
biotransformation of 5‐(Hydroxymethyl) furfural into FDCA, 2,5‐furandicarboxylic acid.
Kuenz, A., Gallenmuller, Y., Willke, T., and Vorlop, K.D. (2012). Microbial production of
itaconic acid: developing a stable platform for high product concentrations. Applied
Kwak, S., Park, Y.C., and Seo, J.H. (2013). Biosynthesis of 3‐hydroxypropionic acid from
glycerol in recombinant Escherichia coli expressing Lactobacillus brevis dhaB and dhaR gene
clusters and E. coli K‐12 aldH. Bioresource Technology 135: 432–439.
La Roe, E. (1959). Fumaric acid fermentation process. US Patent 2,912,363.
Li, X., Zhou, J., Ouyang, S. et al. (2016). Fumaric acid production from alkali‐pretreated
corncob by fed‐batch simultaneous saccharification and fermentation combined with
separated hydrolysis and fermentation at high solids loading. Applied Biochemistry and
Liao, H.H., Gokarn, R.R., Gort, S.J. et al. (2007). Production of 3‐hydropropionic acid using
beta‐alanine/pyruvate aminotransferase. Patent Application US2007/0107080A1.
Liao, W., Liu, Y., Frear, C., and Chen, S.L. (2008). Co‐production of fumaric acid and chitin
from a nitrogen‐rich lignocellulosic material – dairy manure – using a pelletized
filamentous fungus Rhizopus oryzae ATCC 20344. Bioresource Technology 99: 5859–5866.
Liew, F., Martin, M.E., Tappel, R.C. et al. (2016). Gas fermentation‐a flexible platform for
commercial scale production of low‐carbon‐fuels and chemicals from waste and renewable
feedstocks. Frontiers in Microbiology 7: 694.
Lin, C.S.K., Luque, R., Clark, J.H. et al. (2011). Wheat‐based biorefining strategy for
fermentative production and chemical transformations of succinic acid. Biofuels,
Bioproducts and Biorefining 6: 88–104.
Liu, X., Lv, J., Zhang, T., and Deng, Y. (2014). Direct conversion of pretreated straw cellulose
into citric acid by co‐cultures of Yarrowia lipolytica SWJ‐1b and immobilized Trichoderma
reesei mycelium. Applied Biochemistry and Biotechnology 173 (2): 501–509.
Lui, Y., Gong, X., Wang, C. et al. (2016). Production of glucaric acid from myo‐inositol in
engineered Pichia pastoris. Enzyme and Microbial Technology 91: 8–16.
Luo, H., Zhou, D., Liu, X. et al. (2016). Production of 3‐hydroxypropionic acid via the
propionyl‐CoA pathway using recombinant Escherichia coli strains. PLoS One 11: e0156286.
Lynch, M.D., Gill, R.T., and Warnecke‐Lipscomb, T. (2011). Method for producing 3‐
hydroxypropionic acid and other products. European Patent: PCT/US2010/050436.
Lynd, L.R., Weimer, P.J., van Zyl, W.H., and Pretorius, I.S. (2002). Microbial cellulose
utilization: fundamentals and biotechnology. Microbiology and Molecular Biology Reviews 66:
506–577.
Magalhães, A.I., de Carvalho, J.C., Medina, J.D.C., and Soccol, C.R. (2016). Downstream
process development in biotechnological itaconic acid manufacturing. Applied Microbiology
Martín, C. and Carrillo, E. (2009). Determination of the chemical composition of tropical
cellulosic materials by the detergent sequential system combined with acid hydrolysis.
Cellulose Chemistry and Technology 40 (6): 399–403.
Matano, Y., Hasunuma, T., and Kondo, A. (2012). Display of cellulases on the cell surface of
Saccharomyces cerevisiae for high yield ethanol production from high‐solid lignocellulosic
biomass. Bioresource Technology 108: 128–133.
Matsakas, L., Hruzová, K., Ulrika Rova, U., and Christakopoulos, P. (2018). Biological production
of 3‐hydroxypropionic acid: an update on the current status. Fermentation 4 (1): 13.
McKenna, S., Leimkühler, S., Herter, S. et al. (2015). Enzyme cascade reactions: synthesis of
furandicarboxylic acid (FDCA) and carboxylic acids using oxidases in tandem. Green
Chemistry 17: 3271–3275.
Moon, T.S., Yoon, S.H., Lanza, A.M. et al. (2009). Production of glucaric acid from synthetic
pathway in recombinant Escherichia coli. Applied and Environmental Microbiology 75: 589–595.
Munasinghe, P.C. and Khanal, S.K. (2010). Biomass‐derived syngas fermentation into biofuels:
opportunities and challenges. Bioresource Technology 101 (1): 5013–5022.
Okabe, M., Lies, D., Kanamasa, S., and Park, E.Y. (2009). Biotechnological production of
itaconic acid and its biosynthesis in Aspergillus terreus. Applied Microbiology and
Oswald, F., Dörsam, S., Veith, N. et al. (2016). Sequential mixed cultures: from syngas to malic
acid. Frontiers in Microbiology 7 (891) https://doi.org/10.3389/fmicb.2016.00891.
Oviedo, J., Castrillón, P., Ramírez, M., and Vanegas, D. (2009). Preliminary study for the
production of L‐aspartic acid in Pseudomonas fluorescens. Revista Investigaciones Aplicadas
5: 26–33.
Perez, J.M., Richter, H., Loftus, S.E., and Angenent, L.T. (2013). Biocatalytic reduction of
short‐chain carboxylic acids into their corresponding alcohols with syngas fermentation.
Biotechnology and Bioengineering 110: 1066–1077.
Picataggio, S., Rohrer, T., Deanda, K. et al. (1992). Metabolic engineering of Candida tropicalis
for the production of long chain dicarboxylic acids. Biotechnology 10: 894–898.
Praveen, K., Mounika, B., and Sarvamangala, D. (2017). Production of aspartic acid‐a short
review. International Journal of Engineering Trends and Technology 45 (6): 254–257.
Raemakers‐Franken, P.C., Schürmann, M., Trefzer, A.C., and de Wildman, S.M.A. (2010).
Preparation of adipic acid. World Patent: WO2010104391.
Rajesh, R.O., Pandey, A., and Binod, P. (2018). Bioprocesses for the production of 2,5‐
furandicarboxylic acid. In: Biosynthetic Technology and Environmental Challenges: Energy,
Environment, and Sustainability (eds. S. Varjani, B. Parameswaran, S. Kumar and S. Khare).
Singapore: Springer.
Ramos, L.P. (2003). The chemistry involved in the steam treatment of lignocellulosic materials.
Química Nova 26: 863–871.
Rathnasingh, C., Raj, S.M., Lee, Y. et al. (2012). Production of 3‐hydroxypropionic acid via
malonyl‐CoA pathway using recombinant Escherichia coli strains. Journal of Biotechnology
157 (4): 633–640.
Rinaldi, R. and Schuth, F. (2009). Acid hydrolysis of cellulose as the entry point into
biorefinery schemes. ChemSusChem 2: 1096–1107.
Roa Engel, C.A., Straathof, A.J., Zijlmans, T.W. et al. (2008). Fumaric acid production by
fermentation. Applied Microbiology and Biotechnology 78 (3): 379–389.
3 Biotech 5 (4): 337–353.
Saska, M. and Martin, C. (2006). Production of fuel ethanol from sugarcane bagasse and
sugarcane trash. Proceedings of IX Congress on Sugar and Sugar Cane Derivatives, Havana,
Cuba (19–22 June).
Sato, K., Aoki, M., and Noyori, R.A. (1998). Green route to adipic acid: direct oxidation of
cyclohexenes with 30 percent hydrogen peroxide. Science 281: 1646–1647.
Shen, Y., Brown, R., and Wen, Z. (2014). Enhancing mass transfer and ethanol production
insyngas fermentation of Clostridium carboxidivorans P7 through a monolithic biofilm
reactor. Applied Energy 136: 68–76.
Shokrkar, H., Ebrahimi, S., and Mehdi Zamani, M. (2018). A review of bioreactor technology
used for enzymatic hydrolysis of cellulosic materials. Cellulose 25 (1): 6279–6304.
Shuguo, W., Zehui, Z., and Bing, L. (2015). Catalytic conversion of fructose and 5‐
hydroxymethylfurfural into 2,5‐furandicarboxylic acid over a recyclable Fe3O4‐CoOx
magnetite nanocatalyst. ACS Sustainable Chemistry & Engineering 3 (3): 406–412.
Shyamkumar, R., Moorthy, I.M.G., Ponmurugan, K., and Baskaret, R. (2014). Production of L‐
glutamic acid with Corynebacterium glutamicum (NCIM 2168) and Pseudomonas reptilivora
(NCIM 2598): a study on immobilization and reusability. Avicenna Journal of Medical
Sim, J.H., Kamaruddin, A.H., Long, W.S., and Najafpour, G. (2007). Clostridium aceticum‐a
potential organism in catalyzing carbon monoxide to acetic acid: application of response
surface methodology. Enzyme and Microbial Technology 40 (5): 1234–1243.
Singh, O.V. (2008). Modulated gluconic acid production from immobilized cells of Aspergillus
niger ORS‐4.410 utilizing grape must. Journal of Chemical Technology & Biotechnology 83:
780–787.
Singh, O.V. and Singh, R.P. (2006). Bioconversion of grape must into modulated gluconic acid
production by Aspergillus niger ORS‐ 4.410. Journal of Applied Microbiology 100: 1114–1122.
Song, C.W., Kim, J.W., Cho, I.J., and Lee, S.Y. (2016). Metabolic engineering of Escherichia coli
for the production of 3‐hydroxypropionic acid and malonic acid through β‐alanine route.
ACS Synthetic Biology 5: 1256–1263.
Su, J., Shen, F., Qiu, M., and Qi, X. (2017). High‐yield production of Levulinic acid from
pretreated cow dung in dilute acid aqueous solution. Molecules 22 (2): 285.
Taherzadeh, M.J. and Karimi, K. (2008). Pretreatment of lignocellulosic wastes to improve
ethanol and biogas production: a review. International Journal of Molecular Sciences 9 (9):
1621–1651.
Tajima, T., Hamada, M., Nakashimada, Y., and Kato, J. (2015). Efficient aspartic acid
production by a psychrophile based simple biocatalyst. Journal of Industrial Microbiology
Takano, J. and Kino, K. (1999). Process for producing aspartase and process for producing L‐
aspartic acid. US Patent US5916782A, 29 June 1999.
Thakker, C., Bennett, G.N., and San, K.Y. (2013). Production of succinic acid by engineered E.
coli strains using soybean carbohydrates as feedstock under aerobic fermentation conditions.
Thakker, C., Martinez, I., San, K.Y., and Bennett, G.N. (2012). Succinate production in
Escherichia coli. Biotechnology Journal 7: 213–224.
Tsuge, Y., Kawaguchi, H., Sasaki, K., and Kondo, A. (2016). Engineering cell factories for
producing building block chemicals for bio‐polymer synthesis. Microbial Cell Factories 15
(19) https://doi.org/10.1186/s12934‐016‐0411‐0.
Van de Vyver, S. and Román‐Leshkov, Y. (2013). Emerging catalytic processes for the
production of adipic acid. Catalysis Science & Technology 3 (6): 1465–1479.
Vidra, A. and Németh, A. (2018). Bio‐based 3‐hydroxypropionic acid: a review. Periodica
Polytechnica, Chemical Engineering 62 (2): 156–166.
Vikramachakravarthi, D., Kumar, R., and Pa, P. (2014). Production of L (+) glutamic acid in a
fully membrane‐integrated hybrid reactor system: direct and continuous production under
non‐neutralizing conditions. Industrial & Engineering Chemistry Research 53 (49):
19019–19027.
Visioli, L.J., Enzweiler, H., Kuhn, R.C. et al. (2014). Recent advances on biobutanol production.
Sustainable Chemical Processes 2 (15) https://doi.org/10.1186/2043‐7129‐2‐15.
Werpy, T., Petersen, G., Aden, A. et al. (2004). Top value added chemicals from biomass volume
I‐results of screening for potential candidates from sugars and synthesis gas top value added
chemicals from biomass volume I: results of screening for potential candidates. US
Department of Energy Efficiency and Renewable Energy.
Wi, S., Chung, B., Lee, Y. et al. (2011). Enhanced enzymatic hydrolysis of rapeseed straw by
popping pretreatment for bioethanol production. Bioresource Technology 102: 5788–5793.
Wierckx, N., Schuurman, T., Kuijper, S.M., and Ruijssenaars, H.J. (2012). Genetically modified
cell and process for use of said cell. European Patent: EP 2 683 815 B1.
Wierckx, N., Schuurman, T.D., Blank, L.M., and Ruijssenaars, H.J. (2015). Whole‐cell
biocatalytic production of 2,5‐furandicarboxylic acid. In: Microorganisms in Biorefineries (ed.
B. Kamm), 207–223. Berlin, Germany: Springer.
Xi, Y.L., Dai, W.Y., Xu, R. et al. (2013). Ultrasonic pretreatment and acid hydrolysis of
sugarcane bagasse for succinic acid production using Actinobacillus succinogenes. Bioprocess
and Biosystems Engineering 36 (11): 1779–1785.
Xu, Q., Li, S., Fu, Y.Q. et al. (2010). Two‐stage utilization of corn straw by Rhizopus oryzae for
fumaric acid production. Bioresource Technology 101: 6262–6264.
Yan, D., Wang, C., Zhou, J. et al. (2014). Construction of reductive pathway in Saccharomyces
cerevisiae for effective succinic acid fermentation at low pH value. Bioresource Technology
156: 232–239.
Yang, C.F. and Huang, C.R. (2016). Biotransformation of 5‐hydroxy‐methylfurfural into 2,5‐
furan‐dicarboxylic acid by bacterial isolate using thermal acid algal hydrolysate. Bioresource
Yelamanchi, S., Jayaram, S., Thomas, J.K. et al. (2016). A pathway map of glutamate
metabolism. Journal of Cell Communication and Signaling 10 (1): 69–75.
Yu, H., Peng, F., Tan, J. et al. (2011). Selective catalysis of the aerobic oxidation of cyclohexane
in the liquid phase by carbon nanotubes. Angewandte Chemie International Edition 50 (17):
3978–3982.
Yu, K.M.K., Hummeida, R., Abutaki, A., and Tsang, S.C. (2006). One‐step catalytic
cyclohexane oxidation to adipic acid using molecular oxygen. Catalysis Letters 111 (1–2):
51–55.
Zanghellini, A.L. (2017). Fermentation route for the production of levulinic acid, levulinate
esters and valerolactone and derivatives therof. US Patent 2017/0275657 A1, 28 September
2017.
Zelle, R.M., de Hulster, E., wan Widen, W.A. et al. (2008). Malic acid production by
Saccharomyces cerevisiae: engineering of pyruvate carboxylation, oxaloacetate reduction,
and malate export. Applied and Environmental Microbiology 74: 2766–2777.
Zhang, H., Zhang, J., and Bao, J. (2016). High titer gluconic acid fermentation by Aspergillus
niger from dry dilute acid pretreated corn Stover without detoxification. Bioresource
Zhang, K. (2012). Fumaric acid fermentation by Rhizopus oryzae with integrated separation
technologies. PhD thesis. Ohio State University, Columbus, OH, USA.
Zhang, Z., Jin, B., and Kelly, J. (2007). Production of lactic acid and by‐products from waste
potato starch by Rhizopus arrhizus: role of nitrogen sources. World Journal of Microbiological
Zou, X., Zhou, Y., and Yang, S.T. (2013). Production of polymalic and malic acid by
Aureobasidium pullulans fermentation and acid hydrolysis. Biotechnology and
Bioengineering 110: 2105–2113.
247
11
Valorization of Lignin Into Value-Added Chemicals

and Materials
Ruly Teran Hilares1, Lucas Ramos1, Muhammad Ajaz Ahmed2, Avinash P.
Ingle1, Anuj Kumar Chandel1, Silvio Silvério da Silva1, Jeon Woon Choi2, and
Julio Cesar dos Santos1
1
2
Graduate School of International Agricultural Technology, Institute of Green-Bio Science and Technology, Seoul National
University, Pyeongchang, Republic of Korea
11.1 Introduction
Interest in the use of bio‐based resources is continuously increasing due to the limited
availability of fossil fuels and the price of a barrel of oil. Resources of fossil origin are run-
ning out and there are problems related to the political instability of oil‐producing coun-
tries. In addition, global dependency on fossil resources leads to environmental concerns
such as global warming and resulting climate change (Cherubini 2010; Boudet 2011;
Ahorsu et al. 2018).
A vital long‐term alternative is to develop products derived from renewable natural
resources. This solution is environmentally friendly and a great indicator of sustainable
development. In fact, several governments have recently approved legislation on the expan-
sion of gross domestic energy and chemical production from renewable resources, espe-
cially biomass (Ahorsu et al. 2018; Ramesh et al. 2019). As one of the most abundant
renewable carbon sources on earth, lignocellulosic biomass attracts most interest.
The chemical composition of lignocellulosic materials consists mainly of cellulose
(semicrystalline homopolymer), hemicellulose (group of totally amorphous heteropoly-
mers), and lignin (a polyphenolic macromolecule). Among these, lignin is responsible for
the mechanical resistance of plants and is intertwined between the cellulose and hemicel-
lulose molecules, giving rigidity to the plant (Tobimatsu and Schuetz 2019).
Lignin has the most complex structure and its utilization for value‐added chemicals is a
severe challenge compared to the use of cellulose and hemicelluloses (Shen et al. 2017).
Even so, great attention is given to lignin because it is the only abundant renewable natural
aromatic macromolecule, which can be used as a sustainable feedstock to produce aro-
matic chemicals. Indeed, researchers are recognizing the importance of lignin and hence

studies on lignin conversion into platform chemicals and alternative fuels have proliferated
in the last decade.
There are several countries which are rich in different lignocellulosic materials. For
example, Brazil has a lot of agro‐industrial activities that produce great quantities of agri-
cultural residues. Sugarcane is abundant in Brazil and it is considered as a raw material
with high potential for full use of all its constituents in industrial processes. For historical
and cultural reasons, Brazil is the world’s largest producer of sugarcane, which is mainly
destined for the sugar and alcohol industries. In this sector, the juice extracted from the
sugarcane is used to obtain sugar and ethanol, and sugarcane bagasse is used by the indus-
try to generate electricity and steam. Even after this use, there is an excess of bagasse and
thus the use of this by‐product in different processes has become the target of many
research studies.
In recent years, efforts have been made to develop viable multiproduct integrated indus-
trial facilities for the processing of lignocellulosic materials, commonly called “biorefiner-
ies.” They use a range of production techniques and facilities for the processing of biological
and renewable raw materials, operating in a fully integrated manner. Through chemical,
enzymatic or fermentative physical processes, biorefineries propose to transform these raw
materials into products that meet the needs of modern society, in a sustainable way and
with minimum environmental impact (NOVACANA 2013).
The lignin fraction in lignocellulosic biomass like sugarcane bagasse is about 20–25%
(Sporck et al. 2017) and has been used mainly in the production of energy due to its high
calorific value. However, there are several reports demonstrating other applications for this
fraction which can contribute to the economic feasibility of biorefinery with the generation
of different products of interest (Aro and Fatehi 2017). Lignin can be used as a green alter-
native in many petroleum‐derived substances such as fuels, resins, rubber additives, ther-
moplastic blends, nutra‐ and pharmaceuticals. It can be used in many fields due to its
dispersing, binding, complexing, and emulsion‐stabilizing properties.
One of the most recent applications of lignin, reported by Lee et al. (2019), is its use as a
natural UV adsorption agent. In that study, lignin obtained from Miscanthus sacchariflorus
via mild organosolv conditions was applied in the formulation of sunscreens and cosmetics
products in liquid and cream formulations, resulting in high absorption of UV light.
The main applications of lignin are divided into three groups.
●● Fuel (power): to produce energy.
●● Macromolecules: polymers, carbon fibers, and binders.
●● Aromatics: phenols, vanillin, and polymer building blocks.
The diversity and complexity of the lignin molecule are a great challenge and the possi-
ble applications of lignin are dependent on the way it is obtained after biomass pretreat-
ment. The type of process used to produce lignin will determine the value of the products
that can be derived from it. Although efforts have been put into lignin valorization and
considerable progress has been made in this field, fully unlocking lignin’s potential for the
economical production of high value‐added products faces multiple challenges.
The aim of this chapter is to discuss the potential of lignin as an important source to
obtain various interesting products. In addition, aspects like the chemistry of lignin and
various processes for its extraction are discussed.
Valorization of Lignin Into Value-Added Chemicals and Materials 249
11.2 Composition
Lignin is a high molecular weight (MW) macromolecule that presents ring‐shaped struc-
tures in a complex network composed of polyphenols. It is constituted of three main units,
namely monolignols linked together with an array of different intrinsic bindings. These
units, guaiacyl (G), syringyl (S), and p‐hydroxyphenyl (H), are the most common ones and
thus participate in lignin construction in most plants (Fengel and Wegener 1984; Calvo‐
Flores and Dobado 2010). The quantity as well as the proportions of units in lignin vary
depending on different plant species (Table 11.1). Usually, lignin found in conifer types of
wood has more G units than the other two monolignols; broad‐leaf wood only has G and
S units; and lignin found in grass, interestingly, has a mixture of the three monolignols
(Mei et al. 2019).
Because of the composition and abundance, lignin can be classified into three main cat-
egories: G‐lignin, G‐S‐lignin and G‐S‐H‐lignin. These basic building blocks of lignin are
further connected to a series of bonds of C─O and C─C linkages, these more specifically
including the bond types of β‐O‐4 (β‐aryl ether), 4‐O‐5 (diaryl ether), α‐O‐4 (α‐aryl ether),
β‐5 (phenylcoumaran), β‐β (resinol), β‐1 (spirodienone) or 5‐5, and thus form a more com-
plicated macromolecule (Zakzeski et al. 2010). Figure 11.1 depicts the basic structures of
lignin and its various linkages.
One of the most significant obstacles in utilization of lignin for high‐value products is its
structural heterogeneity, which varies depending on the biomass source, type, and even
some growing parameters. Breakdown of lignin macromolecules into their main mono-
meric constituents provides an opportunity to generate products with manageable and
consistent quality. However, selective depolymerization of lignin to valuable compounds
has been a well‐recognized challenge (Beson et al. 2014).
Although it has been investigated for decades, the native lignin structure is still uncer-
tain; several works have been directed to the development of lignin isolation protocols.
Lignin isolation and purification methods usually produce structural modification of the
lignin macromolecular matrix, to varying degrees, and this obscures the lignin native
structure. Aiming to facilitate subsequent conversion, the ideal lignin isolation proce-
dure requires low temperature, high yield, and less condensation of lignin structures
(Li et al. 2019).
Table 11.1 Typical composition of lignin monomers in different types of lignocellulosic species.
p-Coumaryl alcohol Coniferyl Sinapyl alcohol

Monolignol (H) (%) alcohol (G) (%) (S) (%) References
Grass 10–25 25–50 25–50 Li et al. (2019)

Conifer wood 0.5–3.4 90–95 0–1 Li et al. (2019)
Broad‐leaf wood Trace 25–50 50–75 Li et al. (2019)
Sugarcane bagasse 2 38 60 del Río et al. (2015)
Sugarcane straw 4 68 28 del Río et al. (2015)
p-Coumaryl alcohol
fragment
OH
OH O HO
HO O O
O O O
OH HO OH
HO OH O O OH
HO O OH
O HO OH HO O HO
O OH HO O
HO O O p-Coumaryl alcohol
HO Grass Lignin
O O HO HO
HO OH
HO OH O O OH
O OH O
O OH O
O O O HO
HO O O
O O O HO
OH O HO O O Coniferyl alcohol
OH OH
O OH
Coniferyl alcohol Softwood Lignin OHOH HO
O fragment O OH O
O O OH
OH OH O
OH O OH
OH HO
O O O
O O OH O
O OH O O Sinapyl alcohol
O HO O OH
O Sinapyl alcohol O
OH O O
fragment
OH O OH
Hardwood Lignin
O
R
R
HO R
HO R O OH R
HO O O O
O HO HO O HO R
OH O O
R R
O
R O
R R R R R R R O
R R R
O O O R R O R
O
β-O-4 α-O-4 4-O-5 β-5 β-β β-1 5–5
Figure 11.1 Exhibition of lignin along with its intersubunit linkages.

11.3 Lignin Extraction
The branched structure of lignin contains various functional groups (carbonyl, carboxyl, and
even methoxy) and some common linkages including α‐O‐4, 4‐O‐5, β‐O‐4, etc. (Hatakeyama
and Hatakeyama 2009). This lignocellulosic fraction is connected with other major sugars of
cellulose and hemicellulose and, collectively, this linkage is called a lignin‐carbohydrate com-
plex (LCC) (Yuan et al. 2011). Lignin can be extracted from biomass by numerous methods,
but its chemical structure is modified in the process. The classic technique is that used in the
pulp and paper industry but other technologies, specifically in the context of pretreatment of
biomass for biorefineries, have been developed and are discussed below.
11.3.1 Kraft Processing for Lignin Isolation

This is a typical and widely (>95%) adopted strategy in the wood market, more precisely in
the pulp and paper industry (da Silva et al. 2009). It employs the combination of sodium
sulfide (Na2S) and sodium hydroxide (NaOH) followed by thermal treatment up to 180 °C
for a couple of hours of process time (van Heiningen 2006). A black liquor, the spent prod-
uct of the process containing dissolved lignin as well as hemicellulose, is then obtained.
Being a high‐energy density material, lignin is usually burnt off after evaporation to ~50%
solid contents to fulfill the energy demands. The surplus extraction reactants can also be
recovered and lignin can be extracted from this solution (alkaline solution) for other appli-
cations via a lignoboost process.
This process takes advantage of the solubility of lignin and uses a mineral acid to lower
pH and consequently precipitate out the lignin from the black liquor (Zhu et al. 2009). The
lignin from the kraft process has also been recovered by ultrafiltration, as reported by
Jönsson and Wallberg (2009). In that report, ceramic membrane was used for recovery of
lignin from the solution without altering the pH or temperature of the liquor, resulting in
190 g/L and 50 g/L of lignin and hemicellulose in the ultrafiltration retentate, respectively.
11.3.2 Sulfite Process

This process was employed for the first time in 1874, and is considered the oldest but most
promising process for lignin extraction (Zhang et al. 2013). Typically, it produces dissolved
lignin, called lignosulfonate, up to 1 million tons per year solely based on this method. This
method uses sulfite bases such as sodium, calcium, and/or ammonium or magnesium salts
along with the aqueous sulfur dioxide (SO2). During the process, sulfur (largely as SO3
group) is linked with lignin to generate lignosulfonate, mainly with cyclic phenylpropene
units. Thus, this process contains more sulfur with lignin compared to the previously dis-
cussed method (Calvo‐Flores and Dobado 2010).
11.3.3 Application of Soda-Anthraquinone for Lignin Extraction

This process was developed even before the kraft process – it was commercialized in 1853
(Jiménez et al. 2009). It is now rarely employed and is more precisely used for nonwoody
biomasses. It typically includes the treatment of lignocelluloses at less than 165 °C with a
high‐pressure alkali (NaOH) solution. Moreover, further addition of a small amount of

anthraquinone (0.1%) aids the delignification process and preserves the carbohydrate frac-
tion in the material. The main advantage of using this method is the production of pure
lignin (Ghatak 2008).
Recently, this method was used for lignin extraction from paulownia (Zamudio et al.
2015). In that study, delignification of paulownia with soda–anthraquinone extraction
prior to autohydrolysis resulted in higher performance than an alkaline process without
prior autohydrolysis. Additionally, the resulting lignin showed high calorific value, at
14 822 kJ/kg, and when it was used for polylacticacid–lignin composites production, the
elastic (storage) modulus (E’) property improved by increasing the lignin content from
12.5% to 30% in the composite.
11.3.4 Extraction of Lignin Via Acid Treatment

This is actually a lignin extraction cum pretreatment method mainly used for the fractiona-
tion of biomass matrix and hydrolysis of cellulose sugar polymers. The process can be
employed with either mild acid under prolonged thermal conditions or with concentrated
acid (>30%) for a limited period of time (Lloyd and Wyman 2005). The lignin is obtained in
the form of a by‐product through this process.
This method, with its holistic approach, is quite widely adopted, being more specific for
cellulose hydrolysis and leaving lignin in the residual biomass (Miller and Hester 2007).
Unfortunately, it still has some economic constraints due to the high cost required for the
construction of corrosion‐resistant reactors as well as for downstream neutralization. A
wide range of acids have been utilized for biomass feedstock treatment and thus for lignin
separation (Zhao and Liu 2007). These include both organic and inorganic acids such as
phosphoric acid, hydrochloric acid, sulfuric acid, formic acid, nitric acid, and acetic acid.
The mode of action of these acids includes creation of slits in the raw biomass and hydroly-
sis of the amorphous and even crystalline portion of biomass (depending on acid severity),
leaving the lignin in the biomass as a residue. Of all the aforementioned acids, sulfuric acid
is the nearest to commercialization (Cao et al. 2009).
11.3.5 Organosolv Process for Lignin Extraction

This process employs organic solvents and is classically used in the pulp and paper indus-
try (Zhao et al. 2009). It uses solvents (the most frequent are acetone, ethanol, methanol,
and even a dilute mixture of any of them) with a temperature constant of 100–250 °C. Its
typical mechanism results in soluble forms of hemicellulose and lignin fractions (Zhang
et al. 2016). This process can be further intensified by the addition of acid catalysts, either
organic and/or mineral, with mild thermal conditions to further promote the breakage of
intermolecular bonds, resulting in a more complicated lignin structure (similar to the
native one) (Sannigrahi et al. 2010). The solvents usually employed in this process can be
separated and recovered based on boiling point differences whereas the lignin is precipi-
tated out by acids. Some of the widely adopted commercial processes in this regard include
the Milox method (with peroxyformic acid), the Alcell process (employing ethanol), and
the application of methanol called the Organocell process (Sridach 2010).
11.3.6 Lignin Extraction Via Ammonia Fiber Explosion (AFEX)

This is an alkaline pretreatment process involving exposure of biomass to ammonia at a
pressure as high as 40 bar and temperature ranging from 90 to 180 °C (Gollapalli et al.
2002). This process lasts only for a short period of time and is associated with the sudden
compression and expansion (pressure cycles) of ammonia (operational pressure 7–40 bar)
(Tolbert et al. 2014). This pretreatment releases the hemicelluloses and promotes decrystal-
lization of cellulose along with the release of a small amount of lignin and offers the pos-
sibility of high ammonia recovery rates. This method can be used for hardwoods, whereas
for softwoods its efficiency is reduced to some extent (Zhang et al. 2011).
11.3.7 Steam Explosion for Lignin Extraction

This method was pioneered by Mason in 1924 when it was employed for the production of
chipboard panels. Generally, this method uses steam at high pressure to break down the
biomass to release the acetyl content present in the hemicellulose chain (Li et al. 2007).
This then forms acetic acid to hydrolyze the glycosidic bonds of the biomass. Usually, the
process is completed when alkaline delignification is employed for the complete solubiliza-
tion of lignin (Cherian et al. 2010).
11.3.8 Other Methods for Lignin Extraction

Other techniques for lignin extraction include the dioxane‐based extraction process
(Rencoret et al. 2015; Chen et al. 2019) and ionic liquids (IL) (Achinivu et al. 2014, 2018).
In the work of Rencoret et al. (2015), lignin was extracted from brewer’s spent grain (BSG)
using the dioxane‐based extraction process (14 g of extractive free material, 150 mL of
0.05 M HCl in 82:18 v/v of dioxane:water), resulting in 50% klason lignin extraction yield.
The obtained lignin had 32% p‐hydroxyphenyl, 50% guaiacyl and 18% syringyl aromatic
units, with presence of β‐O‐4′ alkyl‐aryl ethers (77–79% of all interunit linkages), followed
by β‐5′ phenylcoumarans (11–13%). Dioxane‐based lignin extraction also can be performed
in an alkaline medium, as recently published by Chen et al. (2019). In that report, 29.5% of
lignin extraction yield was achieved with 80% alkaline dioxane (0.05 M NaOH) with 1:20
(g/mL) solid/liquid ratio, 80 °C and six hours of processing.
The IL are also known as green solvents and are composed of various cations (usually the
organic part) and anions (inorganic part) with low melting points, sometimes even in nega-
tive degrees Celsius. Among the different ILs, the protic ionic liquids (PIL) have been used
efficiently for lignin extraction from lignocellulosic biomass. This kind of IL can be easily
synthetized using relatively cost‐competitive chemicals (Achinivu et al. 2018). In the work
of Achinivu et al. (2018), ionic pyrrolidinium acetate ([Pyrr][Ac]) was used for selective
extraction of lignin from corn stover, resulting in lignin with a low average molecular
weight (Mw 900 g/mol) compared to kraft lignin (Mw 6500 g/mol), with better dispersity.
Additionally, extensive information about IL for solubilization of lignocellulosic biomass
can be found in the review published by Zhu et al. (2018).
The lignin obtained using the above‐mentioned processes are used for production of
several high‐value products, as discussed in the following section.
11.4 Valorization of Lignin in High-Value Products
The application of lignin in the production of different materials is envisaged as one accept-
able strategy for a sustainable world (de Wild et al. 2012). The potential uses of lignin are
illustrated in Figure 11.2. For example, lignin from the pulp and paper industry has been
used for the production of lignosulfonates, value‐added materials due to their capacity to
retain water. Lignosulfonates can be used for concrete mixtures, as surfactants in dyes, pig-
ments, coatings, lubricants, in detergent formulation, as animal feed, and in the formula-
tion of urea and formaldehyde (Stewart 2008).
11.4.1 Biocomposites
The concept of sustainability is always associated with the application of renewable raw
materials for a biodegradable end‐product (Ragauskas et al. 2006). For the production of
sustainable materials, such as bioplastics, blends, or biocomposites, lignin has been used as
a binding matrix for natural fibers. A well‐known material produced in this way is
Arborform, which is composed of a matrix based on lignin (~60%) and flax (natural fibers).
It has been made via a typical fossil‐based thermoplastic polymer manufacturing technique
at the German Fraunhofer‐Institut für Chemische Technologie (Wegener et al. 2006).
Lignin has also been used in the formulation of bioplastics or composites (Spiridon et al.
2015; Dorrstein et al. 2018). Spiridon et al. (2015) used lignin obtained from softwood and
hardwood for the manufacturing of polylactic acid (PLA)‐based composite. The resulting
materials increase the strength and thermal stability of PLA and, by using softwood lignin,
the water sorption capacity was increased. Lignin obtained by organosolv of a grass (Poacae)
silage press cake (PC) batch was also used in a bioplastic (lignin/polyethylene‐co‐vinyl
acetate ‐EVA) composite (Dorrstein et al. 2018). In that study, an increase of dynamic stiff-
ness Cdyn from φL = 0.59 (Cdyn ≈ 350 N/mm) to φL = 0.71 (Cdyn ≈ 550 N/mm) was observed
when using high lignin content (75 wt%).
Biocomposites
Aerogels Filler for

Resins Lignin materials
Thermosetting
Carbon fiber polymers
Activated carbon
Chemicals
Figure 11.2 Some products that can be produced from lignin.

Thermoplastic blends and composite materials are improved by the addition of lignin
(Akato et al. 2015). Microbial characteristics of lignin are of particular interest for the produc-
tion and preparation of packing materials for foodstuff (Akkus et al. 2014). For example, lignin
nanoparticles (LNP) and chitosan (CH) were used as filler in polyvinyl alcohol‐chitosan (PVA‐
CH), as reported by Yang et al. (2016). In that study, the mechanical (tensile strength and
Young’s modulus) and thermal properties of PVA‐CH blend were enhanced using LNP (3%).
Additionally, in binary (PVA/3LNP and CH/3LNP films) and ternary nanocomposite systems
(PVA/CH/3LNP blend), a remarkable antibacterial activity was observed on plant pathogenic
bacteria Pectobacterium carotovorum subsp. odoriferum (Pco) (CFBP 1115).
Recently, blow‐molded containers made of high‐density polyethylene containing 5 wt%
of 1:5 ratio ZnO/lignin (dual fillers) were prepared with a single‐screw extruder
(Klapiszewski et al. 2019). The filler obtained using a 1:5 ratio of ZnO/lignin showed higher
antimicrobial activity on Staphylococcus and Bacillus compared to a positive control (anti-
biotic tetracycline). Besides, a significant twofold increase in the compression force for
blends with a ZnO:lignin ratio of 1:5 was reported in the blow‐molded container.
11.4.2 Production of Carbon Fibers and Activated Carbon

One of the major applications of lignin is the synthesis of engineering materials as carbon
fibers which have received much attention in the last 50 years due to their improved mechan-
ical properties and light weight (Kadla et al. 2002). Carbon fiber has been produced largely
from polyacrylonitrile (PAN) which constitutes almost half of the total production cost
(Baker and Rials 2013). As an alternative, lignin‐based carbon fibers (LCFs) are expected to
be affordable (saving possibly up to 49%) (Maradur et al. 2012). However, lignin is quite brit-
tle which will require modification to enhance its spinning and stretching properties.
Typical carbon fiber production methods include modification (upon addition of chemi-
cals), mixing (with other polymeric compounds), and thermal processing (Zhang and
Ogale 2014). Kraft lignin has been successfully spun through a thermal treatment followed
by carbonization without applying any chemical treatment. Researchers at the University
of Iowa adapted a quick method for the production of carbon fiber using a lignin precursor
via a pyrolization process (Sudo and Shimizu 1994). Typically, for the conversion of lignin
into carbon fibers, it is first converted into fibers using a solvent‐swollen gel followed by air
oxidation and pyrolysis in an inert environment.
An important role of lignin in the production of electrodes for batteries was reported due
to it low cost and high pyrolysis yields (Jin et al. 2014). For example, LCFs for renewable
and multifunctional lithium ion battery (LIB) electrodes were reported by Nowak et al.
(2018). In that report, LCF was synthesized using softwood kraft lignin via oxidative ther-
mostabilization of pure melt‐spun lignin and carbonization at 1000 °C. The obtained mate-
rial had a strength of 628 MPa and stiffness of 37 GPa and when it was used for the negative
electrodes in a LIB, it showed capacity of 335 mAh/g.
Another interesting lignin application, activated carbon is extensively used as a commer-
cial absorbent. The use of lignin precipitated from kraft black liquors for preparation of
activated carbons and their use as adsorbents to remove different water pollutants (phenol,
2,4,5‐trichlorophenol and Cr(VI)) was reported by Gonzalez‐Serrano et al. (2005). In other
recent work by Ibeh et al. (2019), activated carbon monoliths were produced from Alcell®,
kraft lignin, and olive stone by chemical activation with H3PO4, (ratio of 1 H3PO4/raw
material) and they were used in electrochemical applications.
11.4.3 Lignin-Based Polyurethane, Resins, and Aerogels

Lignin can replace phenol to make lignin‐based resins such as phenol formaldehyde (Wang
et al. 2009). However, this is still a burdensome cost for biorefineries which are facing a
challenge of high cost compared with the fossil‐based refineries. Another interesting appli-
cation of lignin could be in the production of lignin‐based polyurethane (PLU), replacing
the petroleum‐based raw materials. Some studies showed a significant improvement in the
production of plywood even with up to 70% addition of lignosulfonates (Alonso et al. 2004).
Lignin has a good affinity to cross‐link in the production of PLU, and can be used in
crude form (Mahmood et al. 2016). In this case, the addition of polyols, as polybutadiene
glycol, polyethylene glycol and ethylene adipate, can lead to softening of the product
(Ciobanu et al. 2004). Addition of soda lignin (5~23%) along with softening agents (poly-
ethylene glycol and polyethyleneadipate) resulted in improved mechanical properties but
with lower elasticity and decomposition temperature (Mousavioun et al. 2013). Other
materials where lignin is used correspond to biodegradable lignin‐based foams in both
rigid and flexible forms (Amaral et al. 2012). Moreover, a one‐pot process intensification
approach was proposed for the production of lignin‐based foams from lignin powders with
the addition of polyols (Campanella et al. 2009). The foams are quite rigid in nature and are
biodegradable, with improved thermal and mechanical properties (Cateto et al. 2014).
Many studies have suggested pathways for the production of monomeric products of
lignin (e.g., γ‐valerolactone). They mainly include the incorporation of vanillin and alco-
hols from the lignin for the production of epoxy resins (Sasaki et al. 2013). These mono-
meric degradation products of lignin can be obtained from the deconstruction of biomass
structure. They are released from biomass and react with epichlorohydrin to form the
epoxy functionality and then are further polymerized with aldehydes and/or amines
(Cavdar et al. 2008). The materials thus produced are of improved quality in terms of their
thermal, mechanical, and even chemical/adhesive properties.
Lignin has also been used for the production of aerogels, specifically to replace oil‐based
raw materials in the production line due to properties such as low density and quite high
surface area. Aerogels typically can be employed for the production of energy storage mate-
rials (Xu et al. 2015). Recently, studies have been done on the successful synthesis of lignin‐
modified graphene aerogels (LGAs). These materials have also been studied for their
application in membranes for oil separation (Chen et al. 2018).
Another form of lignin‐based aerogel is resorcinol formaldehyde‐based carbon aerogels
which have been successfully utilized for energy storage applications due to their nontoxic
nature, high chemical stability, and sustainable nature (Li et al. 2019).
11.5 Economic and Environmental Aspects of Lignin Use
Lignin, due to its aromatic structure and relatively high energy density, has been used for
production of many lignin‐based products in a biorefinery context. Conventionally, after its
recovery from the biorefinery waste, the lignin is merely subjected to energy production by
combustion, thus restricting its economic worth to no more than $50/dry ton on a market
competitive basis (Ragauskas et al. 2014).
Lignin could contribute much more to the bioeconomy if it was utilized for other applica-
tions beyond combustion only. In this regard, a mammoth challenge would be the produc-
tion of high‐value chemicals to replace their petroleum‐based counter parts in the
forthcoming bioeconomy market. For example, BTX (benzene, toluene and xylene) alone is
a $100 billion market with an ever‐increasing demand of 100 million tons per year with a
$1200 per ton market price (Biddy et al. 2016; Niziolek et al. 2016). In addition to BTX, phe-
nols are another attractive chemical feedstock that can be produced from lignin. The current
phenol market, on an annual production of 8 million tons, is ~$1500 per ton per annum and
is expected to expand at a 3.9% compound annual growth rate in the next decade (Biddy
et al. 2016). Phenols extracted from lignin are sustainable raw materials for the production
of value‐added materials such as composites, activated carbon and carbon fibers with utili-
zation in manufacturing, energy production, and pollutant removal (Li et al. 2015).
Carbon fiber is another key player with a worth of $4.5 billion market value and is
expected to rise in near future. The extent of its worth can be imagined when in a previous
report the United States Department of Energy mentioned that almost half of the indige-
nous passenger vehicle steel could potentially be replaced by utilizing only 10% of the
lignin available in the country.
Life cycle analysis (LCA) is a technique that compiles and evaluates the inputs (materials
and energy) and outputs (pollutants, emission, product and by‐product) of a product in
order to estimate and finally measure its potential impact on the environment (Manandhar
and Shah 2017). Investigations on the potential environmental impact (PEI) of lignin‐based
products of four lignin extraction processes – kraft, organosolv, soda, and sulfite pro-
cesses – were performed. Among these processes, organosolv was found to have the highest
total PEI per kg of lignin with a value of 0.25 PEI per kg, followed by the kraft process with
approximately 0.09 PEI per kg of product (Wang et al. 2007). The soda process had the low-
est pollution potential with a PEI value of 0.02 per kg compared with the sulfite process
that had a value of 0.03 PEI per kg of lignin. Moreover, lignin extraction via different strate-
gies considers various economic factors as well.
Beyond the costs associated with lignin‐based products and their environmental impacts,
the choice of extraction method is also strongly linked to the value addition of extracted
lignin (Benali et al. 2013).
11.6 Conclusion
Lignin is the one of the most abundant macromolecular fractions of lignocellulosic biomass
feedstocks; it has a rigid structure due to the presence of various intermolecular linkages in
the cell wall matrix. Pretreatment of lignocellulosic material is essential to release it from
the complex structure of biomass. Among the various pretreatment approaches, organosolv
methods allow the recovery of high‐quality lignin but incur high costs. Kraft and sulfite
approaches are considered cost‐effective but the lignin thus obtained contains impurities. In
contrast, soda extraction is considered an economically viable approach for the production
of pure lignin with a very low environmental impact. Recently, other sustainable processes
involving the use of IL have been proposed for lignin extraction with better properties.
However, their technical and economic viability in large‐scale processes is still uncertain.
The valorization of lignin in high‐value products is a necessity for the economic viability
of lignocellulosic biorefineries. The potential of this lignocellulosic fraction is great, with
different possibilities of use beyond burning for energy generation. For example, lignosul-
fonates can be obtained from lignin and used for concrete mixtures, as surfactants in dyes,
pigments, coatings, lubricants, in detergent formulation, as animal feed, and in the formu-
lation of urea and formaldehyde. Biocomposites can be produced using lignin to modify
the mechanical and thermal properties of materials in a desirable way, including the use of
nanolignin as a filler. Other interesting applications include the use of lignin to produce
carbon fiber for use as electrodes for batteries. Active carbon can be produced from lignin
and used to remove pollutants from effluents and for electrochemical applications. Other
alternatives for valorization of this biomass polyphenolic fraction include its use for polyu-
rethane, resins, and aerogel production. Biodegradable foams with different rigidity prop-
erties and aerogels for energy storage can be obtained.
The economic and environmental benefits of different uses of lignin have been recog-
nized and in the near future, the potential of this raw material will be fully exploited to aid
the transition of society from a fossil‐based economy to an environmentally friendly
bioeconomy.
Acknowledgment
The authors gratefully acknowledge the FAPESP (grant numbers #2017/11086‐4,

#2016/23758‐4, #2016/10636‐8 and #2016/22086-2) for financial support.
References
Achinivu, E.C. (2018). Protic ionic liquids for lignin extraction: a lignin characterization study.
International Journal of Molecular Sciences 19 (2): 428.
Achinivu, E.C., Howard, R.M., Li, G. et al. (2014). Lignin extraction from biomass with protic
ionic liquids. Green Chemistry 16: 1114–1119.
Ahorsu, R., Medina, F., and Constantí, M. (2018). Significance and challenges of biomass as a
suitable feedstock for bioenergy and biochemical production: a review. Energies 11: 3366.
Akato, K., Tran, C.D., Chen, J., and Naskar, A.K. (2015). Poly (ethylene oxide)‐assisted
macromolecular self‐assembly of lignin in ABS matrix for sustainable composite
applications. ACS Sustainable Chemistry & Engineering 3: 3070–3076.
Akkus, M., Bahcegul, E., Ozkan, N., and Bakir, U. (2014). Post‐extrusion heat‐treatment as a
facile method to enhance the mechanical properties of extruded xylan‐based polymeric
materials. RSC Advances 4: 62295–62300.
Alonso, M.V., Oliet, M., Rodriguez, F. et al. (2004). Use of a methylolated softwood ammonium
lignosulfonate as partial substitute of phenol in resol resins manufacture. Journal of Applied
and Polymer Sciences 94: 643–650.
Amaral, J.S., Sepúlveda, M., Cateto, C.A. et al. (2012). Fungal degradation of lignin‐based rigid
polyurethane foams. Polymer Degradation and Stability 97 (10): 2069–2076.
Aro, T. and Fatehi, P. (2017). Production and application of lignosulfonates and sulfonated
lignin. Chemsuschem Reviews 9: 1861–1877.
Baker, D.A. and Rials, T.G. (2013). Recent advances in low‐cost carbon fiber manufacture from
lignin. Journal of Applied Polymer Science 130: 713–728.
Benali, M., Prin‐Levasseur, Z., Savulescu, L. et al. (2013). Implementation of lignin‐based
biorefinery into a Canadian softwood kraft pulp mill: optimal resources integration and
economic viability assessment. Biomass and Bioenergy 7: 2–11.
Beson, M., Gallezo, P., and Pinel, P. (2014). Conversion of biomass into chemical over metal
catalyst. Chemical Reviews 114: 1827–1870.
Biddy, M.J., Scarlata, C., and Kinchin, C. (2016). Chemicals from biomass: a market assessment
of bioproducts with near‐term potential. Technical Report. National Renewable Energy
Laboratory (NREL/TP‐5100‐65509). www.nrel.gov/publications.
Boudet, A.M. (2011). Editorial: a new era for lignocellulosics utilization through biotechnology.
Comptes Rendus Biologies 334: 777–780.
Calvo‐Flores, F.G. and Dobado, J.A. (2010). Lignin as renewable raw material. ChemSusChem
3: 1227–1235.
Campanella, A., Bonnaillie, L.M., and Wool, R.P. (2009). Polyurethane foams from soyoil‐based
polyols. Journal of Applied and Polymer Sciences 112: 2567–2578.
Cao, G., Ren, N., Wang, A. et al. (2009). Acid hydrolysis of corn stover for biohydrogen
production using Thermoanaerobacterium thermosaccharolyticum W16. International
Journal of Hydrogen Energy 34: 7182–7188.
Cateto, C.A., Barreiro, M.F., Ottati, C. et al. (2014). Lignin‐based rigid polyurethane foams with
improved biodegradation. Journal of Cellular Plastics 50: 81–95.
Cavdar, A.D., Kalaycioglu, H., and Hiziroglu, S. (2008). Some of the properties of oriented
trandboard manufactured using kraft lignin phenolic resin. Journal of Material Processing
Chen, C., Li, F., Zhang, Y. et al. (2018). Compressive, ultralight and fire‐resistant lignin‐
modified graphene aerogels as recyclable absorbents for oil and organic solvents. Chemical
Chen, W.J., Zhao, B.C., Cao, X.F. et al. (2019). Structural features of alkaline dioxane lignin and
residual lignin from eucalyptus grandis × E. Urophylla. Journal of Agriculture and Food
Cherian, B.M., Leão, A.L., de Souza, S.F. et al. (2010). Isolation of nanocellulose from
pineapple leaf fibres by steam explosion. Carbohydrate Polymers 81: 720–725.
Cherubini, F. (2010). The biorefinery concept: using biomass instead of oil for producing
energy and chemicals. Energy Conversion and Management 51: 1412–1421.
Ciobanu, C., Ungureanu, M., Ignat, L. et al. (2004). Properties of lignin–polyurethane films
prepared by casting method. Industrial Crops & Products 20: 231–241.
da Silva, E.A.B., Zabkova, M., Araújo, J.D. et al. (2009). An integrated process to produce
vanillin and lignin‐based polyurethanes from Kraft lignin. Chemical Engineering Research &
Design 87: 1276–1292.
De Wild, P.J., Huijgen, W.J.J., and Heeres, H.J. (2012). Pyrolysis of wheat straw‐derived
organosolv lignin. Journal of Analytical and Applied Pyrolysis 93: 95–103.
del Río, J.C., Lino, A.G., Colodette, J.L. et al. (2015). Differences in the chemical structure of
the lignins from sugarcane bagasse and straw. Biomass Bioenerginering 81: 322–338.
Dorrstein, J., Scholz, R., Schwarz, D. et al. (2018). Dataset on the structural characterization of
organosolv lignin obtained from ensiled Poaceae grass and load‐dependent molecular
weight changes during thermoplastic processing. Data Brief 17: 647–652.
Fengel, D. and Wegener, G. (1984). Wood: Chemistry, Ultrastructure, Reactions. New York:
Walter de Gruyter.
Ghatak, H.R. (2008). Spectroscopic comparison of lignin separated by electrolysis and acid
precipitation of wheat straw soda black liquor. Industrial Crops Products 28: 206–212.
Gollapalli, L.E., Dale, B.E., and Rivers, D.M. (2002). Predicting digestibility of ammonia fiber
explosion (AFEX)‐treated rice straw. Applied Biochemistry and Biotechnology 98–100: 23–35.
Gonzalez‐Serrano, E., Cordero, T., Rodriguez‐Mirasol, J. et al. (2004). Removal of water
pollutants with activated carbons prepared from H3PO4 activation of lignin from Kraft black
liquors. Water Research 38: 3043–3050.
Hatakeyama, H. and Hatakeyama, T. (2009). Lignin structure, properties, and applications. In:
Biopolymers: Lignin, Proteins, Bioactive Nanocomposites (eds. A. Abe, K. Dusek and S.
Kobayashi), 1–63. Berlin, Germany: Springer.
Ibeh, P.O., García‐Mateos, F.J., Ruiz‐Rosas, R. et al. (2019). Acid mesoporous carbon monoliths
from lignocellulosic biomass waste for methanol dehydration. Materials 12 (2394).
Jiménez, L., Serrano, L., Rodríguez, A., and Sánchez, R. (2009). Soda‐anthraquinone pulping of
palm oil empty fruit bunches and beating of the resulting pulp. Bioresource Technology 100:
1262–1267.
Jin, J., Yu, B., Shi, Z. et al. (2014). Lignin‐based electrospun carbon nanofibrous webs as
free‐standing and binder‐free electrodes for sodium ion batteries. Journal of Power Sources
272: 800–807.
Jönsson, A.S. and Wallberg, O. (2009). Cost estimates of kraft lignin recovery by ultrafiltration.
Desalination 237 (1‐3): 254–267.
Kadla, J.F., Kubo, S., Venditti, R.A. et al. (2002). Lignin‐based carbon fibers for composite fiber
applications. Carbon 40: 2913–2920.
Klapiszewski, Ł., Bula, K., Dobrowolska, A. et al. (2019). A high‐density polyethylene
container based on ZnO/lignin dual fillers with potential antimicrobial activity. Polymer
Testing 73: 51–59.
Lee, S.C., Tran, T.M.T., Choi, J.W., and Won, K. (2019). Lignin for white natural sunscreens.
Li, C., Zhao, X., Wang, A. et al. (2015). Catalytic transformation of lignin for the production of
chemicals and fuels. Chemical Reviews 115: 11559–11624.
Li, J., Henriksson, G., and Gellerstedt, G. (2007). Lignin depolymerization/repolymerization
and its critical role for delignification of aspen wood by steam explosion. Bioresource
Li, K., Zhou, M., Liang, L. et al. (2019). Ultrahigh‐surface‐area activated carbon aerogels
derived from glucose for high‐performance organic pollutants adsorption. Journal of Colloid
and Interface Science 546: 333–343.
Lloyd, T.A. and Wyman, C.E. (2005). Combined sugar yields for dilute sulfuric acid
pretreatment of corn stover followed by enzymatic hydrolysis of the remaining solids.
Mahmood, N., Yuan, Z., Schmidt, J., and Xu, C.C. (2016). Depolymerization of lignins and
their applications for the preparation of polyols and rigid polyurethane foams: a review.
Renewable and Sustainable Energy Reviews 60: 317–329.
Manandhar, A. and Shah, A. (2017). Life cycle assessment of feedstock supply systems for
cellulosic biorefineries using corn stover transported in conventional bale and densified
pellet formats. Journal of Cleaner Products 166: 601–614.
Maradur, S.P., Kim, C.H., Kim, S.Y. et al. (2012). Preparation of carbon fibers from a lignin
copolymer with polyacrylonitrile. Synthetic Metals 162: 453–459.
Mei, Q., Shen, X., Liu, H., and Han, B. (2019). Selectively transform lignin into value‐added
chemicals. Chinese Chemical Letters 30: 15–24.
Miller, S. and Hester, R. (2007). Concentrated acid conversion of pine sawdust to sugars. Part ii:
high‐temperature batch reactor kinetics of pretreated pine sawdust. Chemical Engineering
Communications 194: 103–116.
Mousavioun, P., Halley, P.J., and Doherty, W.O.S. (2013). Thermophysical properties and
rheology of PHB/lignin blends. Industrial Crops & Products 50: 270–275.
Niziolek, A.M., Onel, O., Guzman, Y.A., and Floudas, C.A. (2016). Biomass‐based production
of benzene, toluene, and xylenes via methanol: process synthesis and deterministic global
optimization. Energy & Fuels 30 (6): 4970–4998.
NOVACANA (2013). Biorrefinaria: futuro para o completo aproveitamento da biomassa de
cana. www.novacana.com/estudos/biorrefinaria‐futuro‐para‐o‐completo‐aproveitamento‐
da‐biomassa‐de‐cana‐241013.
Nowak, A.P., Hagberg, J., Leijonmarck, S. et al. (2018). Lignin‐based carbon fibers for
renewable and multifunctional lithium‐ion battery electrodes. Holzforschung 72 (2): 81–90.
Ragauskas, A.J., Beckham, G.T., Biddy, M.J. et al. (2017). Lignin valorization: improving lignin
processing in the biorefinery. Science 344 (1246843).
Ragauskas, A.J., Williams, C.K., Davison, B.H. et al. (2006). The path forward for biofuels and
biomaterials. Science 311: 484–489.
Ramesh, D., Muniraj, I.K., Thangavelu, K., and Karthikeyan, S. (2019). Chemicals and fuels
production from agro residues: a biorefinery approach. In: Sustainable Approaches for
Biofuels Production Technologies (eds. N. Srivastava, M. Srivastava, P. Mishra, et al.), 47–71.
Cham, Switzerland: Springer.
Rencoret, J., Prinsen, P., Gutierrez, A. et al. (2015). Isolation and structural characterization of
the milled Wood lignin, dioxane lignin, and cellulolytic lignin preparations from Brewer’s
spent grain. Journal of Agriculture and Food Chemistry 63: 603–613.
Sannigrahi, P., Miller, S.J., and Ragauskas, A.J. (2010). Effects of organosolv pretreatment and
enzymatic hydrolysis on cellulose structure and crystallinity in loblolly pine. Carbohydrate
Research 345: 965–970.
Sasaki, C., Wanaka, M., Takagi, H. et al. (2013). Evaluation of epoxy resins synthesized from
steam‐exploded bamboo lignin. Industrial Crops & Products 43: 757–761.
Shen, X.J., Wen, J.L., Huang, P.L. et al. (2017). Efficient and product‐controlled
depolymerization of lignin oriented by Raney Ni cooperated with Cs x H3 − x PW12O40.
Bioenergy Research 10: 1155.
Spiridon, I., Leluk, K., Resmerita, A.M., and Darie, R.N. (2015). Evaluation of PLA–lignin
bioplastics properties before and after accelerated weathering. Composites: Part B 69:
342–349.
Sporck, D., Reinoso, F.A.M., Rencoret, J. et al. (2017). Xylan extraction from pretreated sugarcane
bagasse using alkaline and enzymatic approaches. Biotechnology for Biofuels 10: 296.
Sridach, W. (2010). The environmentally benign pulping process of non‐wood fibers. Suranaree
Journal of Science & Technology 17: 105–123.
Stewart, D. (2008). Lignin as a base material for materials applications: chemistry, application
and economics. Industrial Crops & Products 27: 202–207.
Sudo, K. and Shimizu, K. (1994). Method for manufacturing lignin for carbon fiber spinning.
US Patent US5344921A. https://patents.google.com/patent/US5344921.
Tobimatsu, Y. and Schuetz, M. (2019). Lignin polymerization: how do plants manage the
chemistry so well? Current Opinion in Biotechnology 56: 75–81.
Tolbert, A., Akinosho, H., Khunsupat, R. et al. (2014). Characterization and analysis of the
molecular weight of lignin for biorefining studies. Biofuels, Bioproducts & Biorefining 8:
836–856.
Van Heiningen, A. (2006). Converting a kraft pulp mill into an integrated forest biorefinery.
Pulp & Paper Canada 107: 38–43.
Wang, M., Leitch, M., and Xu, C.C. (2009). Synthesis of phenol–formaldehyde resol resins
using organosolv pine lignins. European Polymer Journal 45: 3380–3388.
Wang, M., Wu, M., and Huo, H. (2007). Life‐cycle energy and greenhouse gas emission impacts
of different corn ethanol plant types. Environmental Research Letters 2: 024001.
Wegener, G., Windeisen, E., Scholz, G. et al. (2006). Material alliance of lignin with natural
fibres. Proceedings of the 9th European Workshop on Lignocellulosics and Pulp, Wien,
pp. 126–129.
Xu, X., Zhou, J., Nagaraju, D.H. et al. (2015). Flexible, highly graphitized carbon aerogels based
on bacterial cellulose/lignin: catalyst‐free synthesis and its application in energy storage
devices. Advanced Functional Materials 25: 3193–3202.
Yang, W., Owczarek, J.S., Fortunati, E. et al. (2016). Antioxidant and antibacterial lignin
nanoparticles in polyvinyl alcohol/chitosan films for active packaging. Industrial Crops and
Products 94: 800–811.
Yuan, T.Q., Sun, S.N., Xu, F., and Sun, R.C. (2011). Characterization of lignin structures and
lignin–carbohydrate complex (LCC) linkages by quantitative 13C and 2D HSQC NMR
spectroscopy. Journal of Agriculture & Food Chemistry 59: 10604–10614.
Zakzeski, J., Bruijnincx, P.C.A., Jongerius, A.L., and Weckhuysen, B.M. (2010). The catalytic
valorization of lignin for the production of renewable chemicals. Chemical Reviews 110:
3552–3599.
Zamudio, M.A.M., Alfaro, A., de Alva, H.E. et al. (2015). Biorefinery of paulownia by
autohydrolysis and soda‐anthraquinone delignification process. Characterization and
application of lignin. Journal of Chemical Technology and Biotechnology 90: 534–542.
Zhang, D.S., Yang, Q., Zhu, J.Y., and Pan, X.J. (2013). Sulfite (SPORL) pretreatment of
switchgrass for enzymatic saccharification. Bioresource Technology 129: 127–134.
Zhang, K., Pei, Z., and Wang, D. (2016). Organic solvent pretreatment of lignocellulosic
biomass for biofuels and biochemicals: a review. Bioresource Technology 199: 21–33.
Zhang, M. and Ogale, A.A. (2014). Carbon fibers from dry‐spinning of acetylated softwood
kraft lignin. Carbon 69: 626–629.
Zhang, X., Tu, M., and Paice, M.G. (2011). Routes to potential bioproducts from lignocellulosic
biomass lignin and hemicelluloses. BioEnergy Research 4: 246–257.
Zhao, L. and Liu, D.X.W. (2007). Effect of several factors on peracetic acid pretreatment of
sugarcane bagasse for enzymatic hydrolysis. Journal of Chemical Technology and
Zhao, X., Cheng, K., and Liu, D. (2009). Organosolv pretreatment of lignocellulosic biomass for
enzymatic hydrolysis. Applied Microbiology and Biotechnology 82: 815–827.
Zhu, J.Y., Pan, X.J., Wang, G.S., and Gleisner, R. (2009). Sulfite pretreatment (SPORL) for
robust enzymatic saccharification of spruce and red pine. Bioresource Technology 100:
2411–2418.
Zhu, X., Peng, C., Chen, H. et al. (2018). Opportunities of ionic liquids for lignin utilization
from biorefinery. ChemistrySelect 3: 7945–7962.
265
12
Conversion of Lignocellulosic Biomass Through Pyrolysis

to Promote a Sustainable Value Chain for Brazilian
Agribusiness
Genyr Kappler1, Débora Machado de Souza2, Carlos Alberto Mendes Moraes1,2,
Regina Célia Espinosa Modolo1,2, Feliciane Andrade Brehm2, Paulo Roberto
Wander1, and Luís António da Cruz Tarelho3
1
Graduate Program of Civil Engineering, University of Vale do Rio dos Sinos – UNISINOS, São Leopoldo, RS, Brasil,
São Leopoldo, Rio Grande do Sul, Brazil
2
Graduate Program of Mechanical Engineering, University of Vale do Rio dos Sinos – UNISINOS, São Leopoldo, RS, Brazil
3
Department of Environment and Planning/Centre for Environmental and Marine Studies (CESAM), University of Aveiro
(UA), Aveiro, Portugal
12.1 Introduction
The global population is currently around 7.37 billion, and it is estimated that it is growing
at the rate of 0.87% per year, which represents an increase of 673 million people in the next
10 years (USDA 2018). The growing demand for food explains the impressive 1.87 billion
hectares used for this purpose on the planet. For this reason, the production of different
types of biomass reaches 1.8 trillion tons (ANEEL 2008). Although there is no world rank-
ing of biomass producers by country, several nations have adopted measures to reuse these
biomaterials. The USA leads in the generation of electrical energy from biomass, Germany
stands out as an important producer of biodiesel, and Brazil is the second largest producer
of ethanol (Minas and Energia 2016).
In Brazil, about 77 million hectares of land is under cultivation which generates plenty
of agricultural waste and hence the country set the implementation of new technologies to
reuse and optimize such biomass waste as a priority. One of the most important agricul-
tural crops in Brazil is sugarcane, in third place in volume produced after soybean and
maize. Approximately 700 million tons of sugarcane was harvested in the country in 2017,
most of which was used to produce sugar, ethanol, and a typical Brazilian alcoholic bever-
age called cachaça (IBGE 2017a,b). The processing of sugarcane generates about 12–15% of
bagasse, which is classified as agro‐industrial organic waste (Cortez et al. 2008). In addition,
yearly production of about 11.7 million tons of rice plays an important role in the country’s
economy, placing it seventh in the list of rice‐producing nations. Rice is another important
source of plant biomass: the husk accounts for roughly 20% of the raw weight of the

roduct. Rice husk mainly contains cellulose (50%), lignin (30%), and an inorganic fraction
p
(20%) (Pouey 2006).
Coconut is another important agricultural product in Brazil; annual coconut production
was reported to be approximately 1.7 million tons, which usually reaches the market in
the form of dehydrated coconut and coconut water. The production cycle of fresh coconut
is six months while the time necessary to produce dehydrated coconut products may be
around 11–12 months (Martin 2013). Approximately 50% of the coconut produced in Brazil
is used to produce coconut water, and around 80–85% of the initial mass is represented by
the shell.
The above‐mentioned three types of biomass are commonly used for the production of
biochar in Brazil using different processes, although the major focus has been on thermo-
chemical conversion.
Briefly, biochar is produced by the thermal decomposition of organic materials in an
oxygen‐starved atmosphere and at fairly low temperatures, usually below 700 °C. The
chemical composition of biochar varies with the biomass used as raw material and the
pyrolysis conditions. In addition to the carbon‐rich organic fraction, biochar has an inor-
ganic fraction that may contain elements like calcium, magnesium, potassium, and inor-
ganic carbonates. Reports have shown that biochar may play an important role in the
development and implementation of measures to improve soil quality (Lehmann et al.
2006), influencing important parameters such as productivity, carbon retention capacity,
and water filtration potential (Lehmann and Joseph 2009). Due to its large specific surface
area, biochar functions as an absorbent, controlling the mobilization of organic and inor-
ganic pollutants (Tong et al. 2014).
The production of biochar from different kinds of lignocellulosic biomass is a promising
alternative in the development of strategies to reuse waste on a local basis (Kumarathilaka
et al. 2016). However, this advantage may only become feasible when the material is
obtained as a co‐product of pyrolysis or gasification. The economic feasibility of these pro-
cesses is a consequence of the fact that these biomasses generate not only biochar itself but
also energy, which may actually be the most important variable in an organization’s com-
petitive edge in its market (Rajapaksha et al. 2016). As a mechanism to sequester carbon,
biochar is a feasible alternative that can contribute to decreasing greenhouse gas emissions.
Due to its molecular structure, which is similar to that of graphite, biochar functions as a
carbon sink in the soil for thousands of years, avoiding the emission of these elements to
the atmosphere (Rezende et al. 2011).
The wide application spectrum of biochar has prompted considerable research across the
globe. Biochar is produced from different kinds of biomass for commercial applications in
an increasing number of countries. As a rule, forest biomass is the most used for the pro-
duction of biochar. However, organic waste from maize farming as well as rice husk, wal-
nut shell, and bagasse are also used, though on smaller scales, depending on the agricultural
profile of the region (UC Davis 2017).
The aim of this chapter is to discuss the potential of the thermochemical conversion of
agricultural residues under slow pyrolysis processes to improve the use of raw lignocellu-
losic biomass. The chapter covers the current panorama of biomass production in Brazil,
the concepts of lignocellulosic biomass and its thermochemical conversion, the products
obtained through slow pyrolysis process, and a thorough examination of some potential
Conversion of Lignocellulosic Biomass Through Pyrolysis 267
applications for the biomass‐derived products of pyrolysis. Additionally, carbonization,

which is being increasingly investigated in the context of cleaner production and circular
economy, is discussed.
12.2 The Concept of Thermochemical Conversion
Plants absorb atmospheric carbon in the form of carbon dioxide (CO2), converting the
molecule into macromolecules based on an intricate chain of specialized reactions called
“photosynthesis” (Rocha et al. 2009). For this reason, the elemental composition of plant
biomass mainly includes carbon, hydrogen, and oxygen (Dufour 2016). During the forma-
tion of plant material, the atoms of these elements bind to form complex chemical
structures such as hemicellulose, cellulose, and lignin (Gómez et al. 2008).
A thermal process can be carried out in the absence or presence of small amounts of
oxygen. This conversion transforms the chemical energy in organic matter and then into
thermal energy. Primary and secondary reactions induce the degradation of biomass,
breaking down lignocellulosic molecules into carbon, gases, and vapors used in the genera-
tion of commercially viable products (Schmal 2017). Table 12.1 shows the reactions taking
place during the degradation of the main molecules in plant biomass (Luengo et al. 2008).
The main thermochemical conversion processes used are roasting, liquefaction, pyroly-
sis, combustion, and gasification (Gómez et al. 2008). Roasting is considered as a thermal
pretreatment of biomass. It is carried out between 200 °C and 300 °C, which characterizes
the devolatilization of hemicellulose and the partial degradation of lignin and cellulose.
The moisture is removed, followed by other compounds of low calorific value. Roasting is
conducted in an inert atmosphere or low oxygen levels in the reaction medium (Gonçalves
et al. 2008). The process reduces oxygen and hydrogen levels in the biomass, increasing its
calorific value and hydrophobicity. In liquefaction, liquid fuels are generated from biomass.
Table 12.1 Thermochemical reactions taking place in plant biomass.
Stage Temperature Biomass reactions
1st Up to 180 °C Heat absorption, heat release, endothermal heat

Between 110 °C Dehydration of –OH groups
and 180 °C
2nd Between 180 °C Degradation of hemicellulose, partial degradation of cellulose,
and 370 °C formation of levoglucosan, release of CO, CO2, and acetic acid
Starting at 250 °C Exothermal reactions
Between 290 °C Total degradation of cellulose, breakage of glucoside bonds in
and 370 °C polysaccharides; high levels of CO, CO2, CH4, H2; formation of acetic
acid, methanol, and ketone
3rd Above 370 °C Complete degradation of lignin, formation of heavy tars and
hydrocarbons
4th Up to 900 °C Increased formation of H2 with longer residence times
Source: Luengo et al. (2008).

Thermochemical conversion occurs at between 300–350 °C and 5–20 MPa (Clark and

Deswarte 2015). In direct liquefaction, biomass is mixed to a solvent at high temperature
over a catalyst. The objective is to produce commercially feasible products with good yield
(Gonçalves et al. 2008).
Pyrolysis is the thermochemical decomposition of organic matter into noncondensable
gases, condensable liquids, and a solid residual product, biochar, or charcoal in the absence
of oxygen. The final product of pyrolysis can be controlled by optimizing the process
parameters, such as temperature and residence time. Depending on the settings, pyrolysis
generates solid (14–45%), gaseous (14–45%), and liquid (50–75%) products (Schmal 2017).
Pyrolysis is characterized by the break of carbon–carbon bonds and the formation of
carbon–oxygen bonds. Part of the biomass is reduced to carbon, while a part is oxidized and
hydrolyzed, thereby producing carbonates, alcohols, carboxylic acids, ketones, and alde-
hydes (Gómez et al. 2008). Several pyrolysis methods such as conventional (or carboniza-
tion), intermediary, and fast pyrolysis have been developed (Schmal 2017). The main
variables in these processes are residence times, heating rates, maximum temperatures,
and desired products. In carbonization, the mean temperature used is 400 °C, residence
times may last for a few days, and the main product obtained is charcoal or biochar.
Based on the partial oxidation of carbon, gasification converts solid carbonaceous mate-
rials into gases used in energy generation processes. The process is carried out in a wide
temperature range, from 550 °C to 1000 °C (Rada 2016). The gaseous product generated is
formed by carbon monoxide (CO), hydrogen (H2), methane (CH4), carbon dioxide (CO2),
nitrogen (N2), and tar. These gases have several applications, such as ingredient in the pro-
duction of chemicals and fuel for internal combustion engines (Gonçalves et al. 2008).
Combustion is an exothermic reaction between oxygen and the hydrocarbons present in
biomass. It is one of the most used thermochemical conversion processes which converts
biomass to energy. The process takes place at high temperatures and excess air, producing
CO2 and steam (Basu 2010). During devolatilization, which is one of the stages of combus-
tion, volatile fuel gases like hydrocarbons, CO, H2, and a liquid phase are formed (Clark
and Deswarte 2015).
12.3 Characterization of Biomass and Its Bioproducts
12.3.1 Biomass as a Renewable Resource

The broadest definition of the term “biomass” is any organic matter derived from nonfos-
silized organisms, whether they are alive or have died recently. The formation of biomass
is directly or indirectly associated with photosynthesis and includes all the animals, plants,
microorganisms, and the organic matter produced or metabolized by them (Basu 2013;
Wang and Luo 2017; Santos 2018). The different types of biomasses play key roles in natu-
ral ecosystems, such as cycling of chemical elements and nutrition of organisms in the
trophic chain. In addition, biomass works as shelter to several forms of life. These bio-
masses are considered as renewable resources generated constantly and are neutral in CO2
emissions due to the fast cycling of elements, which may take days or even years to mobilize
across the various compartments of the biosphere (Martini 2009).
Biomass has always been an important source of materials and energy. It was the main
source of energy until the mid‐nineteenth century when 70% of the world used it as a raw
material in processes devised to produce energy (Grubler and Nakicenovic 1987). However,
recently, biomass has been more extensively used as a result of the development of more
sophisticated processes and technologies to convert it into useful forms of energy, for example
heat and electricity (Gent et al. 2017). One of the oldest processes applied for the conversion
of biomass was carbonization to produce charcoal for combustion purposes (FAO 2018).
From the energy standpoint, biomass is all the renewable resources obtained from non-
fossil organic matter that may be employed to produce useful energy (Brasil 2007; EIA
2011). In this context, biomass includes wood, agricultural crops, grass and woody species
grown for energy, municipal organic waste, and manure. As far as it used as a raw material
source, biomass is the only renewable source of carbon, which highlights it against other
sources of renewable energy. In other words, the technologic applications of biomass are
not restricted to energy generation but also include the production of chemicals
(Bridgewater and Bridge 1991; Hakeem et al. 2015).
Bioproducts may be obtained by either biochemical, physicochemical, or thermochemi-
cal processes. The selection of the method is based mainly on the kind of biomass used and
the final product desired. In biochemical methods, generally enzymes and microorganisms
are used in processes which can take place at only a few dozen Celsius degrees above the
environment temperature (which may take hours or even days to finish, even in the pres-
ence of biocatalysts), whereas thermochemical processes occur at high temperatures (start-
ing at 200–1000 °C). Most thermochemical processes use a variety of raw materials and
convert carbohydrates and lignin into commercially viable products and produce an array
of fuels (Brown 2011). Nevertheless, although they have only recently been developed, pro-
cesses based on pyrolysis afford the extraction of both energy and chemical products from
the biomass, increasing its value.
Despite its enormous potential, biomass by itself does not meet the world’s demand for
energy. However, it has been estimated that biomass will meet the demand for raw materials in
the generation of products used in the synthesis of carbon since its unique composition means
that biomass is especially suitable to play a similar role to that of petrochemical compounds
(Shafizadeh et al. 1976; Klass 1998). New technologies are continuously being researched and
developed to improve the value of biomass, and the drop in fossil fuel resources that is accom-
panied by the rise in prices of these commodities will eventually prompt the move toward
replacing oil refineries by biorefineries in the following decades, with important impacts on all
industrial sectors based on raw material processing (Wild 2011; Pandey et al. 2015).
12.3.2 Lignocellulosic Biomass

Lignocellulosic biomass is one of the most abundant resources on the planet, being widely
distributed geographically. As one of resulting drawbacks from this distribution, its mor-
phology and composition stand as a major challenge in the management and efficient use
of this material. For example, the structure of biomass can be resistant, similar to that of a
fiber, but the material may also be viscous in consistency, like a paste. The density of most
kinds of biomass is low due to the large amounts of water present, with consequences for
shipping and storage logistics.
Forestry Biomass waste generation Products
Agriculture
Figure 12.1 Types of lignocellulosic biomass and a few of its products.
Lignocellulose is the main component of such biomass, accounting for roughly half the
amount of the plant material produced by photosynthesis. The material is the most abun-
dant renewable organic resource (Pérez et al. 2002; Basu 2013). Lignocellulosic biomass is
formed largely by agriculture waste like shells, bagasse, and straw as well as wood scraps
like sawdust, residues from pruning, and general forest residues (Figure 12.1). Lignocellulose
represents an excellent opportunity in the generation of carbon products and second‐
generation primary renewable energy, also shown in Figure 12.1, since this raw material is
generated in large amounts and it does not compete with the food chain. Improving the
value of this biomass raises important issues in the development of sustainable energy and
the mitigation of climate change (ETH 2014; Embrapa 2018).
12.3.3 The Main Characteristics of Lignocellulosic Biomass and Its

Behavior in Pyrolysis
The low energy density, low weight, and high levels of humidity of biomass result in higher
handling, storage, and shipping costs. Therefore, the efficiency of biomass conversion is a
function of the geographic distance between the site of generation and the site where the
biomass will be used. Shipping costs may be reduced using methods to increase density,
such as palletization, torrefaction, and pyrolysis, where the last two also promote decrease
in the hydrogen‐to‐carbon and oxygen‐to‐carbon ratios.
Nonetheless, due to the complexity inherent in thermochemical conversion processes
used to obtain representative yields of quality products, numerous studies have endeavored
to analyze the main components of biomass and the way these behave during pyrolysis.
Generally, these studies are based on gravimetric analysis and bench‐top tests (Neves et al.
2011; Collard and Bin 2014).
Lignocellulosic biomass is formed mainly by three elements: carbon, hydrogen, and oxy-
gen. Together, these elements account for over 95% of the total mass of a plant (de Jong and
van Ommen 2015; Wang and Luo 2017). The atoms of these elements are bound as complex
carbohydrate molecules such as cellulose and hemicellulose as well as aromatic compounds
like lignin, forming the structure of plants. The nonstructural part of plants is formed by
extractives, including waxes, lipids, resins, tannins, sugars, starches, pigments, fatty acids,
phenols, and phytosteroids in addition to ashes, which represent the inorganic fraction of
the plant biomass that remains after total oxidation at high temperatures. Ashes are com-
posed of the main elemental nutrients, like phosphorus and potassium in addition to
smaller levels of sulfur, chlorine, silicon, alkaline earth metals, transition metals, and sev-
eral oligoelements (Lehmann and Joseph 2009; Basu 2013; Wang and Luo 2017). The main
biomass macrocomponents are shown in Figure 12.2.
Another compound found in biomass is water. The removal of water is one of the main
obstacles in the thermochemical conversion of the material, and humidity may potentially
consume, directly or indirectly, most energy contained in biomass. Two forms of water are
present in biomass: free water (or external water) and chemically bound water. Free water
is detected in cell invaginations and intercellular voids in biomass, influencing density,
combustion settings, and permeability of the material. Chemically bound water is detected
in cell walls and comprises roughly 30% of the weight of the dry biomass, affecting most
physicochemical properties of the material.
Pyrolysis (dry distillation) is an efficient thermochemical process compared to other bio-
chemical processes. It converts lignocellulosic materials into products that have an addi-
tional conversion potential: they may be converted into fuels and chemicals that enjoy
increasing commercial importance (Basu 2013; Pandey et al. 2015). During pyrolysis, the
transient heat added to the reactor is conducted by the surrounding gas to the fuel particles,
causing the thermal decomposition of the biomass components. Besides lignin, hemicellulose,
Components of
lignocelulosic biomass
Cell wall
Extractives Ash
components
Non-structural Structural Inorganics
Figure 12.2 Main components of lignocellulosic biomass.

and cellulose, lipids and starches are thermally broken down into three fractions, namely
bio‐oil (condensed vapors), char (the solid fraction), and noncondensable gases (Mohan
et al. 2006). The yield of each of these fractions depends on process settings (heating rate,
reactor temperature, residence time), characteristics of the biomass (particle size, volatiles,
free humidity, ash, fixed carbon), and type of reactor.
The pyrolysis process is carried out in three steps. In the first, the particle is gradually
heated, and, with the increasing temperature, humidity evaporates (drying stage). Next, the
pyrolytic volatiles are progressively released while the chemical bonds of biomass
constituents, such as cellulose, hemicellulose, lignin, and extractives, are broken down
(primary pyrolysis stage).
Pyrolytic gases are composed of permanent gases like CO2, CO, and CH4 and species that
condense at environment temperature and pressure (such as water and a number of organic
compounds). Although each constituent of the biomass decomposes following a specific
temperature and reaction rate profile, primary pyrolysis is completed at relatively low tem-
peratures that generally do not exceed 500 °C. Primary pyrolysis produces a nonvolatile
solid rich in carbon called “char,” or “biochar.” This solid fraction also contains all required
mineral materials which are present in the original biomass fuel. However, if the process is
carried out at high temperatures, some of the primary volatiles that are released inside the
structure of the particle may take part in a variety of secondary reactions, such as cracking,
reforming, dehydration, condensation, polymerization, oxidation, and gasification.
Figure 12.3 shows the products obtained from primary and secondary pyrolysis reactions
(Fagbemi et al. 2001).
Pyrolysis reactions (mostly endothermic)
Drying phase Dehydration Primary and secondary reactions Products
Ambient conditions 100 to 300 °C 300 up to 800 °C Ambient conditions
Water
Chemical Bio-oil
Moisture bond and
free water
Tar
Light gases
Dry
biomass
Permanent gas
Char Char
As-received Torrefied
biomass particle biomass
Figure 12.3 The general mechanism of the pyrolysis process.

12.3.4 Pyrolysis Products

As mentioned above, pyrolysis is a term that describes the process of thermochemical
decomposition or change of carbonaceous materials by the action of heat with restriction
or absence of the oxidizing agent. However, the term is generically used in allusion to pro-
cesses that take place at a wide range of temperature, which starts at 200 °C and goes up to
over 1000 °C. For this reason, depending on the process settings, pyrolysis may be classified
under various categories. The most common categories are “slow pyrolysis” (as in roasting,
carbonization, and conventional pyrolysis) and “fast pyrolysis” (used in fixed, fluidized,
and circulating bed reactors, for example).
The thermochemical decomposition of the biomass generates three products: pyrolysis gas,
pyroligneous extract (also called condensed extract, formed by water and tar), and char, also
called biochar. The type and settings of the process used as well as the kind and geometry of
the biomass particles define the heating rates, yields, and chemical characteristics of each of
the products generated. Generally, long residence times (of around hours or even days) are
used in slow pyrolysis (that is, a process carried out at a heating rate of approximately 2 °C/s
or even less); it occurs at comparatively low temperatures (up to 550 °C) and produces mostly
solid products (i.e., char). In turn, fast pyrolysis, which is carried out at higher temperature
and heating rates (of up to 1000 °C/min) and shorter residence times (a few seconds in some
cases), favors the production of bio‐oil (Klass 1998; Bridgwater and Bridge 1991; Naik et al.
2010; Bridgwater 2012). Pyrolytic gases, which are the permanent gases formed from the
volatilization of organic compounds, are often used to generate the heat required by pyrolysis
or remove the humidity of the biomass fuel (Bridgwater 2006; Liu et al. 2018).
Due to its highenergy content, bio‐oil is becoming a commercially attractive product
because it is easily transported and used with the possibility to decouple production and
application processes (Bridgwater 1994). According to Bertero and Sedran (2015), although
processes based on fast pyrolysis seem to have reached a consolidated position in the indus-
try, the technologies used to convert bio‐oil into fuels or by‐products have not attained such
a high development stage. The routes used for perspective improvement, also called
“upgrading,” include hydrodeoxygenation, cracking with zeolites, mixing with diesel oil to
form an emulsion, and steam reforming to produce hydrogen or synthesis gas (Almeida
2008). Other routes include co‐processing in conventional refineries, gasification followed
by Fischer–Tropsch synthesis, combustion to generate heat (also to feed the pyrolysis pro-
cess itself), and generation of electricity in boilers (Broust and Girard 2006).
The solid products of pyrolysis include the torrefied biomass, fuel char, biocoke, and
biochar (Basu 2013; Gent et al. 2017). The main advantage of the roasting process is the
increase in energy density, as shown in studies published by Khalsa et al. (2016), Thrän
et al. (2016), Ndibe et al. (2015), and Fagernäs et al. (2015). In the steel industry, the char-
coal produced through a carbonization process may be used as a reducing agent (biocoke to
replace coke); the material is used in melting and sintering of iron ores, thermal treatment
of steel, and purification of nonferrous metals (Basu 2013). Charcoal is also widely used as
fuel in the generation of heat to prepare food.
The char has high levels of fixed carbon (between 85% and 87% dry base) and a small
amount of volatiles (between 10% and 12%, in mass dry base) (Basu 2013; Ronsse et al.
2015). When the objective is to use char as biocoke, its physical properties should be favora-
ble to allow using it in a blast furnace. For instance, the compressive strength of char
should be high enough to withstand the heavy load of solids in the blast furnace. In addi-
tion, the material should exhibit good fracture toughness and thus maintain the permeabil-
ity of the load in the furnace to the air blast (FAO 1985). One of the main advantages of
biocoke in blast furnaces is the possibility to use it to replace mineral coal, which in turn
enables the reduction of net CO2 emissions from the steel industry.
The research performed by Dr Wim Sombroek in the 1950s represented an important step
in the investigation of the potential applications of charcoal. Recent studies have looked into
its use as a substrate to improve the physical and chemical characteristics of the soil and
stock carbon (Woods et al. 2009). This was the foundation for the concept of “biochar”
(Lehmann et al. 2003, 2006). For Lehmann and Joseph (2009), the term may be used to high-
light the biological origin of char and thus differentiate it from the carbonized material
originating from plastic and nonbiological materials (Lehmann and Joseph 2009). The
European Biochar Certificate (EBC) defines biochar as a heterogeneous substance rich in
carbon and aromatic compounds produced from the pyrolysis of biomass obtained from
sustainable processes carried out under controlled conditions and using clean technologies.
Moreover, biochar has a variety of applications that do not require fast realization to form
CO2 and may be used to improve soil attributes in some cases (EBC 2012). The EBC also
explains that biochar is produced by pyrolysis under controlled temperature conditions
between 350 °C and 1000 °C. However, although roasting, hydrothermal carbonization, and
charcoal production are pyrolysis processes, the product formed cannot be named biochar.
The characteristics of biochar are quite varied and depend on the source material and
production settings. In the effort to compare the effect of process settings on specific
biomass, Crombie et al. (2013) carried out several studies and concluded that stability of
biochar is a function of the content of fixed carbon in the material. The amount of fixed
carbon, in turn, is affected mostly by heating rate and temperature and the residence time
of the material in the reactor. In other words, if the temperature at which pyrolysis is con-
ducted is higher, the level of fixed carbon and the stability of the product in the soil will be
higher. Ralebitso‐Senior and Orr (2016) assessed the behavior of biochar produced using
pyrolysis processes carried out at low and high temperatures. In the first scenario, the
authors observed that biochar increased agricultural productivity; however, fixed carbon
was less recalcitrant due to the higher reactivity of biochar. In turn, the biochar produced
at high temperature exhibited better water retention capacity values, larger specific area,
higher cation exchange capacity (CEC), and higher recalcitrance. These characteristics
show that the material performs well in sequestering carbon, despite the decrease in the
ability to retain nutrients in soil.
12.4 Use of Biochar in Energy Generation
The current climate panorama underscores the need to develop more diversified energy
sources based on efficient processes and renewable materials. Nevertheless, although several
countries have been developing and implementing clean energy policies, fossil fuels remain
the main energy source across the globe (WEC 2018). In 2016, bioenergy, that is, the energy
obtained from biomass, accounted for approximately 14% of the primary energy on the planet
(WEC 2016). As a flexible form of energy, bioenergy may be converted into solid fuels or
refined for use as solid, gaseous, or liquid biofuel (WEC 2016). However, the properties of
natural biomass do not ensure its direct use. One example of this situation is the restricted use
of the material as solid fuel since the low density, high level of humidity, and difficulties in
transportation mean that it has to be treated prior to use (Ibrahim et al. 2013). Biochar, which
is produced by thermochemical conversion of biomass, changes some specific attributes of
biomass like moisture and density. This advantage widens the array of applications of the
material, whether it is used as a product or by‐product in energy conversion. This potential of
biochar explains the number of studies carried out about this subject in several countries.
A recent study conducted in the King Abdulaziz University in Saudi Arabia presented
the promising results observed when biochar produced from agricultural biomass is used
in the optimization of energy conversion processes such as anaerobic digestion (Wagas
et al. 2018). The samples analyzed had alkaline pH, high porosity level, and good thermal
stability. According to the study, the alkaline pH of biochar may be a useful characteristic
in the production of methane, neutralizing acidity of the raw material during anaerobic
digestion. In turn, silica, aluminum hydroxide, and other compounds present in the car-
bonized biomass improve degradation and bonds in plastic polymers. These advantages
explain the use of the material as a catalyst (Wagas et al. 2018).
Another study was carried out in China on the production of biodiesel in which the effi-
ciency of CaO catalysts obtained from biochar of rice husks was analyzed. The maximum
yield observed was 93.4%, and the authors ascribed this promising value to the possible
bonding between CaO molecules and the silicon atoms present in the biochar obtained
from rice husk. Additionally, the catalysts had good performance with minimal loss, and
could be reused through 120 cycles (Zhao et al. 2018).
In Greece, the potential of biochar obtained from three kinds of biomass (pistachio
shells, nut shells, and sawdust) used as fuel in direct carbon fuel cells (DCFC) or hybrid
carbon fuel cells (HCFC) was tested. The characterization of materials and the electro-
chemical assays carried out showed that DCFC running on biochar from pistachio shells
had the best yield. This advantage was explained based on the appropriate physicochemical
attributes of the material, such as acidity and levels of volatiles (Konsolakis et al. 2014).
Moreover, a study carried out in the University of Saint Andrews, UK, in partnership
with the Instituto Nacional del Carbon in Spain, investigated the performance of char, bio-
char, and graphite in HCFC. The results showed that graphite produced the best results,
since the material is crystalline, has low sulfur content, and its volatiles and humidity con-
tents do not affect the efficiency of the system. The biochar produced from forest biomass
presented very low sulfur levels, although the content of volatiles and humidity affected
performance negatively. This led the authors to conclude that it may be a feasible alterna-
tive material as long as it undergoes pretreatment (Chien et al. 2014).
Other studies on biochar describe additional features. Since biochar has low production
costs and biomass is widely available, it is suitable to produce catalysts. Kastner et al. (2012)
found that catalysts obtained from biochar present high stability in alkaline and acidic
media, which is an advantage over other catalysts. Lee et al. (2017) suggested that biochar
may be used as catalyst support in transesterification reactions due to its large surface area
and suitable thermal stability (Lee et al. 2017).
12.5 Applications of Biochar in Agriculture
As an essential element for life on the planet, the soil is a nonrenewable natural resource.
The main cycles of elements such as the carbon, nitrogen, and oxygen cycles depend on the
soil. Besides being the essential substrate for food production, the soil plays various
important roles; for example, it contributes, though indirectly, to the regeneration of water
sources. It is also fundamental in the hydrologic cycle, nutrient cycling, and support to
biodiversity. In addition, the soil has more obvious functions, such as the base for every
construction ever built by humans.
Nevertheless, the prolonged use of inappropriate practices negatively affects the intrinsic
properties of the soil. As a result, the condition of 30% of the soils on the planet has
degraded a great deal in the second decade of the twenty‐first century (FAO 2015). These
practices worsen compaction, erosion, loss of biota, and of organic matter. In addition,
water retention capacity and pH of soils are also influenced. These issues underscore the
need for rational management and use of soil and the development and implementation of
technologies to optimize the use of natural resources extracted from it.
In the effort to meet the demand for food for the over 7 billion inhabitants of the planet,
currently, almost 1.87 billion hectares are used for cultivation worldwide (USGS 2018).
However, the efficiency of cultivation is a direct function of soil fertility, which is character-
ized by favorable environmental conditions and adequate amounts of nutrients for the spe-
cies cultivated. The bioavailability of macronutrients like carbon, hydrogen, oxygen,
nitrogen, phosphorus, potassium, calcium, and magnesium as well as micronutrients such
as boron, manganese, copper, zinc, iron, molybdenum, and chlorine is essential for the
growth and development of plant species (Lehmann and Joseph 2009). In this effort, the
use of substrates and soil conditioners represents an attempt to meet the physical, chemi-
cal, and biological needs of soils.
The properties of biochar may improve soil quality by increasing soil fertility and the
efficiency of plants to use the available nutrients (Ronquim 2010). The main relationships
between biochar and soils are described below.
12.5.1 pH
Briefly, pH indicates the amount of hydrogen ions in the soil. High levels of hydrogen ions
accompanied by low levels of calcium, magnesium, and potassium ions indicate that the
soil is acidic (Manahan 2010). In agriculture, pH is an important parameter because most
plant species require almost neutral media to develop (Ronquim 2010). Generally, the pH
of biochar is higher than 7, which means that adding it to acidic soils helps to increase the
pH (Mukome and Parikh 2016).
12.5.2 The Carbon-To-Nitrogen Ratio

The carbon‐to‐nitrogen ratio (C:N) is another important indicator of chemical changes in
soil. It is used to predict the mineralization and release of nitrogen in soils. As with most
chemical properties of biochar, C:N values vary considerably with the kind of raw material
used to produce it and pyrolysis conditions (Mukome and Parikh 2016).
12.5.3 Cation Exchange Capacity and Water-Holding Capacity

According to Brazilian regulations, for a mineral to be considered an agricultural soil con-
ditioner, its CEC has to be at least 200 mmol c/kg, while the minimum water‐holding
capacity (WHC) has to be 60% (Brasil 2017). The CEC represents the total amount of cations
retained on the surface of these materials that may be exchanged (namely Ca2+, Mg2+, K++,
H++, and Al3+), and has direct correlation with pH. Also, WHC is one of the main charac-
teristics of biochar, and is a result of porosity and surface area which has the capacity to
retain nutrients in addition to water adsorption (Lee et al. 2013).
12.6 Biochar: Traditional Applications and Future

Perspectives
The use of charcoal and biochar is a very old custom, which is making a comeback. One of
the main advantages of biochar is its wide array of applications, which sometimes are
adopted as a chain, to optimize and recycle material, nutrient, and energy flows (Schmidt
and Wilson 2014). The main applications of biochar may be used in conjunction with new
organic agriculture systems, in addition to its uses in construction, electronics, and a whole
array of consumer goods. Other applications include heating, lighting, food preparation,
metallurgy, and waste confinement. Biochar is also used in medicine and as an additive in
food products (EBC 2012). Today, biochar is considered one of the most important materi-
als in the development and implementation of sustainability strategies across the world.
The interest in biochar has been growing and, due to the large diversity of technologies
and raw materials used to produce it, certification guidelines and production and quality
control mechanisms have been developed based on recent research and empirical observa-
tion of the uses of biochar. In this sense, the European Biochar Certificate (EBC 2012)
defines the guidelines for the sustainable production of biochar in the European
Community. The adoption of these guidelines is not compulsory, except in Switzerland,
where they have to be complied with by agricultural organizations. The International
Biochar Initiative (IBI) has fostered research, development, implementation, and commer-
cialization of technologies to produce biochar since 2006 in many countries. Established in
the USA and relying on its own research groups across the globe, the IBI launched a volun-
tary certification program for biochar aiming at the sustainable development of the indus-
try (IBI 2015).
Biochar is a stable carbon source with singular properties. Besides being an effective
means to implement carbon sequestration, it has high capacity to absorb elements and a
porous structure that may work as habitat for microorganisms. The electrical and thermal
conductivities of biochar have become the object of extensive research. These attributes
explain the recent interest manifested by researchers and policy makers to mitigate some of
the most pressing problems worldwide, such as soil degradation, food security, and climate
change (Woolf et al. 2010).
However, one of the disadvantages of biochar is its high final cost, due to the considera-
ble investment in equipment required to meet certification criteria. On the other hand,
the material is marketed for a wide array of applications. Schmidt and Wilson (2014)
mentioned 55 different uses for biochar and pointed out that their list is not exhaustive.
Several of the applications listed have been extensively adopted, such as use in composting
and as a filtering element to obtain drinking water, industrial adsorbent to treat waste
water, effluents, and gases, food additive in cattle farming, and ingredient in cleaning for-
mulations in animal farming facilities. Nevertheless, a number of applications of biochar
in the production of technologic materials should be investigated in greater depth, such as
its use in the electronics and cosmetics industries, in medicine, well‐being, food preserva-
tion, construction, textiles, and energy (Schmidt and Wilson 2014).
12.7 Conclusion and Final Considerations
Production of certified biochar uses renewable biological materials and emits very low lev-
els of pollutants. Biochar is used in the manufacturing of carbon‐based products, and it is
an energy source promoting carbon neutrality. Concerning applications in the improve-
ment of soil quality and minimization of CO2 accumulation in the atmosphere, the sustain-
ability of biochar production depends on factors that affect the whole production chain, the
logistics of distribution, and use of the product. In this sense, the use of biochar following
a hierarchy of environmental and economic values is gaining increasing relevance today. It
has also been claimed that biochar is too valuable to be added to the soil before its eco-
nomic and environmental potentials are fullyexplored.
The cascade use of biochar adds a series of advantages to the product. For instance, the
production cost is spread across the chain of applications such as the production of animal
feed and the treatment of manure followed by composting and application on the soil. In
these processes, the material adds further benefits, such as the high level of adsorbed nutri-
ents and microbial activation. This adds environmental management advantages and
includes clean production concepts by removing smells and other contaminants from the
source material, while materials that were considered contaminants become products with
other applications.
In addition, considering that biochar closes the loop in agricultural activities by return-
ing its residues to the origin, the idea of a circular economy becomes yet another variable
in this discussion. The circular economy is based on the premise of the efficient use of
materials throughout their life cycle, therefore reducing environmental impacts. These are
key issues in the implementation of sustainability in agribusiness practices. Therefore, the
adoption of technologies that enable the efficient management of materials following cir-
cular economy premises establishes a new value chain. The production of biochar from
residual biomass and its use in several potential environmental applications is an edge
topic in this context.
References
Almeida, M.B.B. (2008). Bioóleo a partir da pirólise rápida, térmica ou catalítica, da palha de
açúcar e seu coprocessamento com gasóleo em craqueamento catalítico. Masters thesis.
Escola de Química da Universidade Federal do Rio de Janeiro, Rio de Janeiro.
ANEEL (Agência Nacional de Energia Elétrica) (2008). Atlas de Energia Elétrica do Brasil,
3rde. Brasilia, Brazil: ANEEL www2.aneel.gov.br/arquivos/PDF/atlas3ed.pdf.
Basu, P. (2010). Biomass Gasification and Pyrolysis: Practical Design and Theory. Burlington,
VT: Academic Press.
Basu, P. (2013). Biomass Gasification, Pyrolysis and Torrefaction: Practical Design and Theory,
2e. Burlington, VT: Academic Press.
Bertero, M. and Sedran, U. (2015). Coprocessing of bio‐oil in fluid catalytic cracking. In: Recent
Advances in Thermochemical Conversion of Biomass (eds. A. Pandy, T. Bhaskar, M. Stocker
and R.K. Sukumaran), 355–382. Amsterdam, The Netherlands: Elsevier.
Brasil, Ministério da Agricultura e Produção Agrária (MAPA) (2017). IN 35, de 11/09/2017.
Estabelece procedimentos para comercialização de substâncias sujeitas à controle especial.
www.agricultura.gov.br/assuntos/insumos‐agropecuarios/insumos‐agricolas/fertilizantes/
legislacao/in‐35‐de‐4‐7‐2006‐corretivos.pdf.
Brasil, Ministério de Minas e Energia (2007). Plano Nacional de Energia 2030. Ministério de
Minas e Energia; colaboração Empresa de Pesquisa Energética. Brasília, Brazil: MME/EPE.
Bridgwater, A.V. (1994). Catalysis in thermal biomass conversion. Applied Catalysis A: General
116 (1–2): 5–47.
Bridgwater, T. (2006). Review: biomass for energy. Journal of the Science of Food and
Agriculture 86: 1755–1768.
Bridgwater, A.V. (2012). Review of fast pyrolysis of biomass and product upgrading. Biomass
Bioenergy 38: 68–94.
Bridgwater, A.V. and Bridge, S.A. (1991). Pyrolysis technologies. In: Biomass Pyrolysis Liquids
Upgrading and Utilization (eds. A.V. Bridgwater and G. Grassi), 11–92. New York: Elsevier
Applied Science.
Broust, F. and Girard, P. (2006). Preconditioning of biomass through fast pyrolysis for different
biofuels applications (PRECOND). 18th European Biomass Conference, Lyon, France.
Brown, R.C. (2011). Fundamentals of pyrolysis. In: Thermochemical Processing of Biomass:
Conversion into Fuels, Chemicals, and Power, 1e (ed. R.C. Brown), 175–206. Ames, IA:
Wiley.
Chien, A.C., Arenillas, A., Jiang, C., and Irvinea, J.T.S. (2014). Performance of direct carbon
fuel cells operated on coal and effect of operation mode. Journal of the Electrochemical
Society 161: 588–593.
Clark, J.H. and Deswarte, F. (2015). Introduction to Chemicals from Biomass, 2e. Chichester,
UK: Wiley.
Collard, F.X. and Blin, J. (2014). A review on pyrolysis of biomass constituents: mechanisms
and composition of the products obtained from the conversion of cellulose, hemicelluloses
and lignin. Renewable and Sustainable Energy Reviews 38: 594–608.
Cortez, L.A.B., Lora, E.E.S., and Gómez, E.O. (2008). Biomassa para Energia. Campinas, SP,
Brazil: Editora da Unicamp.
Crombie, K., Masek, O., and Sohi, S.P. (2013). The effect of pyrolysis conditions on biochar
stability as determined by three methods. Global Change Biology Bioenergy 5: 122–131.
De Jong, W. and van Ommen, J.R. (2015). Biomass as a Sustainable Energy Source for the
Future: Fundamentals of Conversion Processes. Hoboken, NJ: Wiley.
Dufour, A. (2016). Thermochemical Conversion of Biomass for the Production of Energy and
Chemicals. Ames, IA: Wiley.
EBC (2012). European Biochar Certificate – Guidelines for a Sustainable Production of Biochar.

Arbaz, Switzerland: European Biochar Foundation www.zora.uzh.ch/id/
eprint/125910/1/2016_ebc‐guidelines.pdf.
Embrapa (2018). www.embrapa.br/busca‐de‐imagens.
Energy Information Administration (EIA) (2011). Annual Energy Review 2011. www.eia.gov/
totalenergy/data/annual/pdf/aer.pdf.
ETH (2014). www.ethz.ch/en/news‐and‐events/eth‐news/news/2014/04/biochar‐is‐there‐a‐
dark‐side.html.
Fagbemi, L., Khezami, L., and Capart, R. (2001). Pyrolysis products from different biomasses:
application to the thermal cracking of tar. Applied Energy 69: 293–306.
Fagernäs, L., Kuoppala, E., and Arpiainen, V. (2015). Composition, utilization and economic
assessment of torrefaction condensates. Energy & Fuels 29 (5): 3134–3142.
FAO (1985). Industrial charcoal making. Wood carbonization and the products it yields. www.
fao.org/docrep/x5555e/x5555e03.htm.
FAO (2015). Status of the World’s Soil Resources. Intergovernmental Technical Panel on Soils.
www.fao.org/3/a‐i5199e.pdf.
FAO (2018). Energy for Development. www.fao.org/docrep/Q4960E/q4960e04.htm#energy%20
for%20development.
Gent, S., Twedt, M., Gerometta, C., and Almberg, A. (2017). Theoretical and Applied Aspects of
Biomass Torrefaction: For Biofuels and Value‐Added Products.
Oxford, UK: Butterworth‐Heinemann.
Gómez, E.O., Rocha, J.D., Pérez, J.M.M., and Pérez, L.E.B. (2008). Pirólise rápida de materiais
lignocelulósicos para a obtenção de bioóleo. In: Biomassa para Energia (eds. L.A.B. Cortez,
E.E.S. Lora and E.O. Gómez), 353–418. Campinas, SP, Brazil: Editora da Unicamp.
Gonçalves, A.R., Benar, P., and Schuchardt, U. (2008). Liquefação de biomassas. In: Biomassa
para Energia (eds. L.A.B. Cortez, E.E.S. Lora and E.O. Gómez), 419–434. Campinas, SP,
Brazil: Editora da Unicamp.
Grubler, A. and Nakicenovic, N. (1987). The dynamic evolution of methane technologies.
IIASA Working Paper, IIASA, Laxenburg, Austria.
Hakeem, K.R., Jawaid, M., and Alothman, O.Y. (2015). Agricultural Biomass Based Potential
Materials. Cham, Switzerland: Springer International Publishing.
Ibrahim, R.H.H., Darvell, L.I., and Jones, J.M. (2013). Physicochemical characterisation of
torrefied biomass. Journal of Analytical and Applied Pyrolysis 103: 21–30.
Instituto Brasileiro de Geografia e Estatística (IBGE) (2017a). A geografia da cana‐de‐açúcar.
Coordenação de Geografia, IBGE, Rio de Janeiro, RJ, Brazil.
Instituto Brasileiro de Geografia e Estatística (IBGE) (2017b). LSPA, Levantamento Sistemático
da Produção Agrícola: pesquisa mensal de previsão e acompanhamento das safras agrícolas
no ano civil, 2017. IBGE, Rio de Janeiro, pp.1–81.
International Biochar Initiative (IBI) (2015). The standardized product definition and product
testing guidelines for biochar that is used in soil (IBI Biochar Standards). Version 2.1. www.
biochar‐international.org/wp‐content/uploads/2018/04/IBI_Biochar_Standards_V2.1_Final.pdf.
Kastner, J.R., Miller, J., and Geller, D.P. (2012). Catalytic esterification of fatty acids using solid
acid catalysts generated from biochar and activated carbon. Catalysis Today 190: 122–132.
Khalsa, J.H.A., Leistner, D., Weller, N. et al. (2016). Torrefied biomass pellets – comparing
grindability in different laboratory mills. Energies 9: 794.
Klass, D.L. (1998). Biomass for Renewable Energy, Fuels, and Chemicals, 1e. London, UK:
Academic Press.
Konsolakis, M., Kaklidis, N., and Marnellos, G.E. (2014). Assessment of biochar as feedstock in
a direct carbon solid oxide fuel cell. RSC Advances 5: 73399–73409.
Kumarathilaka, P., Mayakaduwa, S., and Herath, I. (2016). Biochar. In: Biochar: Production,
Characterization, and Applications (eds. S.O. Yong, S.M. Uchimiya and X.C. Scott). New
York.: CRC Press.
Lee, Y., Park, J., and Ryu, C. (2013). Comparison of biochar properties from biomass residues
produced by slow pyrolysis at 500°C. Bioresource Technology 148: 196–201.
Lee, J., Kim, K.H., and Kwon, E.E. (2017). Biochar as a catalyst. Renewable and Sustainable
Lehmann, J. and Joseph, S. (2009). Biochar for environmental management: an introduction.
In: Biochar for Environmental Management: Science and Technology (eds. J. Lehmann and S.
Joseph), 1–12. London: Earthscan.
Lehmann, J., Kern, D.C., and Glaser, B. (2003). Amazonian Dark Earths: Origin, Properties,
Management. Amsterdam, The Netherlands: Springer.
Lehmann, J., Gaunt, J., and Rondon, M. (2006). Biochar sequestration in terrestrial
ecosystems – a review. Mitigation and Adaptation Strategies for Global Change 11:
403–427.
Liu, X., Chang, F., and Wang, C. (2018). Pyrolysis and subsequent direct combustion of pyrolytic
gases for sewage sludge treatment in China. Applied Thermal Engineering 128: 464–470.
Luengo, C.A., Felfli, F.E.F., and Bezzon, G. (2008). Pirólise e torrefação de biomassa. In:
Biomassa para Energia (eds. L.A.B. Cortez, E.E.S. Lora and E.O. Gómez), 333–352.
Campinas, SP. Brazil: Editora da Unicamp.
Manahan, S.E. (2010). Environmental Chemistry, 9e. London: CRC Press.
Martini, P.R.R. (2009). Conversão pirolítica de bagaço residual da indústria de suco de laranja e
caracterização química dos produtos. MSc thesis. Universidade Federal de Santa Maria,
Santa Maria, RS, Brazil.
Martins, C.R. (2013). Produção e comercializaçõao de coco no Brasil frente ao comércio
internacional: panorama 2014. Embrapa Tabuleiros Costeiros, Aracaju, SE, Brazil.
Minas e Energia (2016). Biomassa e Bioenergia. http://minasenergia.rs.gov.br/upload/
arquivos/201603/17083210‐13‐sme‐biomassa‐e‐bioenergia.pdf.
Mohan, D., Pittman, C.U. Jr., and Steele, P.H. (2006). Pyrolysis of wood/biomass for bio‐oil: a
critical review. Energy Fuels 20: 848–889.
Mukome, F.N.D. and Parikh, S.J. (2016). Chemical, physical, and surface characterization of
biochar. In: Biochar: Production, Characterization, and Applications (eds. Y.S. Ok, S.M.
Uchimiya and S.X. Chang), 70–99. New York: CRC Press.
Naik, S.N., Goud, V.V., and Prasant, K.R. (2010). Production of first‐ and second‐generation
biofuels: a comprehensive review. Renewable Sustainable Energy Review 14: 578–597.
Ndibe, C., Maier, J., and Scheffknecht, G. (2015). Combustion, cofiring and emissions
characteristics of torrefied biomass in a drop tube reactor. Biomass and Bioenergy 79: 105–115.
Neves, D., Thunman, H., and Matos, A. (2011). Characterization and prediction of biomass
pyrolysis products. Progress in Energy and Combustion Science 37 (5): 611–630.
Pandey, A., Bhaska, T., and Stöcker, M. (2015). Recent Advances in Thermochemical Conversion
of Biomass. Amsterdam, The Netherlands: Elsevier BV.
Pérez, J., Muñoz‐Dorado, J., and De‐la‐Rubia, T. (2002). Biodegradation and biological treatments
of cellulose, hemicellulose and lignin: an overview. International Microbiology 5: 53–63.
Pouey, M.T.F. (2006). Beneficiamento da cinza de casca de arroz residual com vistas à produção
de cimento composto e/ou pozolânico. PhD thesis. Universidade Federal do Rio Grande do
Sul, Porto Alegre, RS, Brazil.
Rada, E.C. (2016). Thermochemical Waste Treatment Combustion, Gasification, and Other
Methodologies. Toronto, Canada: Apple Academic Press.
Rajapaksha, A.U., Mohan, D., and Igalavithana, A. (2016). Definitions and fundamentals of
biochar. In: Biochar: Production, Characterization, and Applications (eds. Y.S. Ok, S.M.
Uchimiya and S.X. Chang), 4–17. New York: CRC Press.
Ralebitso‐Senior, T.K. and Orr, C.H. (2016). Biochar Application: Essential Soil Microbial
Ecology. Amsterdam, The Netherlands: Elsevier.
Rezende, E.I.P., Angelo, L.C., and dos Santos, S. (2011). Biocarvão (biochar) e sequestro de
carbono. Revista Virtual de Química 3 (5): 426–433.
Rocha, J.C., Rosa, A.H., and Cardoso, A.A. (2009). Introdução à química ambiental, 2e. Porto
Alegre, Brazil: Bookman.
Ronquim, C.C. (2010). Conceitos de fertilidade do solo e manejo adequado para as regiões
tropicais. Campinas, SP, Brazil: Embrapa Monitoramento por Satélite.
Ronsse, F., Nachenius, R.W., and Prins, W. (2015). Carbonization of biomass. In: Recent
Advances in Thermochemical Conversion of Biomass (eds. A. Pandey, T. Bhaskar, M. Stocker
and R.K. Sukumaran). Amsterdam, The Netherlands: Elsevier BV.
Santos,V.S.S. (2018). Ecologia: Ciclos biogeoquímicos. https://mundoeducacao.bol.uol.com.br/
biologia/ciclos‐biogeoquimicos.htm.
Schmal, M. (2017). Cinética e reatores: Aplicação à engenharia química: teoria e exercícios, 3e.
Rio de Janeiro, Brazil: Synergia COPPE/UFRJ.
Schmidt, H.P. and Wilson, K. (2014). The 55 uses of biochar. Biochar Journal, Arbaz,
Switzerland. www.biochar‐journal.org/en/ct/2.
Shafizadeh, F., Sarkanen, K.V., and Tillman, D.A. (1976). Thermal Uses and Properties of
Carbohydrates and Lignins. New York: Academic Press.
Thrän, D., Witt, J., Schaubach, K. et al. (2016). Moving torrefaction towards market
introduction – technical improvements and economic‐environmental assessment along the
overall torrefaction supply chain through the SECTOR Project. Biomass and Bioenergy 89:
184–200.
Tong, H., Hu, M., and Li, F.B. (2014). Biochar enhances the microbial and chemical
transformation of pentachlorophenol in paddy soil. Soil Biology and Biochemistry 70:
142–150.
UC Davis Biochar Database (2017). Characterization database. http://biochar.ucdavis.edu/
download.
USDA (United States Department of Agriculture) (2018). USDA Agricultural Projections to
2027. www.ers.usda.gov/publications/pub‐details/?pubid=87458.
USGS (2018). Science for a changing world. www.usgs.gov/special‐topic/science‐a‐changing
‐world.
Wagas, M., Aburiazaiza, A.S., and Miandad, R. (2018). Development of biochar as fuel and
catalyst in energy recovery technologies. Journal of Cleaner Production 188: 477–488.
Wang, S. and Luo, Z. (2017). Pyrolysis of Biomass: Green Alternative Energy Resources. Berlin,
Germany: Walter de Gruyter.
Wild, P., Reith, H., and Heeres, E. (2011). Biomass pyrolysis for chemicals. Biofuels 2 (2).
Woods, W.I., Teixiera, W.G., and Lehmann, J. (2009). Amazonian Dark Earths: Wim Sombroek’s
Vision. Heidelberg, Germany: Springer.
Woolf, D., Amonette, J.E., and Street‐Perrott, F.A. (2010). Sustainable biochar to mitigate global
climate change. Nature Communications 1: 56.
World Energy Council (WEC) (2016). World Energy Resources 2016. London: World Energy
Council.
World Energy Council (WEC) (2018). Issues Monitor 2018: Perspectives on the Grand Energy
Transition. London: World Energy Council.
Zhao, C., Yang, L., and Xing, S. (2018). Biodiesel production by a highly effective renewable
catalyst from pyrolytic rice husk. Journal of Cleaner Production 199: 772–780.
285
13
Integrated Process of Biomass Thermochemical

Conversion to Obtain Pyrolytic Sugars for Biofuels
and Bioproducts
Victor Haber Perez1, Nathalia Ribeiro Ferreira da Silva1, Euripedes Garcia
Silveira Junior1, Diego Cunha Rocha1, Oselys Rodriguez Justo2, Geraldo
Ferreira David3, Diana Catalina Cubides Roman3, Valdemar Lacerda, Jr3, and
Manuel Garcia-Perez4
1
State University of Northern Rio de Janeiro, Campos dos Goytacazes, Rio de Janeiro, Brazil
2
Estácio de Sá University, Campos dos Goytacazes, Rio de Janeiro, Brazil
3
Federal University do Espírito Santo, Vitoria, Espírito Santo, Brazil
4
Biological System Engineering, Washington State University, Pullman, WA, USA
13.1 Introduction
The sugar and alcohol industries traditionally use co‐generation systems in which sugar-
cane bagasse is burned in boilers to produce steam and electricity, but there is still a large
surplus of bagasse available, and this excess has become an economic and environmental
problem. However, this by‐product can be effectively used for the production of second‐
generation ethanol, and nowadays intense research efforts are ongoing for the development
and/or improvement of the several stages involved in this process and hence, it is believed
that it is possible to increase bioethanol production without expanding the cultivation areas.
In Brazil, the technology adopted for second‐generation bioethanol production is based on
the hydrolysis of biomass. However, this process presents many drawbacks encountered in the
pretreatment methods for total delignification of lignocellulosic biomass to obtain the sugars
present in its structure (Sarkar et al. 2012; Shahsavarani et al. 2013; Tran et al. 2013). Thus, the
main technologic challenges are focused on pretreatment of the biomass, its hydrolysis to
obtain the sugars (from cellulose and hemicellulose) and the stage of fermentation (i.e., find-
ing microorganisms for the conversion of sugars, mainly pentoses, into bioethanol).
On the other hand, biomass thermochemical conversion is commonly used as alternative
technology to make these sugars available for fermentation. In this process, biomass decom-
poses into three main products, bio‐oil, biocarbon, and noncondensable gases, and depending
on the process parameters, different proportions of these products can be obtained. Table 13.1
shows the different types of thermochemical processes as well as the percentage of the forma-
tion of their principal products. Among the thermochemical processes, combustion, roasting,
pyrolysis, and gasification are considered most important. These represent more than 95% of

Table 13.1 Product formation during the different biomass thermochemical conversion processes.
Condensable and noncondensable

gases
Thermochemical Process Biochar Liquid Noncondensable

process parameters (%) fraction (%) gases (%)
Torrefaction 200–300 °C, low heating rate, long 80 0 20

residence time of vapors
Fast pyrolysis 350–600 °C, high heating rate, 12 75 13
short residence time of vapors (‹2 s)
Gasification 800–900 °C, high heating rate, long 10 5 85
residence time of vapors
Source: Bridgwater (2003, 2012).
biomass energy valorization (Ortiz et al. 2013). However, among these technologies, pyrolysis
and gasification are the most advantageous because it is possible to achieve a greater conver-
sion of energy compared with combustion processes (Garcia‐Perez et al. 2002). In addition,
fast pyrolysis optimizes the production of liquid product (bio‐oil), gasification maximizes gas
formation (syngas), and roasting maximizes the formation of solids and biochar.
Pyrolysis is a thermal decomposition process that occurs at moderate temperatures with a
high heat transfer rate to the biomass particles and a short hot vapor residence time in the reac-
tion zone (Czernik and Bridgwater 2004). The pyrolysis process can be slow or fast and is prom-
ising for converting biomass into several compounds of industrial interest. The reactor type to
be used in the pyrolysis depends on the product desired. Reactors used for a fast pyrolysis sys-
tem can be a fluidized bed, rotating cones, entrained flow, vacuum, and ablative (Roy and Dias
2017; Perkins et al. 2018; Sharifzadeh et al. 2019). Various types of biomass such as agricultural
residues, forestry and city wastes are abundantly available and can be used for energy produc-
tion in several ways (Asadullah et al. 2007). With this technology, large volumes of bio‐oil are
produced, approximately 80% on a dry biomass basis (Czernik and Bridgwater 2004; Montoya
et al. 2015; Wang et al. 2018b). The yield and quality of products obtained by pyrolysis depend
on several factors, such as the pyrolysis technology used, the configurations of equipment, oper-
ating conditions, type of sample, pretreatment conditions, etc. (Roy and Dias 2017).
This chapter deals with the general aspects of obtaining pyrolytic sugars from sugarcane
bagasse by fast pyrolysis in order to obtain bioethanol in a process that can be integrated
with an autonomous first‐generation ethanol production plant.
13.2 Production of Pyrolytic Sugars from Biomass

and Applications
The lignocellulosic biomass is mainly composed of cellulose (38–50%), hemicellulose

(23–32%), lignin (15–25%), and small amounts of ash content (Hameed et al. 2019; Neto
et al. 2019). The polysaccharides, cellulose and hemicellulose, form anhydrosugars, furans,
aldehydes, ketones, and carboxylic acids as their primary volatile products. Lignin pyroly-
sis forms low molecular weight and aromatic compounds (Shen et al. 2015).
Integrated Process of Biomass Thermochemical Conversion 287
Cellulose is usually the most abundant component of lignocellulosic biomass and thus it
can provide significantly higher yields of anhydrosugars (Lu et al. 2018). An important
product of cellulose pyrolysis is 1,6‐anhydro‐β‐D‐glucopyranoside (levoglucosan). These
anhydrosugars exhibit a cyclic 1,6‐anhydro ring structure in addition to the canonical pyra-
nose ring of six‐membered carbohydrates containing five carbons and one oxygen atom
(Bacik and Jarboe 2016). Understanding of the mechanisms of levoglucosan formation
during fast pyrolysis is essential for the development of efficient pyrolysis techniques.
There are four mechanistic proposals for the pyrolysis of biomass reaction: free‐radical
mechanism, glucose intermediate mechanism, levoglucosan chain‐end mechanism, and
degradation of cellulose (Meier and Faix 1999; Zhang et al. 2013).
However, fast pyrolysis of raw biomass produces a much lower amount of levoglucosan
than the yield from pure cellulose (Li et al. 2013). Hence, to increase the conversion of cellu-
lose from biomass to levoglucosan, it is essential to perform a pretreatment on the lignocellu-
losic material, seeking to improve the levoglucosan yield during pyrolysis. The depolymerization
of cellulose leads to the formation of a high proportion of anhydrooligosaccharides and anhy-
drosaccharides, especially of levoglucosan whose yield can reach up to 60% (Collard and Blin
2014). The acid treatment removes alkali ions known to decrease levoglucosan yields by two
connected pathways. Ions hinder cellulose depolymerization into anhydrous sugars, and, once
depolymerized, ions serve as catalysts in anhydrosugar fragmentation reactions (Luque et al.
2014). Molecular modeling techniques explain that alkaline and alkaline earth metals
(AAEMs) alter the electronic structure of the carbohydrate by interacting with oxygen, affect-
ing the stereochemistry of the molecules during reactions; because of this, rearrangement and
dehydration reactions are enhanced, followed by fragmentation reactions. It is thought that
AAEMs can directly attack the cellulose chain before and during depolymerization reactions
as well as catalyze reactions in the liquid intermediate phase (Pecha et al. 2015). The observed
effects confirm established opinions about the inhibiting action of metals on the depolymeri-
zation process of cellulose during pyrolysis (Dobele et al. 2005; Pecha et al. 2015).
13.2.1 Main Components Influencing Formation of Levoglucosan

from Biomass
An understanding of the relationship between the changes in physicochemical characteris-
tics and levoglucosan formation is essential for the development of new techniques and
biomass pretreatment strategies. Factors such as biomass composition, interactions between
the constituents of biomass during pyrolysis, structure, degree of polymerization and crys-
tallinity of cellulose, and the presence of AAEMs need to be more extensively studied.
The minerals (AAEMs) present in the structure of lignocellulosic biomass strongly influ-
ence the bio‐oil and levoglucosan production during fast pyrolysis. In some species of biomass,
it is more common to find the following elements: Na, K, Mg, Ca, Al, Mn, Co, Ni, Cu, Zn, and
Ba, and the total mineral content in most biomass can be in the 10% range or higher for straws,
grasses, and leaves (Pecha et al. 2015). Several strategies for pretreatment of biomass can be
employed for the removal or passivation of these minerals before the fast pyrolysis process
(Dobele et al. 2001; Tan and Wang 2009; Wang et al. 2011; Eom et al. 2012; Oudenhoven et al.
2013; Zhou et al. 2013; Dong et al. 2015a; David et al. 2018). The presence of inorganic metals
can be responsible for secondary reactions (cracking and/or condensation) of pyrolytic com-
pounds initially produced by primary pyrolysis reactions, which results in changes in chemi-
cal distributions in bio‐oil and its yield (Mourant et al. 2011; Eom et al. 2012).
During the pyrolysis of biomass, the AAEMs alter the carbohydrate structure by interacting
with the oxygen atoms, affecting the stereochemistry of the molecules during the reactions,
where rearrangement and dehydration reactions are intensified, followed by the fragmentation
reaction as explained previously. In consequence, bio‐oil and levoglucosan yields decrease
(Hameed et al. 2019). Haddad et al. (2017) studied the pyrolysis mechanism of lignocellulosic
biomass decomposition with and without presence of AAEMs. It was observed that potassium
and sodium accelerated the thermal degradation of hemicellulose and modified the catalytic
effects on cellulose degradation followed by condensation of low molecular weight compounds
and consequently an increase in the yield of biochar. In this sense, pretreatment with a Bronsted
acid, for instance acetic acid, is able to remove this negative interference, leading to a ninefold
improvement of levoglucosan production when compared with the untreated biomass (David
et al. 2018).
13.2.2 Pretreatments of Biomass for Levoglucosan Production

For levoglucosan production from biomass, the use of appropriate pretreatments of the
feedstock is essential for success. Several strategies have been performed to improve levo-
glucosan yield, such as acid washing (Mourant et al. 2011; Oudenhoven et al. 2016), acid
impregnation (Patwardhan et al. 2010), calcium hydroxide pretreatment (Hassan et al.
2009), hot water (Liang et al. 2015; Zhuang et al. 2016), microwave (Zheng et al. 2018), and
the use of microorganisms (Yang et al. 2011). The aim of these processes is to remove the
AAEM compounds from the biomass during pretreatment. The acid washing of lignocel-
lulosic material is employed for demineralization, that is, it removes AAEMs from biomass
(David et al. 2018). Acid impregnation is used to passivate the catalytic influence of AAEMs
to form thermally stable salts (Zhou et al. 2013; Pecha et al. 2015). A study using sugarcane
bagasse demonstrated that the combination of acid washing and acid impregnation was the
best approach to increase levoglucosan yield. This study used HNO3 washing followed by
H2SO4 (0.2 wt%) impregnation and achieved yields between five and seven times higher
than untreated biomass (David et al. 2017). Figures 13.1 and 13.2 are chromatograms of
sugarcane bagasse under fast pyrolysis at 400 °C for untreated and previously treated bio-
mass by acetic acid, respectively. These results obtained via pyrolysis–gas chromatography–
mass spectrometry (Py‐GC/MS) showed that levoglucosan formation is improved after
treatment, around nine times higher than untreated biomass (David et al. 2019).
The alkaline pretreatment (calcium hydroxide [Ca(OH)2] and ammonium hydroxide
[NH4OH], among other catalysts) is used for neutralizing the effects of acids present within
the cellulose particles (Hassan et al. 2009; Shaik et al. 2013). In the hot water process, water
penetrates into the plant cell structure, releasing hemicellulose and part of the lignin, and
hydrating cellulose (Johnson et al. 2009; Zhuang et al. 2016). Another strategy used for
pretreatment is the use of microorganisms to alter the structure or chemical composition
of the feedstock, facilitating the ability to liberate sugars (Madadi and Hin 2017). However,
this biopretreatment method is not considered efficient when compared with chemical or
other pretreatments because it has a low yield of levoglucosan when compared with the
control experiments and requires a long pretreatment time.
An efficient pretreatment can promote improved levoglucosan formation during fast
pyrolysis and reduce the generation of toxic inhibitors in the condensed products of pyroly-
sis. Moreover, it can be economically viable, due to the low cost of equipment, being more
environmentally friendly, and reducing the demand for waste disposal. However, more
27 O
O
4 OH
1
24 OH
2 OH
Absorbance
Levoglucosan
18
3 14 34
11
28 38
7
5 33
9 30
22
6 10 26
35 36 39 40
13 19 20 42
41 43
0 5.0 10.0 15.0 20.0 25.0 30.0
Time (min)
Figure 13.1 Pyrolysis-GC/MS chromatogram of condensable compounds at 400 °C for natural

sugarcane bagasse. The identified compounds were: (1) Carbon dioxide, (2) Propanoic acid, 2-oxo,
(3) Acetaldehyde, (4) Acetic acid, (5) 2-Propanone, 1-hydroxy, (6) not identified, (7) 1,2-Ethanediol,
monoacetate, (8) Propanoic acid, 2-oxo-, methyl ester, (9) Furfural, (10) 2-Furanmethanol, (11)
2-Cyclopenten-1-one, 2-hydroxy, (12) not identified, (13) Butanedioic acid, methyl, (14) 2,4(3H,5H)-
Furandione, 3-methyl, (15) 1,2-Cyclopentanedione, 3-methyl, (16) not identified, (17) not identified,
(18) Phenol, 2-methoxy, (19) not identified, (20) ni, (21) Phenol, 3,4-dimethyl, (22) Phenol, 2-methoxy-
4-methyl, (23) not identified, (24) Benzofuran, 2,3-dihydro, (25) 1,4:3,6-Dianhydro-α-d-glucopyranose,
(26) Phenol, 4-ethyl-2-methoxy, (27) 2-Methoxy-4-vinylphenol, (28) Phenol, 2,6-dimethoxy, (29)
Vanillin, (30) Phenol, 2-methoxy-4-(1-propenyl), (31) Phenol, 2-methoxy-4-propyl, (32) Phenol,
3-methoxy-2,4,6-trimethyl, (33) Levoglucosan, (34) 3′,5′-Dimethoxyacetophenone, (35) not identified,
(36) Benzaldehyde, 4-hydroxy-3,5-dimethoxy, (37) not identified, (38) Phenol, 2,6-dimethoxy-4-
(2-propenyl), (39) Ethanone, 1-(4-hydroxy-3,5-dimethoxyphenyl), (40) Desaspidinol, (41) not
identified, (42) not identified, (43) 3,5-Dimethoxy-4-hydroxycinnamaldehyde.
O
O
OH
OH
Absorbance
24 OH
Levoglucosan
11 20
21 28
4 23
7
2 10 29
1 3 17 26
25 30
6 89 14 15 16 18 22 27 31 32
12 13 33
0 5.0 10.0 15.0 20.0 25.0 30.0
Time (min)
Figure 13.2 Pyrolysis-GC/MS chromatogram of condensable compounds at 400 °C for sugarcane

bagasse treated with acetic acid. The identified compounds were: (1) Carbon dioxide, (2) 2Propanoic
acid, 2-oxo, (3) Acetaldehyde, (4) Acetic acid, (5) 2-Propanone, 1-hydroxy, (6) 1,2-Ethanediol,
monoacetate, (7) Furfural, (8) 2-Cyclopenten-1-one, 2-hydroxy, (9) Butanedioic acid, methyl, (10)
2,4(3H,5H)-Furandione, 3-methyl, (11) 1,2-Cyclopentanedione, 3-methyl, (12) Phenol, 2-methoxy,
(13) 2,5-Dimethyl-4-hydroxy-3(2H)-furanone, (14) Phenol, 3,4-dimethyl, (15) ni, (16) Phenol,
2-methoxy-4-methyl, (17) not identified, (18) Benzofuran, 2,3-dihydro, (19) 1,4:3,6-Dianhydro-à-d-
glucopyranose, (20) Phenol, 4-ethyl-2-methoxy, (21) 2-Methoxy-4-vinylphenol, (22) not identified,
(23) Phenol, 2,6-dimethoxy, (24) Vanillin, (25) Phenol, 2-methoxy-4-(1-propenyl), (26) Phenol,
2-methoxy-4-propyl, (27) Phenol, 3-methoxy-2,4,6-trimethyl, (28) Levoglucosan, (29) Phenol,
2,6-dimethoxy-4-(2-propenyl), (30) not identified, (31) Ethanone, 1-(4-hydroxy-3,5-
dimethoxyphenyl), (32) Desaspidinol, (33) 3,5-Dimethoxy-4-hydroxycinnamaldehyde.
investigation is essential to develop new and low‐cost pretreatments suitable for yield of
levoglucosan from lignocellulosic biomass by fast pyrolysis (Jiang et al. 2019).
13.2.3 Extraction of Levoglucosan from Bio-oil and Hydrolysis

Water and several organic solvents such as ethanol, methanol, isopropanol, ether, and ethyl
acetate have been used for sugar extraction from bio‐oil (Oasmaa et al. 2016; Wang et al.
2016). It is reported that the extraction conditions, such as solvent volume, temperature,
and time of extraction, can affect the recovery of levoglucosan. Lian et al. (2010) used
blends of fatty acid methyl esters (FAMEs) and ethyl acetate to extract levoglucosan from
bio‐oil. These extraction studies aimed to identify the optimal conditions required to selec-
tively remove phenols, furans, and carboxylic acids from the dissolved oil, without extract-
ing sugars. The mixture was shaken for 10 minutes at 30 °C and the fractions were separated
by decantation. The anhydrosugars concentrated in the aqueous phase were used in the
fermentation step and the undesirable compounds were separated (Lian et al. 2010). In
another study, the extraction of levoglucosan was carried out using water. The procedure
occurred in two steps: a hot water extraction step and a cold water separation step. In the
first step (at 60 °C), the hydrogen bonds between lignin structures present in the bio‐oil
break, and bound levoglucosan is released into the aqueous phase. After mixing, the solu-
tion was stored at 4 °C overnight to precipitate nonpolar lignin oligomeric material in the
suspension and a rich phase with levoglucosan was obtained (Lian et al. 2013).
Bennett et al. (2009) optimized the extraction of levoglucosan from bio‐oil using just water
as a solvent. The optimal extraction conditions were 41 wt% water at 34 °C and a concentration
of up to 88 g/L of levoglucosan was obtained (Bennett et al. 2009). Wang et al. (2016) used the
response surface technique to optimize parameters to extract levoglucosan. The authors found
conditions that quickly and easily extracted levoglucosan from bio‐oil using a Soxhlet system
for extraction followed by precipitation. NaOH and Ca(OH)2 powders were used to remove
organic colloid compounds and pigments; a mixture of ethyl acetate and water (2:1, v/v) was
used as the solvent for levoglucosan extraction from other compounds into bio‐oil and 78%
levoglucosan with purity of 95.56% was obtained using Ca(OH)2 (Wang et al. 2016).
Bennett et al. (2009) reported the best conditions for levoglucosan hydrolysis (125 °C, for
44 minutes using H2SO4 0.5 M) that resulted in a maximum glucose yield. These authors
observed that temperature was the most significant parameter in this reaction because
increasing the reaction temperature to 130 °C increased the reaction rate, while further
increases resulted in sugar degradation. In addition, two reaction options were proposed,
with shorter exposure times at higher temperatures and longer reactions at lower tempera-
tures. The levoglucosan hydrolysis reaction is a stoichiometrically equimolar reaction, that
is, one mole levoglucosan yields one mole of glucose (Sukhbaatar et al. 2014); however, the
glucose concentration is greater than the initial concentration of levoglucosan. This fact is
attributed to the hydrolysis of cellobiosan or other oligomeric forms of anhydrosugars pre-
sent in bio‐oil (Yu and Zhang 2003).
13.2.4 Detoxification of Levoglucosan

One of the challenges in conversion of levoglucosan from bio‐oil by the fermentation process
is the presence of inhibitory compounds (Shen et al. 2015). These inhibitors must be removed
or inactivated to detoxify the sugar‐containing aqueous fraction enough to be fermented.
Compounds such as 5‐hydroxymethylfurfural, furfural, acetic acid, phenolic compounds,

and others inhibit microbial growth and consequently the fermentation process (Lian et al.
2010). Many of these undesirable compounds are lignin derived (Sukhbaatar et al. 2014). The
removal of inhibitory compounds is essential for a successful fermentation process. In this
context, it is very important to identify the main inhibitors present in the aqueous phase or
hydrolyzate so that an adequate strategy can be chosen to remove them (Jarboe et al. 2011).
Several detoxification strategies have been used to remove undesirable chemicals, which
mainly include activated carbon, air stripping, overliming, solvent extraction, microbial
digestion, and development of microorganisms resistant to inhibitory compounds (Chan
and Duff 2010; Wang et al. 2012). Among these methods, some are specific to certain types
of compounds, and hence sometimes it is necessary to combine two or more methods (Lian
et al. 2010). However, there is a need to develop methods able to remove inhibitors in a selec-
tive way, inexpensively, and with minimal loss of fermentable sugars (Jiang et al. 2019).
Studies of metabolic evolution have been utilized in the development of biocatalysts capa-
ble of fermenting bio‐oil. Yeasts have been adapted to bio‐oil previously detoxified by solvent
extraction and then hydrolyzed to convert anhydrosugars to glucose (Jarboe et al. 2011).
However, more research should focus on the development of more efficient detoxification
methods for a less toxic substrate and robust strains that are highly tolerant of toxicity. In
addition, it can be interesting to understand the interaction mechanisms of these undesirable
compounds with the microorganisms to improve detoxification procedures (Shen et al. 2015).
13.2.5 Levoglucosan Fermentation in Biofuels and Other Chemicals

The levoglucosan can be directly fermented or further hydrolyzed to glucose before the fer-
mentation process (Wang et al. 2010; Shen et al. 2015). Levoglucosan can be converted to glu-
cose by acid hydrolysis or catalysis, but these additional processing steps represent an extra
cost. The development of microorganisms that can directly metabolize levoglucosan is desir-
able (Jarboe et al. 2011). Some microorganisms can utilize levoglucosan naturally as the sole
carbon and energy source, but for most microorganisms it is difficult to utilize levoglucosan
because it is in the anhydrous form (Rover et al. 2014). Some industrial workhorse strains,
such as Escherichia coli, can be engineered for direct utilization of levoglucosan. Levoglucosan
kinase (lgk) is the key enzyme required for these microorganisms to convert levoglucosan into
glucose‐6‐phosphate, which then enters the glycolysis pathway. The modularity of this path-
way has been demonstrated by its functional expression in bacteria such as E. coli (Dai et al.
2009; Layton et al. 2011; Bacik and Jarboe 2016). Other distinct biological pathways have been
identified during levoglucosan degradation. A bacterial levoglucosan dehydrogenase pathway
converts levoglucosan to 3‐keto‐levoglucosan in an NAD+‐dependent reaction, but levoglu-
cosan membrane transporters remain unknown (Ning et al. 2007; Bacik et al. 2015).
Some of the studies summarized in Table 13.2 showed that levoglucosan and hydrolyzate
glucose can be efficiently fermented to ethanol. Sometimes, yields product/substrates were
similar to those attained using glucose (Zhuang et al. 2001; Wang et al. 2010). In addition,
Table 13.3 shows several reports about levoglucosan fermentation to attain other important
products, such as styrene, lipids, and citric acid using different microorganisms. The potential
applications of these sugars are interesting from the perspective of bioengineering. Knowledge
of the existing biological pathways related to these sugars must be expanded if they are to be
used effectively as carbon and energy sources for the fermentative production of renewable
fuels and chemicals (Zhuang and Zhang 2002; Bacik et al. 2015).
Table 13.2 Fermentation processes using levoglucosan as the carbon source for bioethanol production.
Pyrolysis Fermentation
Feedstock Pretreatment conditions Treatment Strain process Product yield References
Pinewood H3PO4 solution Pyrolysis Extraction with n‐butanol + hydrolyzed Saccharomyces Fermentation 0.5 g of ethanol/ g of Sukhbaatar
particles (0.1 wt%) and vapor with H2SO4 (0.5 M) autoclaved at 125 °C pastorianus flask glucose; 98% of the et al. (2014)
heating to stream for 44 min + neutralization with NaOH theoretical yield
100 °C for 1 h (5 M) and CaCO3 + filtered by reverse
osmosis
Poplar wood Acid wash Pyrolysis at LGA extracted with ethyl acetate/ Saccharomyces Erlenmeyer 0.473 g ethanol/g Lian et al.
500 °C in a biodiesel blends + hydrolysis with cerevisiae ATCC flask glucose (2010)
fluidized‐ H2SO4 (0.5 M) at 00062
bed reactor 120 °C/240 min + neutralization by
Ba(OH)2 + detoxification with activated
carbon
Bio‐oil None None LGA hydrolysis at 120 °C with H2SO4 S. cerevisiae T2 Flask in 0.19 g ethanol/g Chan and
(0.5 M) + overliming with Ca(OH)2 shaker glucose at 50% volume Duff (2010)
hydrolyzate and 0.45 (g
ethanol/g glucose) at
40% volume
hydrolyzate
Loblolly pine Acid Pyrolysis at Water extraction + hydrolysis at 120 °C S. pastorianus Erlenmeyer 0.4 g of ethanol/g of Wang et al.
pretreatment 450 °C in an with H2SO4 (0.5 M) for ATCC 2345 flasks glucose; 79% yield (2012)
auger 240 min + activated carbon
reactor
Waste cotton None Pyrolysis at H2SO4 (0.2 M) hydrolysis + Ba(OH)2 S. cerevisiae Stirred 89% theoretical yield; Islam et al.
388 °C in a neutralization + ethyl acetate extraction 2.399 fermenter 14.78 g/L of ethanol (2018)
stainless‐ + activated carbon
steel tube
furnace
c13.indd 292 18-11-2019 14:59:09

Cotton None Pyrolysis at Water extraction + H2SO4 hydrolysis S. cerevisiae Flask 0.44 g ethanol/g Yu and
cellulose 388 °C in a + Ca(OH)2 neutralization + filtration 2.399 glucose Zhang
stainless‐ (0.45 μm membrane) + different (2004)
steel tube nitrogen sources
furnace
Waste cotton None Pyrolysis in H2SO4 hydrolysis + Ba(OH)2 Escherichia coli Fermenter 0.41 g ethanol/g Chang et al.
a tubular neutralization + ethyl ACCC 11177 glucose (2015)
furnace acetate extraction
reactor
Red oak None Pyrolysis at Water extraction + overliming with E. coli Flask 0.9 g/L ethanol Rover et al.
wood 450–500 °C NaOH (2014)
in a
fluidized‐
bed reactor
Pure None None None Engineered E. Flask 0.35 g ethanol Layton
levoglucosan coli KO11 produced/g LGA et al. (2011)
Red oak None Pyrolysis at Fractionating bio‐oil + water extraction Engineered E. Flask 0.24 g ethanol/g LGA Chi et al.
wood 450–500 °C + Ca(OH)2 treatment coli KO11 (2013)
in fluidized‐
bed reactor
LGA, levoglucosan.
c13.indd 293 18-11-2019 14:59:09

Table 13.3 Fermentation processes using levoglucosan as the carbon source to attain several important bioproducts.
Pyrolysis Fermentation Fermentation

Feedstock Pretreatment conditions Treatments Strain process product Product yield Reference
Cotton None Pyrolysis at None Aspergillus Flask Citric acid 87.5% of citric acid Zhuang et al.
cellulose 388 °C in a niger‐CBX‐209 (2001)
stainless‐
steel tube
furnace
Levoglucosan None None None Engineered Flask Styrene 0.24 g/L styrene; Xiong et al.
Escherichia 0.021 g/g LG (2016)
coli
Pure None None None Rhodotorula Flask Lipid R. glutinis: 2.7 g Lian et al.
levoglucosan glutinis lipid/L medium (2013)
(20 g/L)
Douglas fir Acid wash Pyrolysis at LGA extracted with ethyl R. glutinis Flask Lipid 0.78 g lipid/ L Lian et al.
500 °C in a acetate/biodiesel blends + medium (20 g/L) (2013)
fluidized‐ hydrolysis with H2SO4 (0.5 M) at
bed reactor 120 °C/240 min + neutralization
with Ba(OH)2 + detoxification
with activated carbon
Poplar wood Acid wash Pyrolysis at LGA extracted with ethyl Cryptococcus Flask Lipid C. curvatus: 68% Lian et al.
500 °C in a acetate + hydrolysis with H2SO4 curvatus or R. lipid mass/cell (2010)
fluidized‐ (0.5 M) at glutinis mass (0.167 g
bed reactor 120 °C/240 min + neutralization lipids/g sugar); R.
with Ba(OH)2 + detoxification glutinis 46% lipid
with activated carbon mass/cell mass
(8.9 g lipid/100 g
glucose)
LGA, levoglucosan.
c13.indd 294 18-11-2019 14:59:09

13.3 Integrated Process for Thermochemical Biomass

Conversion
From the technologic point of view, an attractive alternative for Brazil would be the inte-
gration of processes, considering as a starting point an autonomous first‐generation etha-
nol production plant, generically represented in Figure 13.3, as this technology is well
established. In this process, the excess sugarcane bagasse can be used as raw material for a
thermochemical biomass conversion subprocess (Figure 13.4), in which fast pyrolysis for
biomass conversion is adopted.
Initially, the sugarcane bagasse is pretreated for the removal of AAEMs, its moisture
content is reduced and then it is subjected to milling before the pyrolysis step. The
pyrolysis reactor uses a technology developed in Canada (Pyrovac Process, Pyrovac Inc.,
Canada) and operates under vacuum conditions, using molten salts (HITEC) as a form
of indirect heating of the biomass inside it. However, different prototypes are under
intensive development seeking scale‐up for industrial application (Sharifzadeh et al.
2019). Thereafter, the biocoal produced can be used to generate synthesis gas (syngas),
which is required as a fuel to maintain the salts in their molten form. In addition, two
streams are also generated in the condensation steps of the pyrolytic vapors: the bio‐oil
stream in the first condenser and an aqueous fraction containing the short‐chain organic
C1–C4, very rich in acetic acid, in the second condenser (Shen et al. 2009; Westerhof
et al. 2011). The following steps are related to bio‐oil refining processes and the produc-
tion of bioproducts that add value to this technology. Figure 13.5 shows the refining
steps of the bio‐oil, from which the pyrolytic sugars are initially separated and then
bioethanol is obtained.
At this point, ethanol fermentation and distillation steps can be conducted at the autono-
mous ethanol plant without any technologic change, thus reducing investment costs.
13.4 Oligosaccharide Production from Biomass
Oligosaccharides are carbohydrates composed of 2–10 monomeric units of sugars bound

by glycosidic linkages (BeMiller and Huber 2010). Some oligosaccharides are called non-
digestible because they act in the body as prebiotics, that is, substances that are not
digested by digestive enzymes and are metabolized in the small intestine and colon by
beneficial bacteria such as lactobacilli and bifidobacteria. In addition, oligosaccharides
contribute to sensory characteristics, such as taste and texture, as well as stabilizing
foams and emulsions. In contrast to the biomass oligosaccharides, fructooligosaccha-
rides, inulin, isomaltooligosaccharides, polydextrose, lactulose, galactooligosaccharides,
and glycooligosaccharides are well known (Grajek et al. 2005; Otieno and Ahring 2012;
Al‐Sheraji et al. 2013; Zambelli et al. 2016; Panesar et al. 2018).
Recently, great interest has been shown in the production of oligosaccharides from
biomasses from various sources. For example, several authors reported oligosaccharide
production from olive‐pruning residues, rice straw, corn cob, sugarcane bagasse, and
peanut shells (Teng et al. 2010; Ruiz et al. 2017; Carvalho et al. 2018; Chen et al. 2018; Rico
et al. 2018). In general, the basic principle of obtaining these products consists of the
Sugarcane juice extraction stage
Sugarcane input
Sugarcane
bagasse Turbine for energy
cogeneration
Steam (480°C)
Electricity
Boiler for steam
generation
Fermentation stage Steam (128 °C)
Juice
pretreatment
Ethanol distillation stage Ethanol storage
Ethanol
(94%)
Figure 13.3 The conventional process for ethanol production from sugarcane (autonomous bioethanol plant).
Aqueous phase (C1-C4)
T = 353 K H2SO4 (1%)
P = 101.3 kPa T = 273 K
P = 101.3 kPa T- 01 05
17
Biomass
G - 01
01 P - 03 02
08
W - 01 Ground biomass
Wet biomass Size: 3–10 mm
50 wt%moisture PR - 01
Aqueous phase
P - 01 P - 02 (C1-C4) Pyrolytic gas
Biomass
03
T=
T= 16
P = 101.3 kPa
08 P = 101.3 kPa 14 P - 07
Air
T = 298 K T = 298 K
Pyrolytic T = 353 K C - 02 P=
P = 101.3 kPa vapors C - 01
17
P=
T = 773 K
22 Founded salt P= H2O
T = 843 K H2O T = 298 K
10 T = 298 K
B - 02 Salt
FH - 01 T = 273 K
P = 101.3 kPa E - 02
R - 01
Pyrolytic gas 23 E - 01
09 H2O
H2O P - 05 T=
Syngas T - 02 T=
(CO2, CO, N2, H2) P - 04
12
20
15
T=1073 K Crude bio-oil

P = 101.3 kPa P - 06 Biochar T = 353 K
T = 273 K P=101.3 kPa
P = 101.3 kPa
Biochar
Air 10 T = 773 K
T= 298 K P = 101.3 kPa
P=101.3 kPa
18
SC - 02
Ash
B - 03 19
R - 02
Figure 13.4 Flow diagram for biomass fast pyrolysis processing under vacuum operation. Symbols: B 01–03 Blowers; C 01–02 Condensers; E 01–02
Heat exchangers for bio-oil and aqueous phase; E 03–04 Heat exchangers for air; FH 01 Firer heater; G 01 Grinder; P 01–05 Centrifugal pumps; P 06
Positive displacement pump; P 07 Vacuum pump; PR 01 Presser; R 01 Vacuum pyrolysis reactor; R 02 Gasification reactor; SC 02 Screw conveyor;
T 01–03 Tanks; W 01 Biomass washer.
Microorganisms
Other substrates
(Nitrogenous and mineral sources)
H2O 36 Air
Ba(OH)2 35
37
Detoxified 30
33 H2O
H2O pyrolytic sugars T=
24 29 H2O
25
Pyrolytic sugars
T=
Crude bio-oil ST - 01 T = 303 K
T =298 K T - 01 pH = 5
P = 101.3 kPa H2O
T - 02 120 rpm
T= F - 01
15 38
H2O Inoculum
T = 277 K 31 (10–20%)
Substrates
CO2
Neutralized H2O 39
26 FT - 01 aqueous 40 41
28 phase
27 Pyrolytic 34
sugars
Aqueous phase Phenols
P - 03
P - 01 (pyrolytic sugars) P - 02 FT - 02
32
Bio-oil Precipitated
salts
42
F - 02
CT - 01 Yeast biomass
(Single cell protein)
Fermentation broth
(8–10% ethanol content)
for distillation in the
Autonomous plant
Figure 13.5 Flow diagram for bio-oil processing to pyrolytic sugars extraction. Symbols: CT 01 Centrifuge; F 01–02 Fermenters; FT 01–02 Filters;
P 01–02; Centrifugal pumps; RB 01–03 Reboilers; ST 01 Sedimentation tank.
depolymerization of lignocellulosic materials, in which the hemicellulosic fraction is used

because it is more easily hydrolyzed than cellulose (Otieno and Ahring 2012). In addition,
depending on the source, they can be classified as xylooligosaccharides, arabinooligosac-
charides, and mannooligosaccharides because they are formed from xylans, arabinans,
and mannan, respectively.
Xylooligosaccharides have great importance because xylose is present in greater quantity
in several biomasses. Thus, raw materials with more than 20% of xylans have great poten-
tial for production of xylooligosaccharides (Otieno and Ahring 2012). These compounds
have several applications, among them foods, which has aroused great interest due to their
properties and health benefits, in some cases in a similar way to oligosaccharides available
in the market (Vásquez et al. 2000).
The xylooligosaccharides present a sweet taste and low caloric value, and are odorless.
Some of their benefits are related to their stability in a wide range of pH and tempera-
tures, selective metabolism by bifidobacteria, increased production of volatile fatty
acids, contribution to the reduction of stomach ulcer lesions (Parajó et al. 2004), and
reduction of glucose and blood cholesterol levels and procarcinogenic enzymes. In the
gastrointestinal tract, they increase the absorption of minerals by the large intestine
(Samanta et al. 2015).
There are several physical, chemical, and biochemical methods that can be used for oli-
gosaccharide production from biomass, such as ultrasound, steam explosion, autohydroly-
sis, enzymatic and microbial, and thermochemical (Kawamoto et al. 2003; Otieno and
Ahring 2012; Álvarez et al. 2017; Liu et al. 2018; Zhang et al. 2018b). The advantage of
using any method depends on the type of biomass and the parameters employed resulting
in greater or lesser efficiency in the depolymerization, for example xylans and consequent
formation of oligosaccharides and xylose. In addition, according to Otieno and Ahring
(2012), combination methods, such as the thermochemical transformation of biomass
followed by enzymatic hydrolysis, are reported to obtain high yields of oligosaccharides
because the enzyme has easier access to the substrates.
However, the formation of undesirable compounds such as furfural and hydroxymethyl-
furfural, among others, can inhibit the enzymatic activity of xylanases. Some studies have
also reported the production of oligosaccharides by pyrolysis of monosaccharides. However,
Liu et al. (2018) have argued that such formation occurs by the respective monosaccharides
with intermolecular dehydration followed by a condensation reaction under thermochemi-
cal treatment.
13.5 Application of Biochar Obtained from Biomass

Pyrolysis
Biochar is a carbon‐rich (65–90%) solid product obtained from the thermochemical
conversion of biomass at temperatures below 900 °C, under oxygen‐free conditions
(Xiong et al. 2017; Zhang et al. 2019). Biochar yield as a function of the thermochemi-
cal process used can vary as follows: 50–80% in torrefaction > 20–50% pyrolysis assisted
by microwave > around 35% for slow pyrolysis > about 12% in fast pyrolysis > typically
about 10% in gasification (Qambrani et al. 2017; Zhu et al. 2018; Mutsengerere et al.
2019). Some authors distinguish biochar from hydrochar, which results from
hydrothermal carbonization, and from other biocarbons, restricting the definition of
biochar only to that obtained by pyrolysis (Kambo and Dutta 2015; Liu et al. 2015;
Zhang et al. 2019).
Although biochar is mostly composed of carbon and ash, its overall elemental composi-
tion and properties are highly variable. Usually, the carbon content is 45–60 wt%, the
hydrogen content 2–5 wt%, and the oxygen content about 10–20% (Muhammad et al. 2018).
Compared with other typical amorphous carbon materials (Table 13.4), biochar usually has
abundant surface functional groups (C─O, C═O, COOH, OH, etc.), which being highly
modifiable act as a platform for the synthesis of various functionalized carbon materials
(Liu et al. 2015; Xiong et al. 2017).
The biochar quality can be greatly affected by conditions and circumstances such as
feedstock materials, catalysts, reaction conditions, reactor types, temperature, pressure,
additives, hot vapor residence time, and solids residence time (Kambo and Dutta 2015;
Muhammad et al. 2018; Zhu et al. 2018; Zhang et al. 2019). For example, slow pyrolysis,
in general, tends to produce biochar with greater nitrogen, sulfur, available phosphorus,
calcium, magnesium, surface area, and catalytic exchange capacity than fast pyrolysis
(Lehmann and Joseph 2015). High heating rates are characterized by biochar with high
oxygen content and low calorific value, probably as a result of the relatively short particle
residence time. High temperatures tend to produce greater concentrations of condensed
aromatic carbon, while lower process temperatures may produce biopolymers (Duman
et al. 2011). According to Bridgwater (2018), depending on the reactor configuration and
gas velocities, a large part of the char will be of a comparable size and shape as the bio-
mass feed.
Table 13.4 Differences between biochar, activated carbon, and carbon black.
Char Carbon
type Main source Preparation method content Structure
Biochar Biomass Pyrolysis of the biomass at 40–90% Amorphous carbon

medium temperature with abundant
(400–600 °C) and then surface functional
functionalization with groups,
physical or chemical methods nanostructures, or
porosity
Activated Coal, asphalt, Carbonization of the coal, 80–95% Amorphous carbon
carbon biomass, etc. asphalt, or biomass at high with abundant
temperature (700–1000 °C) porosity
under physical or chemical
activation
Carbon Petroleum, Combustion of petroleum, >95% Microcrystal or
black coal tar, coal tar, or asphalt under air‐ amorphous carbon
asphalt, etc. poor conditions particles
In pyrolysis, biochar acts as a vapor cracking catalyst and reduces liquid yield (~20%);
for this reason, fast and effective removal is essential. Liquid filtration is very difficult
due to the nature of char and pyrolytic lignin, so typically, cyclones are used with fines
passing through and collected in the liquid product. Hot vapor filtration is another
separation process that gives a high‐quality char‐free product from bio‐oil (Sharifzadeh
et al. 2019). It requires careful handling and storage of the fresh biochar because of its
pyrophoric characteristic of spontaneously combusting when exposed to air. This prop-
erty decreases with time due to oxidation of active sites on the char surface (Bridgwater
2018).
The physicochemical characterization of biochar, by a range of techniques, indicates
beneficial characteristics such as high surface‐to‐volume ratios, porosity, easily tuned sur-
face functionality by its abundant surface functional groups and minerals such as nitrogen,
sulfur, magnesium, phosphorus, calcium, and potassium, as well as strong affinity for non-
polar substances such as furans, dioxins, polycyclic aromatic hydrocarbons, and other com-
pounds (Kan et al. 2016; Muhammad et al. 2018; Zhu et al. 2018). In addition, it is known
that biochar has adsorption potential for toxic substances, such as manganese and alu-
minum in acidic soils, and arsenic, nickel, copper, cadmium, and lead in heavy metal‐
contaminated soils (Rizwan et al. 2016; Ho et al. 2017).
Other important properties of biochar include chemical stability, cost‐effectiveness,
water‐retaining capacities and retention of soil nutrients and microbial accumulation
(Ahmad et al. 2014; Xie et al. 2015; Shaheen et al. 2019). It can also be used as a fertilizer
while simultaneously acting as a carbon sequestration medium (Qambrani et al. 2017;
Muhammad et al. 2018), or as a feedstock for the production of activated carbon and other
industrial catalysts (Azargohar and Dalai 2006; Xiong et al. 2017). However, biochar
generally contains various heavy metals and other contaminants, depending on the types
of feedstock and methods used for its production, which carries risk during its application
(Hilber et al. 2017; Zhang et al. 2018a).
The rapid growth in publications on biochar since 2008 is a consequence of the
above‐mentioned characteristics that make biochar a promising platform for the syn-
thesis of many other functional materials and has attracted a great deal of attention
mainly in catalysis, energy storage, and environmental protection. Biochar production
technologies and their general application have been reviewed by many authors includ-
ing Laird et al. (2009), Meyer et al. (2011), Manyà (2012), Meier et al. (2013), Kambo
and Dutta (2015), Qian et al. (2015), Lee et al. (2017), Wang et al. (2018a), and Zhang
et al. (2019),
More specific reviews of biochar utilization for addressing several environmental man-
agement issues have been published by Ahmad et al. (2014), Hyland and Sarmah (2014),
Lehmann and Joseph (2015), Xie et al. (2015), Rizwan et al. (2016), Qambrani et al. (2017),
Muhammad et al. (2018), Zhang et al. (2018a), and Shaheen et al. (2019). Inyang et al.
(2016) and Ho et al. (2017) reviewed the use of biochar in heavy metals removal, while
Elkhalifa et al. (2019) reviewed the use of food wastes to produce biochar. Biofuel produc-
tion using biochar has been reviewed by Xiong et al. (2017), Perkins et al. (2018), and Zhu
et al. (2018). Table 13.5 summarizes some studies that use biochar as a catalyst in transes-
terification and esterification reactions.
Table 13.5 Biochar as a catalyst in transesterification and esterification reactions.
Surface area
(m2/g1)/
pore volume Catalysis recycling and
Biochar origin Catalysis activation (cm3/g) Reactants, reaction conditions, and yields reuse Reference
A mixture of wood KOH and carbonization at 640–990 Canola oil and oleic acid with methanol, 3 h, Successfully with a Yu et al. (2011),
waste, white wood, 675 °C and sulfonation with /0.35–0.90 150 °C, 1.52 MPa. Yield up to 44% slight decrease in the Dehkhoda and
bark, and shavings H2SO4 (>20 wt% free SO3) for reaction yield Ellis (2013)
15 h at 150 °C
Karanja (Pongamia None 0.015–0.027 120 °C, glycerol: acetic acid: 1:5, catalyst Can be reused with Rafi et al. (2015)
pinnata) seed shells /3.877–7.963 weight: 0.2 g. consistent activity
Conversion 83–85.5%
Peat KOH solution, mixed with 48.38–73.25 Palm oil and methanol, 65 °C Slight deactivation Wang et al.
CaO impregnating solution, at /39.51–90.58 Yield 90.1–93.4 due to leaching of (2019)
600 °C and 800 °C Ca2+
Rice husk Sulfonation with H2SO4 4.0 Simultaneous esterification and Li et al. (2014)
/7.7 transesterification of waste cooking oil. Yield:
87.57% after 15 h
Palm kernel shell Calcined at 800 °C for 2 h Sunflower oil: methanol (9:1), 3 wt% catalyst, Reused three times Kostic et al.
65 °C. Best FAMEs content of 99% with little drop in (2016)
catalytic activity
Pine chips Sulfonation with H2SO4. 365 Methanol and palmitic and stearic acids, Remained >90% after Kastner et al.
Heating at 100 °C for 12–18 h /0.2 spiked soybean oil or rendered poultry fat 6 cycles (2012)
after decantation (37:1 and 54 : 1). 2 h, 65 °C, 3 and 6 wt%
catalyst, -90–100% conversion
Douglas fir wood Sulfonation with H2SO4 for 3.51 Methanol: refined microalgal oil and FFAs Reused for 10 cycles Dong et al.
chips 24 h at 150 °C produced by hydrolysis of Chlorella without a dramatic (2015b)
sorokiniana refined oil (5:1–30:1), 3–7 wt% loss in activity
catalyst, 80–120 °C, 10–90 min
FAMEs, fatty acid methyl esters; FFA, free fatty acids.
c13.indd 302 18-11-2019 14:59:12

13.6 Conclusion
Bioethanol is conventionally produced in Brazil (first‐generation) on a large scale by using

biotechnologic methods. Around 33 056.44 × 103 L of both anhydrous and hydrated ethanol
were produced in 2018. However, Brazil still has a large potential for bioenergy production
derived from biomass and, in particular, thermochemical conversion of sugarcane bagasse
produces a high yield of pyrolytic sugars that can be used as raw materials to increase bioeth-
anol production by fermentation. Therefore, further efforts should be made to implement at
industrial scale an integrated process of thermochemical conversion of sugarcane bagasse to
autonomous bioethanol and to evaluate the technoeconomic and environmental impact of
this technology.
The debate about the use of land for energy production versus food may be increasingly
attenuated as new biomass that is not competing with the food chain begins to gain more
attention in biofuels production. Thus, the expectation of using sugarcane bagasse contin-
ues to be of great relevance as a raw material to obtain more added‐value products like
biochar, bio‐oil, chemicals, pyrolytic sugars, and oligosaccharides for the food industry,
among others. However, more studies should be carried out to enhance these processes and
seek their further implementation on an industrial scale.
Acknowledgments
We are grateful to the Natural Sciences Graduate Program for the Postdoctoral Position
Grants (No. 001/2018) and Plant Production Program, both from the State University of
Northern Rio de Janeiro, Coordination for the Improvement of Higher‐Level Personnel‐
Brazil (CAPES Finance Code 001), National Council for Scientific and Technological
Development (CNPq – Process No. 433235/2016‐0), Grants Program of the Estácio
University for Research Productivity and Foundation for Research and Innovation Support
of the Espírito Santo (FAPES, Process No. 355/2018) for financial support.
R
eferences
Ahmad, M., Upamali, A., Eun, J. et al. (2014). Biochar as a sorbent for contaminant
management in soil and water: a review. Chemosphere 99: 19–33.
Al‐Sheraji, S.H., Ismail, A., Manap, M.Y. et al. (2013). Prebiotics as functional foods: a review.
Journal of Functional Foods 5: 1542–1553.
Álvarez, C., González, A., Negro, M.J. et al. (2017). Optimized use of hemicellulose within a
biorefinery for processing high value‐added xylooligosaccharides. Industrial Crops and
Products 99: 41–48.
Asadullah, M., Rahman, M.A., Ali, M.M. et al. (2007). Production of bio‐oil from fixed bed
pyrolysis of bagasse. Fuel 86: 2514–2520.
Azargohar, R. and Dalai, A.K. (2006). Biochar as a precursor of activated carbon. Applied
Biochemistry and Biotechnology 129: 762–773.
Bacik, J.P. and Jarboe, L.R. (2016). Bioconversion of anhydrosugars: emerging concepts and
strategies. IUBMB Life 68: 700–708.
Bacik, J.P., Klesmith, J.R., Whitehead, T.A. et al. (2015). Producing glucose 6‐phosphate from
cellulosic biomass: structural insights into levoglucosan bioconversion. Journal of Biological
Chemistry 290: 26638–26648.
BeMiller, J.N. and Huber, K.C. (2010). Caiboidratos. In: Química de alimentos de Fennema, 4e
(eds. S. Damodaran, K.L. Parkin and O.R. Fennema). Porto Alegre, Brazil: Artmed Publisher.
Bennett, N.M., Helle, S.S., and Duff, S.J.B. (2009). Extraction and hydrolysis of levoglucosan
from pyrolysis oil. Bioresource Technology 100: 6059–6063.
Bridgwater, A.V. (2003). Renewable fuels and chemicals by thermal processing of biomass.
Chemical Engineering Journal 91: 87–102.
and Bioenergy 38: 68–94.
Bridgwater, B.T. (2018). Challenges and opportunities in fast pyrolysis of biomass: part I.
Johnson Matthey Technology Review 62: 118–130.
Carvalho, A.F.A., Marcondes, W.F., de Oliva Neto, P. et al. (2018). The potential of tailoring the
conditions of steam explosion to produce xylo‐oligosaccharides from sugarcane bagasse.
Chan, J.K.S. and Duff, S.J.B. (2010). Methods for mitigation of bio‐oil extract toxicity.
Chang, D., Yu, Z., Islam, Z.U., and Zhang, H. (2015). Mathematical modeling of the
fermentation of acid‐hydrolyzed pyrolytic sugars to ethanol by the engineered strain
Escherichia coli ACCC 11177. Applied Microbiology and Biotechnology 99: 4093–4105.
Chen, X., Li, H., Sun, S. et al. (2018). Co‐production of oligosaccharides and fermentable sugar
from wheat straw by hydrothermal pretreatment combined with alkaline ethanol extraction.
Chi, Z., Rover, M., Jun, E. et al. (2013). Overliming detoxification of pyrolytic sugar syrup for
direct fermentation of levoglucosan to ethanol. Bioresource Technology 150: 220–227.
Collard, F.X. and Blin, J. (2014). A review on pyrolysis of biomass constituents: mechanisms
and composition of the products obtained from the conversion of cellulose, hemicelluloses
and lignin. Renewable and Sustainable Energy Reviews 38: 594–608.
Czernik, S. and Bridgwater, A.V. (2004). Overview of applications of biomass fast pyrolysis oil.
Energy and Fuels 18: 590–598.
Dai, J., Yu, Z., He, Y. et al. (2009). Cloning of a novel levoglucosan kinase gene from Lipomyces
starkeyi and its expression in Escherichia coli. World Journal of Microbiology and
David, G.F., Perez, V.H., Rodriguez Justo, O., and Garcia‐Perez, M. (2017). Effect of acid
additives on sugarcane bagasse pyrolysis: production of high yields of sugars. Bioresource
David, G.F., Justo, O.R., Perez, V.H., and Garcia‐Perez, M. (2018). Thermochemical conversion
of sugarcane bagasse by fast pyrolysis: high yield of levoglucosan production. Journal of
Analytical and Applied Pyrolysis 133: 246–253.
David, G.F., Rios‐Rios, A.M., de Fatima, A. et al. (2019). The use of p‐sulfonic acid calix [4]
arene as organocatalyst for pretreatment of sugarcane bagasse increased the production of
levoglucosan. Industrial Crops & Products 134: 382–387.
Dehkhoda, A.M. and Ellis, N. (2013). Biochar‐based catalyst for simultaneous reactions of
esterification and transesterification. Catalysis Today 207: 86–92.
Dobele, G., Meier, D., Faix, O. et al. (2001). Volatile products of catalytic flash pyrolysis of
celluloses. Journal of Analytical and Applied Pyrolysis 58: 453–463.
Dobele, G., Rossinskaja, G., Dizhbite, T. et al. (2005). Application of catalysts for obtaining 1,
6‐anhydrosaccharides from cellulose and wood by fast pyrolysis. Journal of Analytical and
Applied Pyrolysis 74 (1–2): 401–405.
Dong, T., Gao, D., Miao, C. et al. (2015a). Two‐step microalgal biodiesel production using acidic
catalyst generated from pyrolysis‐derived bio‐char. Energy Conversion and Management 105:
1389–1396.
Dong, Q., Zhang, S., Zhang, L. et al. (2015b). Effects of four types of dilute acid washing on
moso bamboo pyrolysis using Py‐GC/MS. Bioresource Technology 185: 62–69.
Duman, G., Okutucu, C., Ucar, S. et al. (2011). The slow and fast pyrolysis of cherry seed.
Elkhalifa, S., Al‐ansari, T., Mackey, H.R., and Mckay, G. (2019). Food waste to biochars through
pyrolysis: a review. Resources, Conservation & Recycling 144: 310–320.
Eom, I.Y., Kim, J.Y., Kim, T.S. et al. (2012). Effect of essential inorganic metals on primary
thermal degradation of lignocellulosic biomass. Bioresource Technology 104: 687–694.
García‐Peréz, M., Chaala, A., and Roy, C. (2002). Vacuum pyrolysis of sugarcane bagasse.
Journal of Analytical and Applied Pyrolysis 65: 111–136.
Grajek, W., Olejnik, A., and Sip, A. (2005). Probiotics, prebiotics and antioxidants as functional
foods. Acta Biochimica Polonica 52: 665–671.
Haddad, K., Jeguirim, M., Jellali, S. et al. (2017). Combined NMR structural characterization
and thermogravimetric analyses for the assessment of the AAEM effect during
lignocellulosic biomass pyrolysis. Energy 134: 10–23.
Hameed, S., Sharma, A., Pareek, V. et al. (2019). A review on biomass pyrolysis models: kinetic,
network and mechanistic models. Biomass and Bioenergy 123: 104–122.
Hassan, E.B.M., Steele, P.H., and Ingram, L. (2009). Characterization of fast pyrolysis bio‐
oils produced from pretreated pine wood. Applied Biochemistry and Biotechnology 154:
182–192.
Hilber, I., Bastos, A.C., Loureiro, S. et al. (2017). The different faces of biochar: contamination
risk versus remediation tool. Journal of Environmental Engineering and Landscape
Management 6897: 86–104.
Ho, S., Zhu, S., and Chang, J. (2017). Recent advances in nanoscale‐metal assisted biochar derived
from waste biomass used for heavy metals removal. Bioresource Technology 246: 123–134.
Hyland, C. and Sarmah, A.K. (2014). Advances and innovations in biochar production and
utilization for improving environmental quality. In: Bioenergy Research: Advances and
Applications (eds. V.K. Gupta, M.G.T.P. Kubicek and J.S. Xu), 435–446. Amsterdam, The
Netherlands: Elsevier.
Inyang, M.I., Gao, B., Yao, Y. et al. (2016). A review of biochar as a low‐cost adsorbent for
aqueous heavy metal removal. Critical Reviews in Environmental Science and Technology 46:
406–433.
Islam, Z.U., Zhang, H., Yu, Z. et al. (2018). Fermentation of detoxified acid‐hydrolyzed
pyrolytic anhydrosugars into bioethanol with Saccharomyces cerevisiae 2399. Applied
Biochemistry and Microbiology 54: 58–70.
Jarboe, L.R., Wen, Z., Choi, D., and Brown, R.C. (2011). Hybrid thermochemical processing:
fermentation of pyrolysis‐derived bio‐oil. Applied Microbiology and Biotechnology 91:
1519–1523.
Jiang, L.Q., Fang, Z., Zhao, Z.L. et al. (2019). Levoglucosan and its hydrolysates via fast
pyrolysis of lignocellulose for microbial biofuels: a state‐of‐the‐art review. Renewable and
Johnson, R.L., Liaw, S.S., Garcia‐Perez, M. et al. (2009). Pyrolysis gas chromatography mass
spectrometry studies to evaluate high‐temperature aqueous pretreatment as a way to
modify the composition of bio‐oil from fast pyrolysis of wheat straw. Energy and Fuels 23:
6242–6252.
Kambo, H.S. and Dutta, A. (2015). A comparative review of biochar and hydrochar in terms of
production, physico‐chemical properties and applications. Renewable and Sustainable
Kan, T., Strezov, V., and Evans, T.J. (2016). Lignocellulosic biomass pyrolysis: a review of
product properties and effects of pyrolysis parameters. Renewable and Sustainable Energy
Reviews 57: 1126–1140.
Kastner, J.R., Miller, J., Geller, D.P. et al. (2012). Catalytic esterification of fatty acids using solid
acid catalysts generated from biochar and activated carbon. Catalysis Today 190: 122–132.
Kawamoto, H., Murayama, M., and Saka, S. (2003). Pyrolysis behavior of levoglucosan as an
intermediate in cellulose pyrolysis: polymerization into polysaccharide as a key reaction to
carbonized product formation. Journal of Wood Science 49: 469–473.
Kostic, M.D., Bazargan, A., Stamenkovic, O.S. et al. (2016). Optimization and kinetics of
sunflower oil methanolysis catalyzed by calcium oxide‐based catalyst derived from palm
kernel shell biochar. Fuel 163: 304–313.
Laird, D.A., Brown, R.C., Amonette, J.E., and Lehmann, J. (2009). Review of the pyrolysis
platform for coproducing bio‐oil and biochar. Biofuels Bioproducts & Biorefining 3: 547–562.
Layton, D.S., Ajjarapu, A., Choi, D.W., and Jarboe, L.R. (2011). Engineering ethanologenic
Escherichia coli for levoglucosan utilization. Bioresource Technology 102: 8318–8322.
Lee, J., Kim, K., and Kwon, E.E. (2017). Biochar as a catalyst. Renewable and Sustainable
Lehmann, J. and Joseph, S. (2015). Biochar for Environmental Management: Science, Technology
and Implementation, 2e. Oxford, UK: Routledge.
Li, M., Zheng, Y., Chen, Y., and Zhu, X. (2014). Biodiesel production from waste cooking oil
using a heterogeneous catalyst from pyrolyzed rice husk. Bioresource Technology 154:
345–348.
Li, Q., Steele, P., Mitchell, B. et al. (2013). The addition of water to extract maximum
levoglucosan from the bio‐oil produced via fast pyrolysis of pretreated loblolly pinewood.
BioResources 8: 1868–1880.
Lian, J., Chen, S., Zhou, S. et al. (2010). Separation, hydrolysis and fermentation of pyrolytic
sugars to produce ethanol and lipids. Bioresource Technology 101: 9688–9699.
Lian, J., Garcia‐Perez, M., and Chen, S. (2013). Fermentation of levoglucosan with oleaginous
yeasts for lipid production. Bioresource Technology 133: 183–189.
Liang, J., Lin, Y., Wu, S. et al. (2015). Enhancing the quality of bio‐oil and selectivity of phenols
compounds from pyrolysis of anaerobic digested rice straw. Bioresource Technology 181:
220–223.
Liu, W., Jiang, H., and Yu, H. (2015). Development of biochar‐based functional materials:
toward a sustainable platform. Carbon Material Chemical Reviews 115: 12251–12285.
Liu, X., Wei, W., Wu, S. et al. (2018). A promptly approach from monosaccharides of biomass
to oligosaccharides via sharp‐quenching thermo conversion SQTC. Carbohydrate Polymers
189: 204–209.
Lu, Q., Hu, B., Zhang, Z. et al. (2018). Mechanism of cellulose fast pyrolysis: the role of
characteristic chain ends and dehydrated units. Combustion and Flame 198: 267–277.
Luque, L., Westerhof, R., Rossum, G.V. et al. (2014). Pyrolysis based bio‐refinery for the
production of bioethanol from demineralized lignocellulosic biomass. Bioresource
Madadi, M. and Hin, P. (2017). Lignin degradation by fungal pretreatment: a review. Journal of
Plant Pathology & Microbiology 8: 1–6.
Manyà, J.J. (2012). Pyrolysis for biochar purposes: a review to establish current knowledge gaps
and research needs. Environmental Science & Technology 46: 7939–7954.
Meier, D. and Faix, O. (1999). State of the art of applied fast pyrolysis of lignocellulosic
materials – a review. Bioresource Technology 68: 71–77.
Meier, D., van de Beld, B., Bridgwater, A.V. et al. (2013). State‐of‐the‐art of fast pyrolysis in IEA
bioenergy member countries. Renewable and Sustainable Energy Reviews 20: 619–641.
Meyer, S., Glaser, B., and Quicker, P. (2011). Technical economical and climate‐related aspects
of biochar production technologies: a literature review. Environmental Science & Technology
45: 9473–9483.
Montoya, J.I., Valdés, C., Chejne, F. et al. (2015). Bio‐oil production from Colombian bagasse
by fast pyrolysis in a fluidized bed: an experimental study. Journal of Analytical and Applied
Pyrolysis 112: 379–387.
Mourant, D., Wang, Z., He, M. et al. (2011). Mallee wood fast pyrolysis: effects of alkali
and alkaline earth metallic species on the yield and composition of bio‐oil. Fuel 90:
2915–2922.
Muhammad, N., Hussain, M., Ullah, W. et al. (2018). Biochar for sustainable soil and
environment: a comprehensive review. Arabian Journal of Geosciences 11: 1–14.
Mutsengerere, S., Chihobo, C.H., Musademba, D., and Nhapi, I. (2019). A review of operating
parameters affecting bio‐oil yield in microwave pyrolysis of lignocellulosic biomass.
Neto, J.M., da Oliveira, L.S.C., da Silva, F.L.H. et al. (2019). Use of sweet sorghum bagasse
Sorghum bicolor L moench for cellulose acetate synthesis. Bioresources 8: 3534–3553.
Ning, J., Yu, Z., Xie, H. et al. (2007). Purification and characterization of levoglucosan
kinase from Lipomyces starkeyi YZ‐215. World Journal of Microbiology and Biotechnology
24: 15–22.
Oasmaa, A., Fonts, I., Pelaez‐Samaniego, M.R. et al. (2016). Pyrolysis oil multiphase behavior
and phase stability: a review. Energy and Fuels 30: 6179–6200.
Ortiz, A.P., Segura, F.J.N., Jabalera, R.S. et al. (2013). Low temperature sugar cane bagasse
pyrolysis for the production of high purity hydrogen through steam reforming and CO2
capture. International Journal of Hydrogen Energy 38: 12580–12588.
Otieno, D.O. and Ahring, B.K. (2012). The potential for oligosaccharide production from the
hemicellulose fraction of biomasses through pretreatment processes: xylooligosaccharides
XOS, arabinooligosaccharides AOS, and mannooligosaccharides MOS. Carbohydrate

Research 360: 84–92.
Oudenhoven, S.R.G., Westerhof, R.J.M., Aldenkamp, N. et al. (2013). Demineralization of
wood using wood‐derived acid: towards a selective pyrolysis process for fuel and chemicals
production. Journal of Analytical and Applied Pyrolysis 103: 112–118.
Oudenhoven, S.R.G., van der Ham, A.G.J., van den Berg, H. et al. (2016). Using pyrolytic acid
leaching as a pretreatment step in a biomass fast pyrolysis plant: process design and
economic evaluation. Biomass and Bioenergy 95: 388–404.
Panesar, P.S., Kaur, R., Singh, R.S., and Kennedy, J.F. (2018). Biocatalytic strategies in the
production of galacto‐oligosaccharides and its global status. International Journal of
Biological Macromolecules 111: 667–679.
Parajó, J.C., Garrote, G., Cruz, J.M., and Dominguez, H. (2004). Production of
xylooligosaccharides by autohidrolysis of lignocellulosic materials. Trends in Food Science &
Patwardhan, P.R., Satrio, J.A., Brown, R.C., and Shanks, B.H. (2010). Influence of inorganic
salts on the primary pyrolysis products of cellulose. Bioresource Technology 101: 4646–4655.
Pecha, B., Arauzo, P., and Garcia‐Perez, M. (2015). Impact of combined acid washing and acid
impregnation on the pyrolysis of Douglas fir wood. Journal of Analytical and Applied
Pyrolysis 114: 127–137.
Perkins, G., Bhaskar, T., and Konarova, M. (2018). Process development status of fast pyrolysis
technologies for the manufacture of renewable transport fuels from biomass. Renewable and
Qambrani, N.A., Rahman, M.M., Won, S. et al. (2017). Biochar properties and eco‐friendly
applications for climate change mitigation, waste management, and wastewater treatment: a
review. Renewable and Sustainable Energy Reviews 79: 255–273.
Qian, K., Kumar, A., Zhang, H. et al. (2015). Recent advances in utilization of biochar.
Rafi, J.M., Rajashekar, A., Srinivas, M. et al. (2015). Esterification of glycerol over a solid acid
biochar catalyst derived from waste biomass. RSC Advances 5: 44550–44556.
Rico, X., Gullón, B., Alonso, J.L. et al. (2018). Valorization of peanut shells: manufacture of
bioactive oligosaccharides. Carbohydrate Polymers 183: 21–28.
Rizwan, M., Ali, S., Qayyum, M.F. et al. (2016). Mechanisms of biochar‐mediated alleviation of
toxicity of trace elements in plants: a critical review. Environmental Science and Pollution
Research 23: 2230–2248.
Rover, M.R., Johnston, P.A., Jin, T. et al. (2014). Production of clean pyrolytic sugars for
fermentation. ChemSusChem 7: 1662–1668.
Roy, P. and Dias, G. (2017). Prospects for pyrolysis technologies in the bioenergy sector: a
review. Renewable and Sustainable Energy Reviews 77: 59–69.
Ruiz, E., Gullón, B., Moura, P. et al. (2017). Bifidobacterial growth stimulation by
oligosaccharides generated from olive tree pruning biomass. Carbohydrate Polymers
169: 149–156.
Samanta, A.K., Jayapal, N., Jayaram, C. et al. (2015). Xylooligosaccharides as prebiotics from
agricultural by‐products: production and applications. Bioactive Carbohydrates and Dietary
Fibre 5: 62–71.
Sarkar, N., Ghosh, S.K., Bannerjee, S., and Aikat, K. (2012). Bioethanol production from
agricultural wastes: an overview. Renewable Energy 37: 19–27.
Shaheen, S.M., Niazi, N.K., Hassan, N.E.E. et al. (2019). Wood‐based biochar for the removal of
potentially toxic elements in water and wastewater: a critical review. International Materials
Reviews 6608: 216–247.
Shahsavarani, H., Hasegawa, D., Yokota, D. et al. (2013). Enhanced bio‐ethanol production
from cellulosic materials by semi‐simultaneous saccharification and fermentation using
high temperature resistant Saccharomyces cerevisiae TJ14. Journal of Bioscience and
Bioengineering 115: 20–23.
Shaik, S.M., Sharratt, P.N., and Tan, R.B.H. (2013). Influence of selected mineral acids and
alkalis on cellulose pyrolysis pathways and anhydrosaccharide formation. Journal of
Sharifzadeh, M., Sadeqzadeh, M., Guo, M. et al. (2019). The multi‐scale challenges of biomass
fast pyrolysis and bio‐oil upgrading: review of the state of art and future research directions.
Progress in Energy and Combustion Science 71: 1–80.
Shen, J., Wang, X.‐S., Garcia‐Perez, M. et al. (2009). Effects of particle size on the fast pyrolysis
of oil mallee woody biomass. Fuel 88: 1810–1817.
Shen, D., Jin, W., Hu, J. et al. (2015). An overview on fast pyrolysis of the main constituents in
lignocellulosic biomass to valued‐added chemicals: structures, pathways and interactions.
Sukhbaatar, B., Li, Q., Wan, C. et al. (2014). Inhibitors removal from bio‐oil aqueous fraction
for increased ethanol production. Bioresource Technology 161: 379–384.
Tan, H. and Wang, S. (2009). Experimental study of the effect of acid‐washing pretreatment on
biomass pyrolysis. Journal of Fuel Chemistry and Technology 37: 668–672.
Teng, C., Yan, Q., Jiang, Z. et al. (2010). Production of xylooligosaccharides from the steam
explosion liquor of corncobs coupled with enzymatic hydrolysis using a thermostable
xylanase. Bioresource Technology 101: 7679–7682.
Tran, D.T., Pole, I.Y., and Lin, C.W. (2013). Developing co‐culture system of dominant
cellulolytic Bacillus sp THLA0409 and dominant ethanolic Klebsiella oxytoca THLC0409 for
enhancing ethanol production from lignocellulosic materials. Journal of the Taiwan Institute
of Chemical Engineers 44: 762–769.
Vazquez, M.J., Alonso, J.L., Domınguez, H., and Parajo, J.C. (2000). Xylooligosaccharides:
manufacture and applications. Trends in Food Science & Technology 1111: 387–393.
Wang, Z., Garcia‐Perez, M., O’Fallon, J. et al. (2010). Separation, hydrolysis and
fermentation of pyrolytic sugars to produce ethanol and lipids. Bioresource
Technology 101: 9688–9699.
Wang, Z., Lu, Q., Zhu, X.F., and Zhang, Y. (2011). Catalytic fast pyrolysis of cellulose to prepare
levoglucosenone using sulfated zirconia. ChemSusChem 4: 79–84.
Wang, H., Livingston, D., Srinivasan, R. et al. (2012). Detoxification and fermentation of
pyrolytic sugar for ethanol production. Applied Biochemistry and Biotechnology 168:
1568–1583.
Wang, J., Wei, Q., Zheng, J., and Zhu, M. (2016). Effect of pyrolysis conditions on levoglucosan
yield from cotton straw and optimization of levoglucosan extraction from bio‐oil. Journal of
Wang, B., Gao, B., and Fang, J. (2018a). Recent advances in engineered biochar productions
and applications. Critical Reviews in Environmental Science and Technology 3389:
2158–2207.
Wang, J.Q., Zheng, J.L., Wang, J.T., and Lu, Z.M. (2018b). A separation and quantification
method of levoglucosan in biomass pyrolysis. Industrial Crops and Products 113:
266–273.
Wang, S., Shan, R., Wang, Y. et al. (2019). Synthesis of calcium materials in biochar matrix as a
highly stable catalyst for biodiesel production. Renewable Energy 130: 41–49.
Westerhof, R.J.M., Brilman, D.W.F., Garcia‐Perez, M. et al. (2011). Fractional condensation of
biomass pyrolysis vapors. Energy and Fuels 25: 1817–1829.
Xie, T., Reddy, K.R., Wang, C. et al. (2015). Technology characteristics and applications of
biochar for environmental remediation: a review. Critical Reviews in Environmental Science
and Technology 3389: 939–969.
Xiong, X., Lian, J., Yu, X. et al. (2016). Engineering levoglucosan metabolic pathway in
Rhodococcus jostii RHA1 for lipid production. Journal of Industrial Microbiology and
Xiong, X., Yu, I.K.M., Cao, L. et al. (2017). A review of biochar‐based catalysts for
chemical synthesis, biofuel production, and pollution control. Bioresource Technology
246: 254–270.
Yang, X., Ma, F., Yu, H. et al. (2011). Effects of biopretreatment of corn stover with white‐
rot fungus on low‐temperature pyrolysis products. Bioresource Technology 102:
3498–3503.
Yu, Z. and Zhang, H. (2003). Pretreatments of cellulose pyrolysate for ethanol production by
Saccharomyces cerevisiae, Pichia sp YZ‐1 and Zymomonas mobilis. Biomass and Bioenergy 24:
257–262.
Yu, J.T., Dehkhoda, A.M., and Ellis, N. (2011). Development of biochar‐based catalyst for
transesterification of canola oil. Energy and Fuels 25: 337–344.
Yu, Z. and Zhang, H. (2004). Ethanol fermentation of acid‐hydrolyzed cellulosic pyrolysate
with Saccharomyces cerevisiae. Bioresource Technology 93: 199–204.
Zambelli, P., Tamborini, L., Cazzamalli, S. et al. (2016). An efficient continuous flow process
for the synthesis of a non‐conventional mixture of fructooligosaccharides. Food Chemistry
190: 607–613.
Zhang, X., Yang, W., and Dong, C. (2013). Levoglucosan formation mechanisms during
cellulose pyrolysis. Journal of Analytical and Applied Pyrolysis 104: 19–27.
Zhang, C., Liu, L., Zhao, M. et al. (2018a). The environmental characteristics and applications
of biochar. Environmental Science and Pollution Research 25: 21525–21534.
Zhang, W., You, Y., Lei, F. et al. (2018b). Acetyl‐assisted autohydrolysis of sugarcane bagasse
for the production of xylo‐oligosaccharides without additional chemicals. Bioresource
Zhang, Z., Zhu, Z., Shen, B., and Liu, L. (2019). Insights into biochar and hydrochar production
and applications: a review. Energy 171: 581–598.
Zheng, A., Zhao, K., Sun, J. et al. (2018). Effect of microwave‐assisted organosolv fractionation
on the chemical structure and decoupling pyrolysis behaviors of waste biomass. Journal of
Zhou, S., Wang, Z., Liaw, S.S. et al. (2013). Effect of sulfuric acid on the pyrolysis of Douglas fir
and hybrid poplar wood: Py‐GC/MS and TG studies. Journal of Analytical and Applied
Pyrolysis 104: 117–130.
Zhu, L., Lei, H., Zhang, Y. et al. (2018). A review of biochar derived from pyrolysis and its
application in biofuel production. SF Journal of Material and Chemical Engineering 1:
1–9.
Zhuang, X. and Zhang, H. (2002). Identification, characterization of levoglucosan kinase, and
cloning and expression of levoglucosan kinase cDNA from Aspergillus niger CBX‐209 in
Escherichia coli. Protein Expression and Purification 26: 71–81.
Zhuang, X.L., Zhang, H.X., Yang, J.Z., and Qi, H.Y. (2001). Preparation of levoglucosan by
pyrolysis of cellulose and its citric acid fermentation. Bioresource Technology 79: 63–66.
Zhuang, X., Wang, W., Yu, Q. et al. (2016). Liquid hot water pretreatment of lignocellulosic
biomass for bioethanol production accompanying with high valuable products. Bioresource
313
14
Life Cycle Analysis of Lignocellulosic Conversion into

Fuels, Energy, and Chemicals
Mahdi Mazuchi
Iranian Sugarcane and By-product Research and Training Institute, Ahvaz, Iran
14.1 Introduction
Environmental sustainability as well as value creation are the goals of process development
by producing energy and materials from various lignocellulosic residues. Major concerns
including declining fossil resources, excessive usage of gasoline, and increasing environ-
mental problems are increasing. Bioresources such as lignocellulosic biomass have a posi-
tive impact on aspects of environmental sustainability and economy (Vohra et al. 2014;
Chandel et al. 2018). According to Biddy et al. (2016), the biotech market share will increase
from 2% in 2016 to 22% in 2025. Based on the current scenario, commercialization should
lead from pilot research to a fully commercial level, which will reduce the capital and oper-
ational costs and also needs a details technoeconomic analysis (Chandel et al. 2019). Raw
materials in the first‐generation biorefinery are mainly crops, grains or juice and in the
second generation biorefinery are agro‐residues and forestry wastes. However, their judi-
cious utilization with a balance between food/feed and fuel should be assessed for the reali-
zation of biorefineries in the near future. However, rising food prices, energy security, and
land use changes should be addressed in legitimate manner (Menon and Rao 2012; Chandel
et al. 2019).
The use of residual biomass (which is not primarily for human consumption and cattle
feed) can be more feasible for second‐generation biorefineries. The usage of biomass
resources such as wheat straw, sugarcane straw, and rice straw which are considered as
food/ fodder should be evaluated properly. According to the analysis by ePURE (2017),
bioethanol from lignocellulosic material represents a 4% share, whil others such as corn,
wheat, sugar, and other grains produce 39%, 30%, 20%, and 7%, respectively. For example,
Granbio, a company in Brazil, has established a cellulosic ethanol production facility which
can produce 21 million gallons of cellulosic ethanol per year from bagasse and straw.
However, this amount is less than 1% of the total bioethanol production in Brazil (AFDC
2018; Granbio 2014).

A promising alternative to starch or sugar (first‐generation) biorefineries is lignocellulosic‐

based feedstock (second generation). Most of the ethanol present in the market is produced
from raw materials such as corn grains, sugarcane molasses, sugar beet, and other cereals,
because lignocellulosic materials have a favorable potential for biorefinery feed in terms of
low costs and availability in different climatic regions (Ghosh et al. 2015; Lamers et al.
2015; Chandel et al. 2018). The important sources of lignocellulosic biomass include forest
and wood residue, municipal solid waste (MSW), paper waste, and agricultural residues. It
has been proposed that use of lignocellulosic biomass could increase the production of
bioethanol by five times compared to current levels of bioethanol production. As a result,
second‐generation biofuels can minimize environmental problems (Stephen et al. 2012;
Lamers et al. 2015).
Life cycle assessment (LCA) is an important approach to studying the different materials,
products, and processes and their impacts on environmental sustainability. A life cycle
with respect to environmental indicators and mass and energy balances can measure the
cycle from feedstock to disposal of a product. Various assessment strategies including the
use of specialized LCA software and other simulations of the processes are necessary. As
far as solutions to optimize sustainability are concerned, integration can increase the effi-
ciency of materials and energy and reduce waste disposal. Here, we present critical infor-
mation on various biomass resources, cell wall components of second‐generation biomass,
and the basic elements of life cycle analysis of biorefineries.
14.2 Basic Components of Lignocellulosic Biomass
Lignocellulosic biomass mainly contains three types of biopolymers: cellulose, hemicellu-

lose, and lignin. Each of these polymers can be used for the production of different chemi-
cals, materials, and fuels (Ladisch et al. 2010).
14.2.1 Cellulose
Cellulose is a well‐known organic polymer with the molecular formula (C6H10O5)n. It is clas-
sified in the polysaccharide category with a linear chain of many glucose linkages. Cellulose
is a vital component of the primary cell wall of green plants and algae (Ladisch et al. 2010).
14.2.2 Hemicellulose
Hemicellulose is a polysaccharide composed of sugars and commonly found with cellulose
in almost all plant cell walls. Cellulose crystals are robust, while hemicellulose has a
random structure, which is weak and amorphous. For this reason, it is easily hydrolyzed.
Typical hemicelluloses include glucuronoxylan, arabinoxylan, glucomannan, and xyloglu-
cans (Cheng et al. 2008; Scheller and Ulvskov 2010). Hemicellulose is often accompanied
by cellulose, but their molecular structures are different. Hemicellulose consists of several
simple sugars, but cellulose has only glucose. In addition to glucose, hemicellulose con-
tains many sugar monomers including xylose, arabinose, rhamnose, galactose, and
mannose (Cheng et al. 2008; Scheller and Ulvskov 2010).
Life Cycle Analysis of Lignocellulosic Conversion into Fuels, Energy, and Chemicals 315
14.2.3 Lignin
Lignin surrounds the cellulose and hemicellulose to protect them. Lignin is a complex
combination of polymers connected to each other. Strong interaction of lignin with other
polymers makes the structure recalcitrant. In addition, the composition of lignin varies
from plant to plant.
Other components of lignocellulosic biomass have no specific structure and therefore do
not affect the initial treatment process. The composition of lignin, cellulose, and hemicel-
lulose varies from biomass to biomass, cultivation conditions, and geographical distribu-
tion. Reliable composition data of lignocellulosic biomass can help in accurate LCA
(Ohgren et al. 2007; Li et al. 2008).
14.3 Sources of Lignocellulosic Biomass
Lignocellulosic materials are the largest renewable biosources available. They include
agro‐residues such as sugarcane bagasse, corn stover, rice straw, wheat straw, sorghum
straw, forest residues like wood, hardwood, softwood, sawdust, prunings, and other MSWs.
Figure 14.1 represents the total percentage of lignocellulosic biomass obtained from vari-
ous sources (USDE 2009).
14.3.1 Forest Waste

There are two types of wood materials, commonly classified as softwood or hardwood.
Unlike agricultural biomass, these raw materials are flexible in harvesting time and have
no long storage time. Moreover, it has been observed that wood waste has higher lignin
content and very little ash content compared to agricultural residues. The special features
of wood biomass, such as high density, low ash, low transportation cost, make it a good
Biomass
4% 4% Agricultural Residues
6%
Forest Resources
31%
Energy Crops
28%
Grain and Corn

27%
Municipal and Industrial

Wastes
Others (i.e. oil seeds)
Figure 14.1 Annual total tonnage of biomass for biofuel. Source: Modified from USDE (2009).
choice for biorefinery. Other wastes like sawdust from sawmills, wood chips, and dead tree
parts can also be used as feedstock. However, differences in biomass composition analysis
should be considered in the process of LCA (Demirbas 2005; Hu et al. 2008).
14.3.2 Agricultureal and Urban Waste

The agricultural residues include different types of waste such as corn stover, rice straw,
wheat straw, sugarcane bagasse, sugarcane straw, etc. Hemicellulosic content is generally
higher in agriculture waste than in wood biomass (about 25–35%). However, agricultural
residues are a promising alternative for forestry biomass to avoid deforestation. Furthermore,
agricultural crops have a short life cycle and can be easily replaced which promotes sus-
tainable development (Demirbas 2005; Epplin et al. 2007; Hu et al. 2008).
Grass is another important source of lignocellulosic biomass because it is nonedible and
hence has no issues with food security and cost. Moreover, grass cultivation can be done
easily with low investment and maintenance cost and high yield. Various types of grasses
like Miscanthus giganteus and Switchgrass are fast‐growing feedstocks of lignocellulosic
biomass (Khanna et al. 2008). Raman and Gnansounou (2015) proposed that vetiver grass
could be used as replacement for wheat straw in a biorefinery. This grass particularly helps
in the reduction of CO2 and also improves soil condition.
In addition to cellulosic material, urban and domestic wastes can be used as raw material
in the production of various high‐value biorefining products. Utilization of such waste also
helps to solve problems associated with their disposal and ultimately in controlling envi-
ronmental pollution. Due to the cost of biomass and land limitation, optimal resource uti-
lization can improve environmental impact as well as economic condition. The use of
nonagricultural wasteland for the cultivation of nonedible crops can be an important solu-
tion for the supply of sufficient feedstock to biorefineries. According to USDOE (2003), if
lignocellulosic feedstocks are used then it may be possible to replace 30% of petroleum‐
based gasoline with biofuels by 2030.
14.3.3 Industrial Waste

Various industries, particularly the paper industries, are important sources of waste which
could be very useful for biorefineries. As a result, European pulp mills are developing
wood‐based biorefineries so that a wide range of valuable chemicals can be produced as
side‐streams from self‐generated wastes, such as black liquor (the residue of the pulp pro-
duction process). Lignosulfonate is an important by‐product which can be obtained via
ultrafiltration of black liquor. Similarly, the waste generated in these industries could be
used in the sustainable production of polymers such as sodium polyacrylate (Gonzalez‐
Garcıa et al. 2011; Gontia and Janssen 2016).
14.4 Biorefinery Products from Lignocellulosic Feedstock
The existing second‐generation biorefineries utilize various kinds of lignocellulosic biomass

to produce different forms of bioenergy like bioethanol and biogas and other high‐value
products which directly support the economy and environment health. However, there is
always scope for further development of attractive commercial processes for the efficient
conversion of these feedstocks to various basic chemicals like bio‐based polymers (biocom-
patible polymers). It is proposed that when the different products are obtained from such
materials, the entire value chain will be covered (Rajendran and Murthy 2017; Chandel
et al. 2019). A wide range of biorefining products can be produced using a variety of ligno-
cellulosic feedstocks (Chandel et al. 2019). Some of these products are discussed here.
14.4.1 Valeric Biofuels

It has been claimed that levulinic acid can be easily produced from lignocellulosic biomass
using a simple, cheap, and robust chemical catalysis process. Moreover, several derivatives
of levulinic acid have been proposed for fuel applications (Hayes et al. 2006). Lange et al.
(2010) demonstrated a process for the production of valeric biofuels; acid hydrolysis of
lignocellulosic materials was performed to produce levulinic acid followed by its hydro-
genation to produce valeric acid and finally its esterification to obtain alkyl (mono/di)
valerate esters. This derivative of valeric acid is recognized as valeric biofuel. Depending on
the ester used in a reaction, the fuel may be biogasoline or biodiesel and blend with cur-
rently used fuel. Also, biofuels can be modified unlike bioethanol which can be utilized in
modern cars without any engine modification.
14.4.2 Dimethyl Furan (DMF)

Dimethyl furan (DMF) is another biorefinery product which can be used as biofuel and a
new resource of energy. However, the impact on the environment, human health, hazards
and overall sustainability of these biofuels must be considered. Large‐scale production of
DMF is possible using lignocellulosic feedstocks which have higher energy content than
ethanol. Comparing DMF with ethanol, it has 40% higher energy content per molecule,
better octane number and hence can provide better combustion performance. Despite all
these benefits, there are some concerns about using DMF as a biofuel which are related to
human health and environmental pollution. These issues need to be addressed in LCA
analysis (Román‐Leshkov et al. 2007).
14.4.3 Bio-oil
Biodegradable oils are commonly known as bio‐oils, which are produced using pyrolysis.
Bio‐oils can be used as a fuel or raw material for the production of various other chemical
intermediates. Usually, the rapid pyrolysis process produces bio‐oil from lignocellulosic
biomasses. Unfortunately, the effect of low oil prices will inevitably lead to low market
value of biofuels (Bridgwater 2012). Moreover, technical and economic analysis of bio‐oil
production and its development pathway as a fuel have not been commercially evaluated
yet. Bio‐oils look profitable, but not attractive enough for investment.
The production of valuable chemicals from bio‐oil can improve its production process as
well as sustainability by reducing process wastes. A significant part of the bio‐oil is the frac-
tion which has phenolic compounds derived from lignin (also known as pyrolytic lignin).
Therefore, a fraction of bio‐oil can be used for bio‐based phenolic resin production. In this
approach, lignin usage can improve sustainability (Brown et al. 2011; Bridgwater 2012; Hsu
2012). Zheng et al. (2018) studied a technoeconomic approach for the production of levo-
glucosan, phenol formaldehyde resins, and noncorrosive bio‐oil from fast biomass pyroly-
sis. In addition to technical and economic studies, they also paid a lot of attention to LCA
and investigation its higher environmental impacts.
14.4.4 Higher Alcohols

Alcohols with a higher molecular weight such as n‐butanol and isobutanol have higher
energy content. They can be produced through thermochemical routes (such as synthesis
gas conversion into a mixture of alcohols) or biochemical pathways (e.g., fermentation)
using various biomass resources. Limited reports are available based on technical and eco-
nomic analysis of n‐butanol produced commercially. The success of the biofuel industry
not only depends on the economy but also on environmental sustainability as determined
by LCA. Among the various quantitative studies performed on production of biobutanol,
the majority are focused on the production of bio‐based biobutanol from corn instead of
cellulosic feedstock (Kumar and Gayen 2011; Swana et al. 2011).
14.4.5 Bioethanol
Bioethanol is generally produced from grains or molasses, but several researchers have sug-
gested that production of cellulosic bioethanol has more environmental benefits compared
to bioethanol from grains. According to LCA, bioethanol produced from agricultural waste
or cellulosic materials significantly reduces greenhouse gas emissions and has higher sus-
tainability than ethanol produced from grains. Most of the research in this field is aimed at
improving and optimizing production methods based on various feedstocks. Efforts have
been directed to develop economically viable processes for production of bioethanol from
both cellulosic and hemicellulosic sources. The main component of cellulose hydrolyzate
is glucose which is a fermentable sugar and essential in the production of ethanol. Xylose
is a pentose sugar present in the hemicellulosic hydrolyzate. The bioconversion of xylose
into ethanol is possible with a satisfactory yield. However, longer fermentation times and
low ethanol titers are the major challenges in xylose conversion into ethanol (Borrion et al.
2012; Brown and Brown 2013).
14.4.6 Biogas and Fuel Cell

The available reports suggest that the production of biogas like methane from lignocel-
lulosic biomasses can be significantly improved by choosing the appropriate pretreat-
ment method (Dererie et al. 2011). It has been proposed that grass mixture can be
fermented to biogas through the anaerobic digestion process. However, a variety of ligno-
cellulosic biomass such as wood, bagasse, straw, willow, etc. can be promising natural
materials for the synthesis of biogas, which can be easily converted into heat and electric-
ity in solid oxide fuel cells (SOFC) with high‐efficiency power (electricity 54% and heat
36%). As renewable sources are available in large quantities, sustainable power genera-
tion using fuel cell technologies could be a feasible option (Calise et al. 2006; Strazza
et al. 2010).
14.5 Production Process
The biorefinery is based on production of various valuable products like biofuel, renewable
energy, and different biochemicals from lignocellulosic feedstocks. One of the critical steps
in this process is biomass pretreatment. The ideal process should include production of
new by‐products with generation of zero waste (Kumar et al. 2009). Various biochemical
and thermal processes have been employed for processing of feedstocks but there are still
some technical challenges which limit the commercialization of the biorefinery concept.
Therefore, there is a need to develop “green chemistry” processes which can efficiently
convert raw materials into bioenergy and other important products in a sustainable man-
ner (Drapcho et al. 2008).
Currently, the National Renewable Energy Laboratory (NREL) is targeting “advanced
green chemistry” to facilitate its wider deployment. Modern biorefineries are trying to
achieve the target through:
●● improved understanding of lignocellulosic structure
●● lignocellulosic structure engineering and its composition to facilitate the removal of
lignin and increase the efficiency of biomass decomposition processes
●● development of advanced biocatalysts to increase the efficiency and reduce the chemical
and biological risks of biofuels
●● methods development for the full usage of all raw biomass, including waste streams or
unused products to complete the production chain (Gnansounou and Dauriat 2010; de
Jong and Jungmeier 2015).
Thermochemical and biochemical methods are mostly commonly used in biorefineries. In
biochemical conversion, carbohydrate polymers (cellulose and hemicellulose) of biomass are
hydrolyzed and broken down into simple sugars (glucose and xylose) which are further fer-
mented to produce bioethanol and various biochemicals. Lignin, which cannot be hydro-
lyzed, is usually combusted to produce heat and electricity. However, in thermochemical
processes, heat is used to break down the raw biomass to syngas (a mixture of carbon monox-
ide and hydrogen), which is further reacted in the presence of a catalyst to produce ethanol
and higher molecular weight alcohols (Demirbas 2007; Ohgren et al. 2007; Mu et al. 2010).
As far as lignocellulosic ethanol production technology is concerned, utilization of the
appropriate approach and its environmental impact are the most important decision‐
making factors. The most common method used for the assessment of environmental
impacts of a product or a process is the LCA method which provides a comprehensive view
of environmental indicators, thus allowing a more precise vision of the real environmental
interactions in order to make decisions (Demirbas 2007; Mu et al. 2010).
14.5.1 Process Simulation

The process simulation is usually used as a mass and energy balance tool for technical and
economic feasibility studies. However, different feeds and processes for the production of
various materials can be used in LCA. The NREL has developed the “Aspen” environmental
model to improve the conceptual design and provide the process knowledge for a biorefinery.
The latest version of this Aspen model was released in May 2011. This model was prepared by
assuming the target product was ethanol based on cellulose with hydrolysis. The NREL model
can be used in case of both acid and enzyme hydrolysis (ASPEN 2000). A novel computer
program, QSAR, was developed to generate and validate QSAR or QSPR (quantitative struc-
ture‐activity or ‐property relationships) models. It is useful for the determination of carcino-
genicity which occurs due to any specific material production process (Fjodorova et al. 2010).
14.6 Life Cycle Assessment

The LCA is a technique used for the assessment of environmental impacts associated with
products and services. For holistic assessment of the life cycle of any product, it is neces-
sary to evaluate all the stages of a product’s life from raw material extraction through mate-
rials processing, manufacture, distribution, use, repair and maintenance, to disposal or
recycling. This information is required to improve existing processes and provide support
to make management decisions. For the LCA process, the ISO 14000 family of standards
are followed (Rebitzer et al. 2004). According to ISO 14040 and 14044, assessment of the
critical life cycle can be performed in four separate stages: goal and scope, inventory analy-
sis, impact assessment, and interpretation (Figure 14.2) (Guinee 2002; ISO 2006a). These
stages are briefly discussed below.
14.6.1 Goal and Scope

According to ISO 14040, the goal of an LCA study should clearly state the intended applica-
tion such as the reasons for carrying out the analysis and the intended audience, that is, to
whom the results of the study are intended to be communicated. However, in case of scope,
various items need to be clearly discussed including function of product, allocation proce-
dures, data requirement, assumptions, limitations, etc. (ISO 2006). As far as lignocellulosic
biorefinery is concerned, the following important goals are reviewed in its LCA.
●● Feedstock comparison
●● Different production scenarios evaluation
●● Production process alternatives
14.6.2 Life Cycle Inventory Analysis

The second stage of LCA is inventory analysis which involves the detailed analysis of the
production cycle from beginning to end (from nature to nature) through the collection of
data and calculation procedures to quantify relevant inputs and outputs of a product sys-
tem. These inputs and outputs may include the use of resources and any releases to air,
water, and land associated with the system. In LCA, it is preferred to consider current
sources and avoid obtaining data from previous research. In LCA, data accuracy and cred-
ibility should be thoroughly checked. Due to regional differences, it is necessary to be aware
that data obtained from other regions or countries cannot be properly used for local condi-
tions (Guinee et al. 2002; ISO 2006; Finnveden et al. 2009).
LCA Framework
Goal and Scope Definition
Purpose of study, intended

audience, functional unit, system
boundaries, data, assumptions
and limitations, etc.
Interpretation
Analysis of results and

findings in relation to
Inventory Analysis defined goal and scope,
conclusions,
Data collection, calculation, recommendations, etc.
quantification of inputs and
outputs, allocation procedures
and calculation of energy flows,
etc.
Impact Assessment
Classification, Characterization,
Weighting
Figure 14.2 Phases of life cycle assessment (LCA), as per ISO 14040. The phases of LCA are
iterative and repetitive. Source: Hyder et al. (2016).
14.6.3 Life Cycle Impact Assessment

The assessment of life cycle impact is the third important stage of LCA and is performed to
evaluate the significance of potential environmental impacts from the findings of the life
cycle inventory analysis. In general, this process aims to associate inventory data with spe-
cific environmental impacts and attempts to understand those impacts. The impact assess-
ment of the classic life cycle is detailed in the following steps (ISO 2006).
●● Class of impact indicators and specifications model
●● The class priority
●● The impacts of evaluation and normalization
14.6.4 Interpretation
In this stage of the LCA, findings obtained from the second stage (inventory analysis) and
third stage (impact assessment) of LCA are combined with the defined goal and scope in
order to reach relevant conclusions and recommendations. One of the key goals of the criti-
cal interpretation is to evaluate the assurance level of the obtained value. The interpreta-
tion of any LCA is not as simple as just suggesting a product and process. It also describes
the concepts that can be determined according to the objectives. Sensitivity analysis is a
tool used to assess the sensitivity of any effective data as well as the sustainability of any
investigation (Menten et al. 2013).
After interpretation, the critical data analysis is validated by the quantity and quality of
the data presented so it is very important that the data used to complete the analysis of the
cycle should be accurate. It is worth mentioning that sufficient data should be available to
compare any production process. The Research and Development (R&D) wing of compa-
nies presents new materials and production methods with updated information obtained
from the LCA. However, new data generation is time‐consuming, and it cannot be extended
to other times and places. If most environmentally harmful things are recognized, then
there is need to focus on production methods. For example, in the biorefinery, the pretreat-
ment of lignocellulosic feedstocks is a crucial process step that can be of major concern
(Finnveden et al. 2009; Menten et al. 2013; Muench and Guenther 2013).
14.7 Important Types of LCA
14.7.1 Cradle to Grave

This type of LCA includes assessment of the entire critical cycle of the resource (cradle) to
the end‐user and then discarding the waste (grave). For example, paper can be produced
from a tree, which can be further recycled as low‐energy cellulose insulation (fibrous
paper). Later, it can be used as an energy‐saving device in the ceiling of a home for many
years, ultimately saving fuel energy. After a specific time, the old ceiling cellulose fibers
can be replaced by new ones and old ones may be disposed of, possibly by incineration
which is released to the environment (Grave) (Singh et al. 2010; Bonin and Lal 2012;
Borrion et al. 2012).
14.7.2 Cradle to Gate

Cradle to gate involves evaluating the critical cycle of a product from source of extraction
(cradle) to factory (gate). The application phase and waste disposal stage have been removed
in this case. When there are the same application and dissipation, it makes calculations
easier. For example, comparing the production of cellulosic alcohol from bagasse, straw
and wood chips, which differs only in the raw material, the cradle to gate evaluation can
have achieve results related to LCA. In this approach, mainly the various raw materials,
stages of production and manufacturing processes are considered and, more simply, the
values of the cradle to their input for the specified products are determined (Brehmer and
Sanders 2008; Swana et al. 2011).
14.7.3 Cradle to Cradle or Closed Loop

Cradle to cradle can be considered as a cradle to grave assessment category in which the
wastes are recycled. One of the most important strategies for sustainability is zero waste
emission. Minimizing the environmental impacts of products necessitates designing prod-
ucts with recycling potential. In the recycling process, there are two approaches: recycling
the same or new products. Unfortunately, 100% of the energy from fuel cannot be recov-
ered in fuel energy form. When the plastic or other chemical recycling process is consid-
ered, the closed loop can reduce sustainability problems (Sobrino et al. 2011).
14.7.4 Well to Wheel

Well to wheel is a LCA applied for transportation. Well to wheel is common in the evalua-
tion of energy conversion efficiency of different fuels while traditional cradle to grave has
more focus on the vehicle itself. For example, when comparing electricity, gasoline, and
bioethanol, the well to wheel approach can be used to compare them in a working automo-
bile scenario (Cherubini and Ulgiati 2010).
14.8 Applications of LCA in Biorefinery
Most LCA is supporting the development of new bio‐based products as raw materials for
the production of conventional bioproducts. Most companies nowadays apply LCA in new
product development research. Governments need to invest in the development of national
databases, so they can support LCA. For example, data about industrial and natural
resources (forests) and land use can be obtained from local governments. Also, information
on demand for products will be required from central governments for strategic planning
(Singh et al. 2010; Muench and Guenther 2013).
14.8.1 LCA in Lignocellulosic Conversion

In bioethanol production, LCA studies are mainly based on emission of greenhouse gases
(GHG) and energy consumption. One of the most important reasons for this approach is to
compare it with fossil fuels. When acidification and eutrophication are considered, there
are some disadvantages in the pretreatment phase. Well to wheel is the most important
approach used in previous studies for bioethanol production from a variety of lignocellu-
losic biomass but the results of LCA are different for each study with regard to factors such
as system boundaries, functional units, and allocation. It is obvious that choosing a func-
tional unit is very important as the same studies with different functional units will show
different results. About 35% of LCA studies include utilization of wood and forest wastes
for the production of alcohol and in 25% of studies agricultural residues are used
(McKechnie et al. 2011; Silalertruksa and Gheewala 2011).
14.8.2 Basic Challenges in Applying LCA in Second-Generation Biorefinery

The second‐generation biofuels are mostly produced from lignocellulosic feedstocks such
as corn, sugarcane bagasse, etc. and some other waste residues from agriculture and for-
estry. Due to huge transportation costs, feedstocks obtained from agriculture and forestry
are generally stored and processed at small to medium‐scale plants situated close to the
source. Unlike oil companies and government agencies, small and medium‐scale plants
have a less organized structure for decision making. The initial stages of biomass produc-
tion might involve a number of decision makers. Using LCA to influence policies that
would alter the behaviors of these distributed decision makers poses different challenges
than when the decision‐making authority is more highly concentrated. It expects that
farmers utilize land to increase their profits. However, pricing and tax policies can influ-
ence farmers to act in ways that may be difficult to predict, especially in light of imperfect
information, uncertain weather and climate conditions, and complex markets (McKone
et al. 2011).
According to McKone et al. (2011), there are some major challenges for applying LCA to
biofuels.
●● Understanding farmers, feedstock options, and land use.
●● Predicting biofuel production technologies and practices.
●● Characterizing tailpipe emissions and their health consequences.
●● Incorporating spatial heterogeneity in inventories and assessments.
●● Accounting for time in impact assessments.
●● Assessing transitions as well as end‐states.
●● Confronting uncertainty and variability.
14.9 Life Cycle Assessment versus Technoeconomic

Assessment
It is well known that LCA is necessary to prove that a technology could contribute to the
mitigation of environmental impacts; however, tecnoeconomic assessment (TEA) will
show how the technology could be competitively delivered in the market. In LCA, financial
costs related to technology are not considered when drawing any conclusion. Mathematical
and statistical modeling is widely used in environmental and biological systems. These
kinds of modeling can help to determine whether the system is technically, economically,
and environmentally feasible or not. Therefore, together, utilization of LCA and TEA can
result in optimum environmental and economical outcomes.
In this context, various models have been proposed and after a technical and economic
review, it was found that most widely accepted scenarios are based on environmental indi-
cators. For example, among different bioethanol pretreatment processes, chemical meth-
ods may be beneficial as far as economic viability is concerned, but biological or
biotechnological methods are found to be environmentally friendly. Moreover, microbio
engineering sustainability and process economics (ESPE) is a commercially available
model which allows engineers to simplify this complex process and to ensure that custom-
ers can make decisions when upgrading or starting a new production line (Lundquist et al.
2010; Guinee et al. 2011).
14.10 Process Integration for Sustainable Design
In addition to economic development, designing of sustainable process can protect the

environment. The principles of designing sustainable processes depend on factors like
resource conservation, material recycling, energy usage, and application of eco‐friendly
technologies. The important steps required in the integration of process are as follows (El‐
Halwagi 2012; Moncada et al. 2014; Oliveira et al. 2018).
●● Problem identification: rectification of the problem is the most important step so that
each activity can be defined in turn.
●● Objectives: the main objective is to set the product target first without considering the
process and technology constraints. Further, any pathway can be used for determination
of the target, in the next step. If some of these pathways are not suitable for the present
situation, they may be useful for future research strategies.
●● Generate alternatives: there will be many ways to reach the target. The determination of
these solutions is the most important part of the work. Therefore, there is a need for pre-
cision and the use of modern technologies so that possible paths can be identified. In
many cases, changes in the order of complementary operations can be achieved in differ-
ent scenarios.
●● Alternative selection: to achieve the best solution, all scenarios should be assessed accord-
ing to the known goal. Then, each parameter is ranked and optimized by using different
optimization tools in the whole process.
●● Optimal point analysis: after determining the optimal point in general, that point is spe-
cifically studied and analyzed both economically and technically, as well as considering
the environmental aspect. Sensitivity analysis is one of the most common techniques
used in analyzing the optimal point.
14.10.1 Process Integration Case Studies

Process integration is the most effective way to increase operational and economic effi-
ciency, as well as sustainability, in biorefineries. In process integration, several products are
produced employing a basic process, and the maximum waste is recovered in the form of
material or energy. By integrating the process lines, energy consumption will be optimized
and therefore fuel usage will be reduced. For example, if after process integration a boiler
in a sugarcane mill uses less bagasse, the extra bagasse can be used to produce second‐gen-
eration ethanol. The important point is that under different conditions, in this case, etha-
nol and electricity can be flexibly managed according to demand (Moncada et al. 2014;
Oliveira et al. 2018). In a recent study, Oliveira and co‐workers have provided an environ-
mental assessment of the whole biojet production chain from sugarcane in addition to
technical and economic analysis (Oliveira et al. 2018).
In another study, Moncada and colleagues recently examined the possibility of integrat-
ing bio‐based products based on all generations (first, second, and third), considering the
ability to use CO2 for the development of algal third‐generation biomass. In this regard, the
growth of microalga, recovery of oil, and then production of biodiesel from oil are impor-
tant factors. To achieve this goal, a technoeconomic and environmental analysis of two
sugar‐based biophysical scenarios were determined. The scenario of joint production of
sugar, ethanol, electricity, biodiesel, and glycerol has also been studied. The integration of
these processes has the potential to use all of its capacity to reduce environmental hazards
and increase sustainability (Moncada et al. 2014). Figure 14.3 showed a schematic repre-
sentation of an integrated biorefinery.
14.10.2 Decision Support System (DSS) and LCA

In a DSS, the LCA and optimization algorithm are used to select optimal technologies
based on indicators (costs, environmental impacts). First, the target is determined and
based on that, production, capacity, cost, and environmental indicators are considered
simultaneously. In this way, the best operating point can be achieved during the operation
time by considering all indicators (Mussati et al. 2012; Pérez‐Navarro et al. 2015).
Since biorefinery systems are in progress, changes in these systems are inevitable.The
study of the technical, economic, and environmental effects of time and decision making is
of great importance so there is a need for decision support tools which can help to solve the
complexity of a balanced system with different, sometimes conflicting the goals. In the
biorefinery, decisions on the survival of the system are based on progressive problems such
as raw materials (biomass), biomass prices, and new process technologies. Also, production
costs and global energy price fluctuations are of concern. The decision‐making support
system in the bioindustry is a supportive system which requires multiple analysis for deci-
sion making. In particular, in the case of a biorefinery with different feeds from various
agricultural, forestry and industry sources, LCA results can change after launch (Mussati
et al. 2012; Pérez‐Navarro et al. 2015; Benali et al. 2018).
There are different approaches to using the results such as the optimal operating point
approach, problem solving, and feed and process substitution. As noted above, decision
making is a complex process, since the technical and economic fields of the environment
are presented along with various parameters and indicators. The reliability of different sce-
narios is the most important thing when deciding on process accuracy and efficiency. In
this way, decision support tools with appropriate models and calculations of environmental
indicators provide a quick analysis with appropriate graphical interpretations (Mussati
et al. 2012; Pérez‐Navarro et al. 2015).
Sugar Heat USP Gypsum
Electricity Ash Syngas Gases
(Steam) Glycerol
Air Gases
Cogeneration CO2 Capture Glycerol
Sugarcane Sugar Production Cane bagasse Gases
System Purification
Water Water
Waste Water
Gases Dilute
CO2 Raw Ca(OH)2
Microalgae Glycerol
Waste Water
Paste
Molasses Gases
Waste Water Waste Water
Anhydrous Anhydrous Anhydrous
Ethanol Ethanol Ethanol
Water Algae Cultivation,

Ethanol Waste Water Microalgae Biodiesel
Waste Water Waste Water Harvesting & Oil
Production Treatment Production
Sulfuric Acid Lipid Extraction Water
Organic Hexane Sulfuric

Ethanol Gypsum Fertilizer Hexane Gases NaOH Ethanol
Water Matter (Oxygen rich) purge acid Biodiesel
+ Water
(Fertilizer)
Figure 14.3 Schematic representation of an integrated biorefinery. Source: Reproduced from Moncada et al. (2014) with permission of Elsevier.
14.10.3 I-BIOREF Tool

I‐BIOREF V.2.0 is a tool to analyze economic and environmental models to provide an
assessment of environmental impacts with economical consideration. Windows‐based
BIOREF is designed and developed using Excel and Aspen Plus. Commercial software can
perform technical and economic feasibility studies as well as LCA simultaneously.
Developing these types of tools can be of great help in the smooth functioning of biorefin-
eries (Benali et al. 2015).
14.11 Conclusion
Lignocellulose feedstock is the base in biorefineries for the production of fuels, materials,
energy, and commodity chemicals. Life cycle assessment employing process configuration
and product analysis is a key element in the sustainability of lignocellulose biorefineries.
Reviews of this assessment using various raw materials and process comparisons for the
production of bioethanol and biochemicals are widely available in the literature. However,
research on nonfuel bioproducts needs more attention. Creating a biorefinery with multi-
ple fuels and products requires optimization of processes with the process integration
approach. Integration can help in reducing operational and capital expenditure and thus
development of a sustainable process in automated biorefineries. This can be accomplished
using process simulation tools. Environmental assessment is not just needed at the begin-
ning of the process, but over time, by changing the decision conditions that are required by
the LCA approach. A DSS can be a decisive tool for achieving and maintaining sustainabil-
ity in the future.
R
eferences
AFDC (Alternative Fuels Data Center) (2018). World Fuel Ethanol Production by Country or
Region (million gallons). https://afdc.energy.gov.
ASPEN Plus™ (2000). Release 10.2, Aspen Technology, Inc, Cambridge, MA.
Benali, M., Jeaidi, J., Mansoornejad, B. et al. (2018). Decision support systems for assessment
of biorefinery transformation strategies. Canadian Journal of Chemical Engineering 96:
2155–2175.
Benali, M., Mansoornejad, B., Ajao, O. et al. (2015). Simultaneous assessment of technical
performance, economic viability and environmental footprint of forest biorefinery systems:
I‐BIOREF Tool. 6th Nordic Wood Biorefinery Conference Proceedings, Helsinki.
Biddy, M.J., Scarlata, C., and Kinchin, C. (2016). Chemicals from biomass: a market assessment
of bioproducts with near‐term potential. NREL Technical Report, NREL/TP‐5100‐65509,
National Renewable Energy Laboratory, Colorado, USA.
Bonin, C. and Lal, R. (2012). Bioethanol potentials and life‐cycle assessments of biofuel
feedstocks. Critical Reviews in Plant Sciences 31: 271–289.
Borrion, A.L., McManus, M.C., and Hammond, G.P. (2012). Environmental life cycle
assessment of lignocellulosic conversion to ethanol: a review. Renewable and Sustainable
Brehmer, B. and Sanders, J. (2008). Implementing an energetic life cycle analysis to prove the
benefits of lignocellulosic feedstocks with protein separation for the chemical industry from
the existing bioethanol industry. Biotechnology and Bioengineering 102 (3): 767–777.
and Bioenergy 38: 68–94.
Brown, T.R. and Brown, R.C. (2013). A review of cellulosic biofuel commercial‐scale projects
in the United States. Biofuels, Bioproducts and Biorefining 7: 235–245.
Brown, T.R., Wright, M.M., and Brown, R.C. (2011). Estimating profitability of two biochar
production scenarios: slow pyrolysis vs fast pyrolysis. Biofuels, Bioproducts and Biorefining 5
(1): 54–68.
Calise, F., Palombo, A., and Vanoli, L. (2006). Design and partial load energy analysis of hybrid
SOFC–GT power plant. Journal of Power Sources 158: 225–244.
Chandel, A.K., Albarelli, J.Q., dos Santos, D.T. et al. (2019). Comparative analysis of key
technologies for cellulosic ethanol production from Brazilian sugarcane bagasse at the
commercial‐scale. Biofuels, Bioproducts and Biorefining 13 (4): 994–1014.
Cheng, K.K., Cai, B.Y., Zhang, J.A. et al. (2008). Sugarcane bagasse hemicelluloses hydrolysate
for ethanol production by acid recovery process. Biochemical Engineering Journal 38: 105–109.
Cherubini, F. and Ulgiati, S. (2010). Crop residues as raw materials for biorefinery systems: a
LCA case study. Applied Energy 87: 47–57.
Demirbas, A. (2005). Bioethanol from cellulosic materials: a renewable motor fuel from
biomass. Energy Source 27: 327–337.
Demirbas, A. (2007). Progress and recent trends in biofuels. Progress in Energy and Combustion
Science 33 (1): 1–18.
Dererie, D.Y., Trobro, S., Momeni, M.H. et al. (2011). Improved bio‐energy yields via sequential
ethanol fermentation and biogas digestion of steam exploded oat straw. Bioresource
Technology 102: 4449–4455.
DOE (US Department of Energy) (2003). Roadmap for Agriculture Biomass Feedstock Supply
in the United States. http://pacificbiomass.org/documents/DOE_RoadmapForAgBio
massFeedstockSupply.pdf.
Drapcho, C.M., Nhuan, N.P., and Wlaker, T.H. (2008). Biofuel Engineering Process Technology.
New York: McGraw Hill.
El‐Halwagi, M.M. (2012). Benchmarking process performance through overall mass targeting.
In: Sustainable Design through Process Integration (ed. M.M. El‐Halwagi), 63–88. Oxford:
Butterworth‐Heinemann.
Epplin, F.M., Clark, C.D., Roberts, R.K., and Hwang, S. (2007). Challenges to the development
of a dedicated energy crop. American Journal of Agricultural Economics 89 (5): 1296–1302.
ePURE (2017). European renewable ethanol – key figures 2017. https://epure.org/
media/1763/180905‐def‐data‐epure‐statistics‐2017‐designed‐version.pdf.
Finnveden, G., Hauschild, M.Z., Ekvall, T. et al. (2009). Recent developments in life cycle
assessment. Journal of Environmental Management 91 (1): 1–21.
Fjodorova, N., Vracko, M., Novic, M. et al. (2010). New public QSAR model for carcinogenicity.
Chemistry Central Journal 4: 1–15.
Ghosh, D., Dasgupta, D., Agrawal, D. et al. (2015). Fuels and chemicals from lignocellulosic
biomass: an integrated biorefinery approach. Energy & Fuel 29: 3149–3157.
Gnansounou, E. and Dauriat, A. (2010). Techno‐economic analysis of lignocellulosic ethanol: a
review. Bioresource Technology 101: 4980–4991.
Gontia, P. and Janssen, M. (2016). Life cycle assessment of bio‐based sodium polyacrylate
production from pulp mill side streams: case study of thermo‐mechanical and sulfite pulp
mills. Journal of Cleaner Production 131: 475–484.
Gonzalez‐Garcıa, S., Hospido, A., Agnemo, R. et al. (2011). Environmental life cycle
assessment of a Swedish dissolving pulp mill integrated biorefinery. Journal of Industrial
Ecology 15: 568–583.
Granbio (2014). www.granbio.com.br.
Guinee, J.B. (2002). Handbook on Life Cycle Assessment: Operational Guide to the ISO
Standards. Springer, Netherlands: Kluwer Academic Publishers.
Guinee, J.B., Heijungs, J., and Huppes, G. (2011). Life cycle assessment: past, present, and
future. Environmental Science and Technology 45: 90–96.
Hayes, D.J., Fitzpatrick, S., Hayes, M.H.B., and Ross, J.R.H. (2006). The biofine process‐
production of levulinic acid, furfural, and formic acid from lignocellulosic feedstocks. In:
Biorefineries: Industrial Processes and Products, vol. 1 (eds. B. Kamm, P.R. Gruber and M.
Kamm), 139–164. Weinheim: Wiley‐VCH.
Hsu, D.D. (2012). Life cycle assessment of gasoline and diesel produced via fast pyrolysis and
hydroprocessing. Biomass and Bioenergy 45: 41–47.
Hu, G., Heitmann, J.A., and Rojas, O.J. (2008). Feedstock pretreatments strategies for
producing ethanol from wood, bark, and forest residues. BioResources 3: 270–294.
Hyder, Z., Ripepi, N.S., and Karmis, M.E. (2016). A life cycle comparison of greenhouse
emissions for power generation from coal mining and underground coal gasification.
Mitigation and Adaptation Strategies for Global Change 21: 515–546.
ISO (2006). 14040 Environmental Management, Life Cycle Assessment, Principles and
Framework. Geneva, Switzerland: International Standards Organization.
de Jong, E. and Jungmeier, G. (2015). Biorefinery concepts in comparison to petrochemical
refineries. In: Industrial Biorefineries & White Biotechnology (eds. A. Pandey, R. Höfer, M.
Taherzadeh, et al.), 3–33. Amsterdam: Elsevier.
Khanna, M., Dhungana, B., and Clifton‐Brown, J. (2008). Costs of producing Miscanthus and
switchgrass for bioenergy in Illinois. Biomass and Bioenergy 32 (6): 482–493.
Kumar, M. and Gayen, K. (2011). Developments in biobutanol production: new insights.
Applied Energy 88 (6): 1999–2012.
Kumar, S., Singh, S.P., Mishra, I.M., and Adhikari, D.K. (2009). Recent advances in
production of bioethanol from lignocellulosic biomass. Chemical Engineering &
Ladisch, M.R., Mosier, N.S., Kim, Y. et al. (2010). Converting cellulose to biofuels. Chemical
Engineering Progress 106 (3): 56–63.
Lamers, P., Tan, E., Searcy, E. et al. (2015). Strategic supply system design a holistic evaluation
of operational and production cost for a biorefinery supply chain. Biofuels, Bioproducts and
Biorefining 9: 648–660.
Lange, J.P., Price, R., Ayoub, P.M. et al. (2010). Valeric biofuels: a platform of cellulosic
transportation fuels. Angewandte Chemie International Edition 49: 4479.
Li, X., Weng, J.K., and Chapple, C. (2008). Improvement of biomass through lignin
modification. Plant Journal 54: 569–581.
Lundquist, T.J., Woertz, I.C., Quinn, N.W.T., and Benemann, J.R. (2010). A Realistic Technology
and Engineering Assessment of Algae Biofuel Production. Berkeley, CA: Energy Biosciences
Institute, UC Berkeley.
McKechnie, J., Colombo, S., Chen, J. et al. (2011). ‘Forest bioenergy or forest carbon? Assessing
trade‐offs in greenhouse gas mitigation with wood‐based fuels. Environmental Science and
McKone, T.E., Nazaroff, W.W., Berck, P. et al. (2011). Grand challenges for life‐cycle
assessment of biofuels. Environmental Science and Technology 45: 1751–1756.
Menon, V. and Rao, M. (2012). Trends in bioconversion of lignocellulose: biofuels, platform
chemicals and biorefinery concept. Progress in Energy and Combustion Science 38 (4): 522–550.
Menten, F., Chèze, B., Patouillard, L., and Bouvart, F. (2013). A review of LCA greenhouse gas
emissions results for advanced biofuels: the use of meta‐regression analysis. Renewable and
Moncada, J., Tamayo, J.A., and Cardon, C.A. (2014). Integrating first, second, and third
generation biorefineries: incorporating microalgae into the sugarcane biorefinery. Chemical
Engineering Science 118: 126–140.
Mu, D., Seager, T., Suresh‐Rao, P., and Zhao, F. (2010). Comparative life cycle assessment of
lignocellulosic ethanol production: biochemical versus thermochemical conversion.
Environmental Management 46: 565–578.
Muench, S. and Guenther, E. (2013). A systematic review of bioenergy life cycle assessments.
Applied Energy 112: 257–273.
Mussati, M.C., Pieragostini, C., and Aguirre, P. (2012). On process optimization considering
LCA methodology. Journal of Environmental Management 96: 43–54.
Ohgren, K., Bura, R., Saddler, J., and Zacchi, G. (2007). Effect of hemicellulose and lignin
removal on enzymatic hydrolysis of steam pretreated corn stover. Bioresource Technology 98
(13): 2503–2510.
Oliveira, C.M., Pavão, L.V., Ravagnani, M.A.S.S. et al. (2018). Process integration of a
multiperiod sugarcane biorefinery. Applied Energy 213: 520–539.
Pérez‐Navarro, Á., Hurtado, E., Peñalvo‐López, E. et al. (2015). Optimization of a hybrid
renewable system for high feasibility application in non‐connected zones. Applied Energy
155: 308–314.
Rajendran, K. and Murthy, G.S. (2017). How does technology pathway choice influence
economic viability and environmental impacts of lignocellulosic biorefineries? Biotechnology
for Biofuels 10: 268.
Raman, J.K. and Gnansounou, E. (2015). LCA of bioethanol and furfural production from
vetiver. Bioresource Technology 185: 202–210.
Rebitzer, G., Ekvall, T., Frischknecht, R. et al. (2004). Life cycle assessment: part 1: framework,
goal and scope definition, inventory analysis, and applications. Environment International
30 (5): 701–720.
Román‐Leshkov, Y., Barrett, C.J., Liu, Z.Y., and Dumesic, J.A. (2007). Production of
dimethylfuran for liquid fuels from biomass‐derived carbohydrates. Nature 447: 982–985.
Scheller, H.V. and Ulvskov, P. (2010). Hemicelluloses. Annual Review of Plant Biology 61:
263–289.
Silalertruksa, T. and Gheewala, S.H. (2011). Long‐term bioethanol system and its implications
on GHG emissions: a case study of Thailand. Environmental Science and Technology 45:
4920–4928.
Singh, A., Pant, D., Korres, N.E. et al. (2010). Key issues in life cycle assessment of ethanol
production from lignocellulosic biomass: challenges and perspectives. Bioresource
Technology 101: 5003–5012.
Sobrino, F.H., Monroy, C.R., and Perez, J.L.H. (2011). Biofuels and fossil fuels: life cycle
analysis (LCA) optimization through productive resources maximization. Renewable and
Stephen, J., Mabee, W.E., and Saddler, J.N. (2012). Will second‐generation ethanol be able to
compete with first‐generation ethanol? Opportunities for cost reduction. Biofuels,
Bioproducts and Biorefining 6: 159–176.
Strazza, C., DelBorghi, A., Costamagna, P. et al. (2010). Comparative LCA of methanol‐fuelled
SOFCs as auxiliary power systems on‐board ships. Applied Energy 87: 1670–1678.
Swana, J., Yang, Y., Behnam, M., and Thompson, R. (2011). An analysis of net energy
production and feedstock availability for biobutanol and bioethanol. Bioresource Technology
102 (2): 2112–2117.
USDE (2009). US Department of Energy Biomass Program. www.energy.gov/sites/prod/
files/2014/03/f14/obp_overview_algae_summit.pdf.
Vohra, M., Manwar, J., Manmode, R. et al. (2014). Bioethanol production: feedstock and
current technologies. Journal of Environmental Chemical Engineering 2: 573–584.
Zheng, J.L., Zhu, Y.H., Zhu, M.Q. et al. (2018). Life‐cycle assessment and techno‐economic
analysis of the utilization of bio‐oil components for the production of three chemicals.
Green Chemistry 20: 3287–3301.
333
15
Technoeconomic Analysis of Biorefinery Processes

for Biofuel and Other Important Products
Harikishan R. Ellamla1 and Srinivas Appari2
1
South African Institute for Advanced Materials Chemistry (SAIAMC), University of Western Cape, Cape Town, South Africa
2
Department of Chemical Engineering, Birla Institute of Technology and Science Pilani, Pilani, Rajasthan, India
15.1 Introduction
A biorefinery is a facility that converts biomass into biofuels, power and/or heat, and other
useful bio‐based value‐added products such as food, feed, chemicals, and other materials.
The biorefinery concept is similar to the petroleum refinery (Banerjee et al. 2013; Jungmeier
et al. 2014). As for petroleum refineries, biorefineries can provide an array of chemicals by
separating the initial biomass into various intermediates such as carbohydrates, proteins,
and triglycerides that can be further treated to obtain more value‐added chemicals
(Jungmeier et al. 2013; Shafiei et al. 2013; Davis et al. 2014; Mejdell et al. 2014; Quinn and
Davis 2015; Soleymani and Rosentrater 2017). The use of biomass as feedstock not only
reduces dependence on oil imports but also significantly minimizes impacts on the envi-
ronment (Kumar et al. 2018).
The biorefinery process is classified into primary and secondary processes. The primary
refining process includes biomass pretreatment, preparation, and separation of constituent
elements in the biomass. The primary refining products are subsequently processed in sec-
ondary refining. The primary refining step involves the pretreatment conditioning of the
biomass and its constituents into intermediates such as cellulose, lignin, plant fibers,
starch, sugar, vegetable oil, biogas, and synthesis gas. In the secondary refining process,
further conversion of intermediate products from primary refining involves various steps to
produce a variety of products. During the first conversion stage, the intermediate materials
are partially or fully converted into more intermediates, and these are further fully or par-
tially converted into refined products. The products from biorefineries can be both finished
and semi‐finished. The intermediates produced as a result of the refining process are used
to supply process energy or are further converted into food or feed.
The conversion of raw material into products requires a different unit operation that
requires various forms of energy such as current, heat in the form of steam, cooling water,

refrigeration, etc. From the choice of reaction path, the quality of the product and number
of unit operations can be defined with related costs and incoming streams. The setting up
of input and output streams and overall material and energy balances play a vital role in the
process economy. Further, the optimization of a process from an economic perspective is
complicated and requires trade‐offs between competing factors.
A variety of technically feasible solutions are available in the literature to meet a particu-
lar objective for a process plant. Methods to evaluate these economics in a standardized
format to facilitate decision making on plant and optimization of designs have been devel-
oped. Technoeconomic evaluations allow organizations to decide on which projects to con-
tinue and how to optimize design to maximize profits. The material and energy balances of
each processing technology are required to approximate the size of process units and the
capital cost. The direct and indirect overhead cost factors along with the equipment costs
are required to calculate the capital investment. Further, plant operating expenses are cal-
culated from the fixed capital investment (FCI) of the inlet/outlet flow of the material of
the plant. Discounted and nondiscounted cash flow estimations are useful to predict the
rate of return and net present value (NPV), and are further useful for the evaluation of a
precommercial process product price. In this chapter we describe the technoeconomic
modeling approach for biomass conversion into various products.
15.2 Biorefinery Classification
Many approaches are used to systematize biorefinery concepts (Wang et al. 2014;
Vijayakumar and Saravanan 2015). Depending on the approach, the biorefinery is classi-
fied into four main categories: platforms, products, feedstocks, and processes. A schematic
representation of a biorefinery process is shown in Figure 15.1. The blend of these catego-
ries indicates each biorefinery system. The various combinations of process systems pro-
vide a summary of the most viable process in a refinery network.
15.2.1 Platforms
The platforms are considered to be the main blocks of the biorefinery system. Platforms
are the intermediates in the refining process that connect various input and output
streams of the processing equipment. Different processing techniques are used in the
platforms to convert the intermediates into marketable products. The energy‐driven plat-
form is considered to be the essential platform in the biorefineries system as shown in
Table 15.1.
15.2.2 Products
The products derived from the biorefinery are classified into energy‐driven and material‐
driven that includes both energetic and nonenergetic products. The secondary energy car-
riers such as transportation biofuels, power and/or heat are considered to be an energy‐driven
biorefinery system. The primarily generated bio‐based products such as food, animal feed,
fine chemicals, biomaterials, lubricants, and other process residues are treated as a mate-
rial‐driven biorefinery system. The residues generated during the process are further
employed or treated to produce a useful form of energy.
Technoeconomic Analysis of Biorefinery Processes for Biofuel and Other Important Products 335
Feedstock
Legend
Primary refining
Secondary refining
Pre-treatment of biomass
Platform
Conversion/Refining
process
Energy products Material products Co-products
Figure 15.1 Schematic representation of a biorefinery.
15.2.3 Feedstocks
Biomass feedstock is considered to be a renewable energy source that can be treated to
produce various forms of fuels and/or energy products. The composition of feedstock
depends on the source of origin and has different ratios of primary components such as
cellulose, hemicellulose, lignin, starch, etc. and the three elements carbon, hydrogen, and
oxygen. Moisture content, calorific value, and specific volume of the material also vary for
each feedstock source. Biomass feedstocks are plant (corn, sugarcane, and other crop resi-
dues) and aquaculture materials (algae, seaweed) which are also used to produce various
green hydrocarbon fuels such as alcohols (ethanol, butanol) and biodiesel.
15.2.4 Processes
An array of technologies have been adapted to convert the various biomass feedstocks into
marketable products. Biorefinery processes can be classified based on final products (e.g.,
fuels, chemicals, materials, food, feed) and can be classified into four main subgroups. The
first classification is based on thermochemical conversion (fast/slow pyrolysis, gasification,
hydrothermal upgrading) that uses heat as the dominant mechanism to convert biomass
into a more valuable fuel (Cherubini et al. 2009). The second classification is based on
chemical conversion that includes processes such as hydrolysis, transesterification, hydro-
genation, oxidation, etc. where a substrate is subjected to chemical changes. The third clas-
sification is based on biochemical conversions, such as anaerobic digestion, aerobic and
anaerobic fermentation, and enzymatic conversion. These processes are generally carried
Table 15.1 The elements of biorefinery classification (Taylor 2008).
Platforms Products Feedstocks Processes
I) C5 sugars I) Energy products I) Dedicated crops I) Thermochemical

(glucose, fructose, ●● Biodiesel ●● Oil crops ●● Combustion
galactose) ●● Bioethanol ●● Sugar crops ●● Gasification
II) C6 sugars ●● Biomethane ●● Starch crops ●● Hydrothermal

(xylose, arabinose) ●● Synthetic ●● Lignocellulosic upgrading
III) Vegetable oils, biofuels crops ●● Pyrolysis
lipids ●● Electricity and ●● Grasses ●● Supercritical

heat ●● Marine II) Biochemical
IV) Biogas
II) Material products biomass ●● Fermentation
V) Syngas ●● Food II) Residues ●● Anaerobic
VI) Hydrogen ●● Animal feed ●● Lignocellulosic digestion
VII) Organic juice ●● Fertilizer residues ●● Aerobic
●● Glycerine ●● Oil‐based conversion

VIII) Pyrolytic residues
●● Biomaterials ●● Enzymatic
liquid
●● Organic processes
●● Chemicals and
IX) Lignocellulose building blocks residues and III) Chemical
components others processes
●● Polymers and
(lignin/cellulose/ ●● Catalytic
hemicellulose) resins
●● Biohydrogen
processes
X) Electricity and ●● Pulping
heat ●● Esterification
●● Hydrogenation
●● Hydrolysis
●● Methanization
●● Steam
reforming
●● Water
electrolysis
●● Water gas shift
IV) Mechanical/
physical
●● Extraction
●● Fiber separation
●● Mechanical
fractionation
●● Pressing/
disruption
●● Pretreatment
●● Separation
out at low temperature and pressure. Finally, the fourth classification includes mechani-
cal/physical processes in which there is no change in the chemical structure of the biomass.
A size reduction or a separation of feedstock components occurs by utilizing various unit
operations such as mechanical pressing, pretreatment, milling, separation, filtration,
extraction, crystallization, adsorption, sieving, and distillation.
15.3 Biorefinery Technoeconomic Evaluations
The biorefinery process transforms raw materials into more marketable products and
therefore, more valuable materials that provide benefits to the end‐users (Brown 2015;
Gubicza et al. 2016). In its simplest form, a biorefinery process consists of a series of raw
material and heat flows which can be represented by a simple block diagram as shown in
Figure 15.2.
The material and energy flows in and out have an economic value either as a cost or a
source of income. The energy consumed in the process will have a cost which reflects its
generation and transmission, but some processes may generate waste that can be utilized
as an energy source within the plant or sold to a third party to make an income.
Transformation of raw material into products can be represented with simple stoichio-
metric chemical reactions. The stoichiometry and thermodynamics of a biochemical
reaction determine the maximum possible conversion of raw materials into products as
well as the energy requirements. The output material from a reactor will contain uncon-
verted raw material, products, and impurities. Unconverted raw material and waste prod-
ucts are separated from the useful products, and unconverted raw material may be reused
to improve overall product yield. Waste products will need to be treated such that they
can be reused, recycled, burned to generate heat energy or disposed of in an environmen-
tally sound manner.
The conversion of feedstock into products requires different unit operations which have
energy requirements in the form of current, heat in the form of steam, cooling water, refrig-
eration, etc. The requirements of steam and electricity in the plant may be generated on site
or may be imported from third parties. The quantity and quality of the feedstock and prod-
ucts are calculated from each process. The amount of feedstock requirement is a function
of the reaction yield and product quantity. The quality of the feedstock/product is defined
by its purity. Undesired by‐products and/or waste products from the reaction can be mini-
mized by maintaining the proper quality of feedstock and operating conditions. Based on
the reaction path, the process flowsheet consists of all necessary unit operations that can be
defined with associated costs and income streams.
The material and energy streams play an important role in the process economy. The set-
ting of the overall energy and material balance is, therefore, a critical step in the manufac-
turing process. Further, the optimization of a process from an economic perspective is
complex and requires trade‐offs between competing factors. In some cases, the selection of
reaction path may require the choice between a process that requires more expensive feed-
stock but is less energy intensive and one for which the feedstock is inexpensive but could
Feedstock
Products
Enzyme/Catalyst Biorefinary Waste products
Energy
Energy
Figure 15.2 Schematic diagram of a biorefinery.

require high temperature and high pressure for the reaction and may involve more
downstream processing steps to separate the final products from a waste product.
There are a variety of technically feasible options available in the literature to meet a
particular process objective for a biorefinery plant (Gutiérrez et al. 2017; Olcay et al. 2018).
Methods to evaluate these economics in a standardized format to facilitate decision making
on required plant and optimization of designs have been developed (Silva et al. 2017).
Technoeconomic evaluations allow organizations to decide on which projects to continue
and how to optimize the design to maximize profits (Shafiei et al. 2011; Vlysidis et al. 2011;
Quintero et al. 2013; Rajendran and Murthy 2017). Figure 15.3 describes the technoeco-
nomic modeling approach used for biomass conversion into products, including process
design and modeling (Davis et al. 2014). This approach is widely used in most of the case
studies, although with a reduced timeline and with extra extrapolations relating to geo-
graphical areas.
A process flow diagram (PFD) is commonly used to represent the flow of inputs and
outputs of a process stream along with the processing equipment. The PFD shows the con-
nectivity between the processing equipment of a plant and a description of the equipment
but not providing piping dimensions and designations. All process flow input and output
streams are identified by a number. All utility streams of the equipment and basic control
strategies are shown in the PFD. The material and energy requirements of each piece of
equipment are needed for estimation of equipment size and capital cost. The thermody-
namic models can be further explored for rigorous evaluation of material and energy bal-
ances of the process. Once the process unit’s costs are known, direct and indirect overhead
cost factors are used to calculate the total capital investment.
Figure 15.3 Process flow sheet of a technoeconomic analysis.

Process flow diagrams
Material and energy

blances
Capital and operating

cost estimation
Economic model
estimation
Minimum product
selling price
Further, plant operating expenses can be estimated from a fixed capital investment of the
inlet/outlet of the flow of the material of the plant. Discounted and nondiscounted cash
flow estimations are useful to predict the rate of return and NPV, and are further useful to
estimate the precommercial process product cost.
15.3.1 Cost Estimation

Financing is one of the essential steps to enable biorefining capacity. The capital costs asso-
ciated with biorefining facilities are high, due to the nonexisting infrastructure, and all
aspects of the project must be built from scratch. The capital cost estimates how much
money will be needed to create a new plant. Capital costs are one‐time expenses incurred
during land acquisition, buildings construction, and pieces of equipment used in the pro-
duction of goods, which provide the total cost needed to start a commercial process under
operating conditions (Peters and Timmerhaus 1991). Whether a particular cost is a capital
or not depends on various parameters such as accounting, tax laws, and materiality. The
operating cost of a plant is known as working capital that estimates the money needed to
operate a plant. Both capital and operating costs indicate the several specific types of com-
posite values that reflect process profitability, and are necessary when evaluating the feasi-
bility of a process. A detailed technoeconomic analysis is discussed in the following
sections.
15.3.2 Capital Cost

The capital cost of a biorefinery includes many costs, such as the cost of the equipment, all
the materials and other associated costs related to the property, material delivery, building
infrastructure, driveaway hooking up facilities, and so on. Table 15.2 represents a summary
of all the associated costs that must be used to calculate the total capital cost of a biorefin-
ery. The capital costs can be divided into four types as shown in Table 15.2. The sum of fixed
capital plus working capital investment is the total capital investment cost.
Table 15.2 Capital costs of biochemical plants.
Direct costs Indirect costs Auxiliary services Contingencies
Preproject study and analysis Engineering and Auxiliary buildings Contractor fees
expenses supervision Land and land Contingencies
Equipment free on‐board cost Construction improvement
Equipment installation expenses Offsite and utilities
Piping (installed) Freight, insurance,
Instrumentation and control and taxes
Electrical installation Construction
(including
Starting‐up costs services)
Interest during
construction
Preliminary economic studies, like market surveys, transportation, laboratory and pilot
plant, are generally carried out before taking a decision or supporting the construction of a
plant. It is necessary to identify the factors that have a significant influence on cost perfor-
mance in the preproject planning phase. Equipment costs at the manufacturer’s site are
provided by the vendor in the form of pro‐forma invoices which give the most accurate
estimate of the equipment cost. The next alternative is to compare the costs of previously
purchased equipment of a similar nature. Another approach, which is sufficient for study
and preliminary estimates of the equipment costs, utilizes summary graphs available for
different types of conventional equipment.
Equipment installation charges are estimated as 20% of total equipment cost. The cost of
installation also includes payment to qualified personnel. In many cases, piping, electrical,
instrumentation, and control installation cost components are calculated separately from
the rest of the equipment. Piping installation cost calculations are made with the help of
diagrams of pipes and their locations. All these installation costs consist of labor charges
and the type of material used. Start‐up costs are also considered between a period from a few
weeks to several months during the formal completion of construction and commencement
of normal production. This cost can be divided into two main groups: one is construction
costs during start‐up, which include loss on construction lines and equipment, mistakes in
design to be solved, need for additional equipment, etc. This is always included in fixed capi-
tal which is depreciating with time in the plant’s useful life. The other main group is start‐up
operational costs that include raw materials, finished or semi‐finished products falling out-
side specifications, salaries, etc. These start‐up costs can be reducible with better design.
Engineering and consultation expenses include payment for technical and administra-
tive services, engineering work, and blueprints of the various jobs or equipment.
Construction expenses are paid for smooth plant operation and usually include field engi-
neering supplies, construction equipment, and temporary services. Freight charges include
all shipping costs for the equipment and material transfer to the site. Insurance cost con-
sists of all insurances associated with the shipping of process units. Sometimes taxes are
added on top of the purchased cost.
Interest to be paid during construction can be established in two situations. First is the
capital required for the development of the project, and the second is that some of the
funds come from external sources. The interest is compounded from the moment the credit
is received until completion of the plant construction. It will be added to the loan amount
to make up the total investment component.
Contractor fees vary according to the condition of the project and can be zero when the
same firm is in charge of the building of the project. Contractor fees differ from one plant
to another. Contingencies are funds which cover unexpected incidents such as sudden
weather changes like storms, or hurricanes. This amount varies and depends on the accu-
racy of the estimation, which also includes unexpected strikes, modifications in the process
design, and unanticipated increases in prices. The contingency can vary from 3% to 10% of
the total direct cost (Humbird et al. 2011). The contingency factors are generally in the
range 5–15% of the fixed capital investment for a plant (Peters and Timmerhaus 1991).
The auxiliary building cost includes administrative and security infrastructure, work-
shop, control rooms, storage, and service buildings for personnel, which consist of the caf-
eteria, meeting rooms, and medical facilities.
The purchase cost of land varies from location to location and country to country. The
land cost can range from 30% to 50% cost factor between a rural area and an industrial area.
The value of land always appreciates with time so the land cost is excluded from the annual
cost of depreciation. Industrial land costs account for 1–2% of total capital investment
(Peters and Timmerhaus 1991). The land improvement cost is part of the investment and is
used for land improvement such as the cost of materials for fences, leveling of the land,
roads, parking lots, electrical, water, and sewer systems, and other similar expenses.
Offsets and utility cost include feedstock and product storage and loading and unloading
amenities, all unit operation utilities (cooling water, steam, fuel, compressed air, electrical
power, etc.), fire protection systems, and other environmental control systems (wastewater
treatment, incinerators, air conditioning systems, etc.).
15.3.3 Capital Cost Estimation

Fixed capital cost investment estimation methods can vary from a quick estimate to detailed
evaluation. The detailed estimation is based on process flowchart detailed information,
product information, time and other plant data available. For the calculation of fixed capi-
tal investment, the cost indexes and cost factors must be considered. A cost index is a num-
ber denoting a ratio between the price of equipment at a time “t” to the price of equipment
at the base time “tb.” Most costs of chemical industry plants are estimated based on the
Marshall and Swift Equipment Cost Index and the Chemical Engineering Plant Cost Index
(Vatavuk 2002; Turton et al. 2012; Dutta et al. 2016). The cost index values for each year are
shown in Table 15.3 (CEPCI 2019).
The purchased equipment cost at a particular time is defined as
Ct It (15.1)
Cb Ib
Table 15.3 Values of the Chemical Engineering Plant Cost Index

and Marshal and Swift Equipment Cost Index from 2001 to 2010.
Chemical Engineering Marshal and Swift

Year Plant Cost Index Equipment Cost Index
2001 1094 394

2002 1104 396
2003 1124 402
2004 1178 444
2005 1244 468
2006 1032 500
2007 1373 525
2008 1449 575
2009 1469 521
2010 1457 550
where Ct is a purchased cost of the equipment at a time “t,” It is a cost index value at a base
time “t,” Cb is a purchased cost of the equipment at the base time “t” and Ib is a cost index
value at the base time.
The present equipment cost can be estimated based on previous data, which vary depend-
ing on equipment size.
Effect of capacity on purchased equipment cost can be defined as:
n
Cr Ar
(15.2)
Cb Ab
where C is a purchased equipment cost, A is an equipment cost attribute, subscript “r” is a

required attribute, subscript “b” is equipment at base attribute cost condition, and n is cost
exponent.
Module costing is the most commonly used cost estimation technique for a new chemical
plant. This technique evaluates the equipment cost by multiplying by a cost factor into
equipment purchased cost at known base conditions. Bare module equipment cost esti-
mates both indirect and direct cost of each equipment.
C BM C p0 FBM (15.3)
where CBM is an equipment bare module cost, FBM is an equipment bare module cost factor
(which depends on construction material and operating pressure), and C p0 is a purchased
cost of equipment at base conditions.
15.3.4 Working Capital

Working capital is the capital fund necessary for the plant to operate at the levels pro-
jected in the technical and financial studies, once it has been constructed and normal
plant operations begin. Working capital is the initial requirement for a few months
before the actual operations of the plant begin. The working capital cost depends on
the financial market of the products, the characteristics of the process, and the avail-
ability of raw materials. Generally, the working capital is recovered at the end of the
project term.
15.3.5 Working Capital Estimation

In the literature, various working capital estimation methods are available. In the first
method, working capital is taken as 10–20% of fixed investment (Chovau et al. 2013). In the
second method, working capital is taken as 10% of annual sales. Table 15.4 shows the per-
centages of each working capital component in terms of yearly sales (Bauman 1964) which
is generally represented as an addition of one month’s raw materials cost plus two months’
finished products cost.
Table 15.4 The average cost of each working capital component

in terms of annual sales.
Percentage of
annual sales
Current assets (A) 18.5

Raw material 1.2
Finished product 4.8
Outstanding bills 10.0
Cash in hand 2.5
Current liabilities (B) Current liabilities 8.64
Taxes 4.80
Salaries 0.42
Services 0.40
Freight 0.02
Raw material 3.00
Working capital (A–B) 9.86
15.3.6 Operating Costs

Operating costs, also called production costs or manufacturing costs, are the costs neces-
sary to maintain day‐to‐day plant operation, processing line or equipment in production.
Gross profit of the company is the difference between income (from product sales and
other resources) and production costs. The production costs are divided into two catego-
ries: direct manufacturing costs, which are proportional to the production rate of the plant,
and fixed costs (including general expenses), which are not dependent on the production
rate. The operating costs of the plant are shown in Figure 15.4. Factors affecting the cost of
operating a plant are shown in Table 15.5.
15.3.7 Operating Costs Estimation

Operating costs estimation allows the selection of operation processes that will save time,
energy, and money. It identifies the costs which most influence project profitability. It helps
to inform decisions and estimates of possible upcoming production costs, which are needed
to determine monetary and commercial studies.
Operating costs = Indirect manufacturing costs + Fixed manufacturing costs
+ General expenses
15.3.7.1 Indirect Manufacturing Costs

The raw material cost can vary from 10% to 50% of the total plant production cost. The total
raw material cost can be calculated from the following equation.
N (15.4)
C RM n * i 1
QRM * PRM
Production costs
ts
cos ut
tur
ing itho
sw
nuf
ac ost
gc
Sal ma urin
es i rect u fact
D an
d m on
Fixe reciati
dep
Biorefinary
Cash flow
Depreciation
er
Oth me Gro
ss p
n c o rofit
i Net Profit
Ta
xe
s
Figure 15.4 Schematic operating cash flow of a biorefinery plant.
Table 15.5 Factors affecting the cost of operating for a plant.
Type of costs Individual cost items Comments
Direct costs
Raw material These costs vary with
Direct labor production rate.
Supervision At low production rate, they are
Maintenance directly proportional to the
production rate
Utilities
Supplies
Royalties and patents
Packaging
Fixed manufacturing
costs
Depreciation These costs do not vary with
Property taxes production rate
Insurance
Credit (financing)
Other obligations
General expenses
Research and development Costs associated with
Public relations management and
Accounting and auditing administrative activities
Legal advice and patents
where CRM is the total raw materials cost, QRM is the raw material quantity needed to make
one unit of the product, PRM is the raw material price of one unit, “N” is the number of
units produced in the process, and “n” is a number of different raw materials involved in
the process.
The operating labor cost varies from 10% to 25% of the total production cost. For highly
mechanized processes, it may be less than 10%. Supervision costs are the salaries paid to
those directly responsible for supervising operations. In most of cases, it is taken as 18% of
the total labor cost. The utility cost includes the cost of electricity, water, and steam, which
are directly influenced by fuel cost. The maintenance cost consists of the materials and
persons employed in regular or subsidiary maintenance in the plant and in some cases, the
renovation of plant equipment and project buildings. In most cases, the maintenance cost
can vary between 4% and 6% of the fixed capital investment. Supplier cost can be taken as
6% of the total labor cost plus 15% of plant maintenance costs. Royalties and patents can be
taken as 1–5% of the total sale price of the product. Packing cost is sometimes estimated
separately, and sometimes is considered as part of the raw material costs.
COL N OS N OOS Ch N OY (15.5)
where COL is the cost of operating labor, NOS is the number of operators required per shift,
NOOS is the number of operators needed for several operating shifts in a year, Ch is the labor
cost per hour, and NOy is the number of working hours in the year per person.
The total number of operators needed for the plant is calculated based on the number of
working hours in a year and the number of plant working days in a year. If one operator
works an average of 49 weeks in a year with five eight‐hour working shifts a week, the
number of shifts per operator in a year is 245 (49 weeks per year × 5 shifts per week). If the
plant runs throughout the year (365 days/year × 3 shifts/day), then the total is 1095 operat-
ing shifts in a year and the total number of operators needed for the plant is 4.5 [(1095
shifts/year)/ (245 shifts/operator/year)].
15.3.7.2 Fixed Manufacturing Costs

Fixed manufacturing costs include property taxes, insurance, and depreciation. These costs
are not dependent on plant production rate, and are charged even during plant shutdown.
Depreciation means a reduction in value as an item increases in age. The main objec-
tive of depreciation estimation is to recover the capital cost invested, and it allows the
estimation of plant indirect production costs over time. The land is one of the fixed
assets but it does not need depreciation estimation, as its value remains typically the
same throughout the life of the plant or may actually increase. Salvage value is calcu-
lated from the difference between the fixed capital investment of the plant and land
value, and is evaluated at the end of the plant life. Most biorefinery plants are built
within the main plant of operation to generate extra income from waste biomass, in
which case initial zero land value is assumed and the corresponding salvage value is
also zero at the end of the plant. The depreciable capital cost of the plant that has not
yet been depreciated is known as book value.
j k
Bk FCI L j 1
dj (15.6)
where dj is the depreciation and k is the number of years.

The depreciation rate can be calculated in four ways: straight line, double declining bal-
ance, sinking fund, and sum of the year’s digits.
In the straight‐line method, the depreciation cost is charged equally in each year over the
depreciation period. It is the simplest and most widely used method.
FCI L S (15.7)
DkSL
n
where FCIL is a fixed capital investment not including land value, S is a salvage value, and
n is the expected useful life of the plant.
The double declining balance method is an accelerated depreciation method. It results in
lower income tax payments for earlier years.
n j k 1
DkDDB FCI L j 0
dj (15.8)
2
The sum of the year’s digits method is based on the sum of the number of years in the plant
asset’s useful life. It can be calculated as:
FCI L S n 1 k
DkSL (15.9)
n
n 1
2
The sinking fund method will provide an amount of depreciation as well as funds for the
replacement of the asset.
Property taxes vary widely according to current laws and location of the plant. The overall
taxation rate varies from 30% to 50%. Insurance is obtained to cover the property, personnel
and merchandise, drop in wages, etc. Credit financing is the compensation paid for the use of
loaned capital. Other obligations include rent of land/equipment, contributions, etc.
15.3.7.3 General Expenses

General expenses include management, sales, legal advice, patents, financing, and research
functions. These are the costs necessary to maintain a plant’s day‐to‐day operations and
administer its corporate business. These costs are not directly attributable to the production
rate of goods and plant services. General expenses rarely vary with production level.
The administrative costs relate to the cost of operating labor and FCI.
Administrative costs 0.009 * FCI 0.177 * COL (15.10)
Sale and distribution costs are estimated at 1% of total sales or 10–11% of total manufactur-
ing costs. Research and development costs are estimated at 5% of total manufacturing costs.
15.3.8 Investment
The investment goal is to make more money which can be achieved by manufacturing
high‐value products from low‐value material (biomass). The biorefinery industry produces
costly chemicals from the waste of low‐value raw materials or biomass, and these are used
as an alternative to conventional products. The economic analysis enables the selection of

alternatives for which capital cost might be required during the period of the project. The
estimation of time effect on capital investment needs to be considered. If the time value
relationship with capital costs is not adequately accounted for, the results of the economic
studies will be inaccurate and might lead to incorrect decisions being made. Direct testing
of alternative operational procedures is generally costly and, in many cases, impossible.
Decision models and simulation processes provide a suitable way in which the assessor can
obtain useful information from the operations under his or her control without disturbing
the activities themselves. The simulation process is a method of indirect experimentation
through which alternative courses of action can be explored before being applied for real.
For the estimation of project profitability, a projection of the lifetime of the project is
needed. Time, cash, and interest rate are three bases required for the evolution of the
project. There are two types of profitability criteria: discounted profitability and nondis-
counted profitability criteria. The discount rate can be 10% for renewable energy invest-
ments (Short et al. 1995). Nondiscounted profitability criteria do not consider the time
value of money invested. A cumulative cash flow diagram of a new production plant
after taxes is shown in Figure 15.5. In Figure 15.5, the y‐axis represents the money value
and the x‐axis represents a number of years. At time value zero, land purchases take
place and after that the construction of the project begins. Based on the size of the plant,
six months to three years may be needed for construction of the plant. During this
period, cash flow is negative, and it is equal to the sum of money paid for the land and
other fixed investments of the plant. When the plant is ready to start production, there
is an extra amount of money needed, known as working capital. After the initial start‐up
of operation, the plant begins to produce final products for sale and the money flow
accumulates, and will move from negative to positive. After some period of operation,
the cash flow rate decreases; this happens because of depreciation. The payback period
is an estimation of the number of years required for the project to return cash equal to
the amount invested for the plant.
25.0
Land
20.0 +
Project construction starts Working Capital
15.0 +
Construction completed Salvage value
Project value
10.0 +
Plant start-up
5.0
0.0
Land
–5.0 Project completed
Fixed Capital or plant shutdown
10.0 Depreciation period
Working Capital
Project life for economic comparisons
–15.0
–1 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14
Project Life (Years)
Figure 15.5 A cumulative cash flow diagram of a new production plant after taxes.
15.3.8.1 Nondiscounted Profitability Criteria

Nondiscounted profitability criteria is a profitability estimation method in which the
amount received in the future is assumed to have equal value to money that was invested
many years earlier. The shorter the payback period (PBP) of the plant, the more profitable
the project.
The payout time, also known as the PBP, is defined as the minimum amount of time
necessary to recover the actual investment of the plant. The original plant investment con-
sists of the initial, fixed, and depreciable costs. The PBP starts after start‐up of the plant and
is the length of time until an investment reaches a break‐even point.
Fixed depreciable investment value Average depreciation of the plant

PBP
average plant profit / year year

(15.11)
The shorter the PBP, the more desirable the investment. Conversely, the longer the pay-
back, the less desirable it is.
Rate of return on investment (ROROI) of the plant is a percentage value, and is normally
expressed as follows.
Average annul net profit
ROROI (15.12)
FCI L
Cumulative cash position is an estimation method, which is the worth of the project at the
end of its life.
Sum of theall positive cash flows
Cumulative cash position (15.13)
Sum
m of theall negative cash flows
Projects with cumulative cash ratios >1 are potentially profitable and those with values <1
cannot be profitable. The NPV is a cumulative discounted cash position at the end of the
project life. The PBP and internal rate of return of the normal manufacturing plant is
shown in Table 15.6.
15.3.8.2 Discounted Profitability Criteria

The NPV is also known as the discounted cumulative cash position. The discounted cash
flow rate of return (DCFRR) is defined as the interest rate at which all the cash flows need
Table 15.6 Typical values of payback period and internal

rate of return of the plant.
Project with Payback period Internal rate

type of risk (years) of return (%)
High risk <2 20

Normal risk <5 15
Slight risk <10
to be discounted for the NPV of the plant to be equal to zero. A trial and error method is
used to estimate the interest rate, such that the initial investment cost would be reduced to
zero during the useful life of the project.
A discounted PBP estimates the number of years needed to break even considering the
initial expenses, discounting future cash flows and the future value of money with respect
to time. The most feasible plant should have the shortest discounted PBP.
15.3.9 Minimum Product Selling Price

If the conversion rate of raw material to product is low, this results in a higher projected
minimum product selling price (Kazi et al. 2010). The minimum price is essentially the
break‐even point for a given sale. Any pricing above the absolute minimum results in prof-
its. At the break‐even point, project profits equal zero.
The net profit can be calculated as:
Net profit Total revenue Total costs (15.14)
The total revenue is the product of the selling price of the product and production rate.
Technoeconomic evaluation of plant can be estimated based on the three cost management
system types: target costing, value engineering, and combined target costing and value engi-
neering. The value engineering technique aims at reduction of the production cost of a product
or specific service (Ellram 2006; Gnansounou and Dauriat 2010). It allows identification of the
main cost reduction possibilities, making cost enhancement replacements and determining the
best among them. For example, in the lignocellulosic production of ethanol, the pretreatment
stage is recognized as one of the vital steps for minimizing overall plant cost. Ammonia fiber
explosion, hot water treatment, and explosion with CO2 are considered as the most promising
pretreatment methods, because improved effectiveness of the process and reduction in pretreat-
ment costs enhance the flexibility of raw materials use and by‐product valorization.
15.4 Conclusion
In this chapter, we described the technoeconomic modeling approach for the conversion of
biomass into products. This approach is widely used in most case studies, although with a com-
pact timeline and additional extrapolations related to geographical area. A standard method of
technoeconomic analysis was presented, which allows organizations to decide on which pro-
jects to continue and how to optimize design to maximize profits. Methods of estimating plant
capital cost, working, and operating expenses were discussed. Discounted and nondiscounted
probability criteria were also discussed for estimation of rate of return, PBP, and NPV.
R
eferences
Banerjee, S., Tiarks, J.A., Lukawski, M. et al. (2013). Techno‐economic analysis of biofuel
production and biorefinery operation utilizing geothermal energy. Energy Fuels 27 (3):
1381–1390.
Bauman, C.H. (1964). Fundamentals of Cost Engineering in the Chemical Industries. New York:
Reinhold Publishing Corporation.
Brown, T.R. (2015). A techno‐economic review of thermochemical cellulosic biofuel pathways.
CEPCI (Chemical Engineering Plant Cost Index) (2019). www.tekim.undip.ac.id.
Cherubini, F., Jungmeier, G., Wellisch, M. et al. (2009). Toward a common classification
approach for biorefinery systems. Biofuels Bioproducts Biorefining 4: 1–13.
Chovau, S., Degrauwe, D., and Van‐Der‐Bruggen, B. (2013). Critical analysis of techno‐
economic estimates for the production cost of lignocellulosic bio‐ethanol. Renewable and
Sustainable Energy Review 26: 307–321.
Davis, R., Kinchin, C., Markham, J. et al. (2014). Process design and economics for the
conversion of algal biomass to biofuels: algal biomass fractionation to lipid‐ and
carbohydrate‐derived fuel products. Technical Report, National Renewable Energy
Laboratory(NREL).www.osti.gov/biblio/1159351‐process‐design‐economics‐conversion‐algal‐biomass‐
biofuels‐algal‐biomass‐fractionation‐lipid‐carbohydrate‐derived‐fuel‐products.
Dutta, A., Dowe, N., Ibsen, K.N. et al. (2016). An economic comparison of different
fermentation configurations to convert corn Stover to ethanol using Z. mobilis and
saccharomyces. Biotechnology Progress 26: 64–72.
Ellram, L.M. (2006). The implementation of target costing in the United States: theory versus
practice. Journal of Supply Chain Management 42 (1): 13–26.
Gnansounou, E. and Dauriat, A. (2010). Technoeconomic analysis of lignocellulosic ethanol: a
review. Bioresource Technology 101 (13): 4980–4991.
Gubicza, K., Nieves, I.U., Sagues, W.J. et al. (2016). Techno‐economic analysis of ethanol
production from sugarcane bagasse using a liquefaction plus simultaneous saccharification
and co‐fermentation process. Bioresource Technology 208: 42–48.
Gutiérrez, C.D.B., Serna, D.L.R., and Alzate, C.A.C. (2017). A comprehensive review on the
implementation of the biorefinery concept in biodiesel production plants. Biofuel Research
Journal 4 (3): 691–703.
Humbird, D., Davis, R., Hsu, T.L.K.C.D., and Aden, A. (2011). Process design and economics
for biochemical conversion of lignocellulosic biomass to ethanol‐dilute‐acid pretreatment
and enzymatic hydrolysis of corn stover. Technical Report, National Renewable Energy
Laboratory (NREL). www.nrel.gov/docs/fy11osti/47764.pdf.
Jungmeier, G., Hingsamer, M., and van Ree, R. (2013). Biofuel‐driven biorefineries: a selection
of the most promising biorefinery concepts to produce huge volumes of road transportation
biofuels until 2025. www.nachhaltigwirtschaften.at/resources/iea_pdf/iea_task_42_biofuel_
driven_biorefineries_lr.pdf.
Jungmeier, G., van Ree, R., Jorgensen, H. et al. (2014). The biorefinery fact sheet. https://www.
iea‐bioenergy.task42‐biorefineries.com/upload_mm/6/2/f/ac61fa53‐a1c0‐4cbc‐96f6‐
c9d19d668a14_BCI working document 20140709.pdf.
Kazi, F.K., Fortman, J.A., Anex, R.P. et al. (2010). Techno‐economic comparison of process
technologies for biochemical ethanol production from corn stover. Fuel 89: S20–S28.
Kumar, D., Long, S.P., and Singh, V. (2018). Biorefinery for combined production of jet fuel and
ethanol from lipid‐producing sugarcane: a techno‐economic evaluation. GCB Bioenergy 10
(2): 92–107.
Mejdell, A.L., Nygard, P., Ochoa‐Fernandez, E. et al. (2014). Sustainable products from economic
processing of biomass in highly integrated biorefineries: techno‐economic assessment and
market analysis. www.ifeu.de/wp‐content/uploads/Statoil__Biogasol_2014_Techno‐economic‐
assessment‐and‐market‐analysis‐of‐SUPRABIO‐biorefineries_2014‐01‐31.pdf.
Olcay, H., Malina, R., Upadhye, A.A. et al. (2018). Techno‐economic and environmental
evaluation of producing chemicals and drop‐in aviation biofuels via aqueous phase
processing. Energy & Environmental Science 11 (8): 2085–2101.
Peters, M.S. and Timmerhaus, K.D. (1991). Plant Design and Economics for Chemical Engineers,
4e. New York: McGraw‐Hill.
Quinn, J.C. and Davis, R. (2015). The potentials and challenges of algae based biofuels: a
review of the techno‐economic, life cycle, and resource assessment modeling. Bioresource
Quintero, J.A., Moncada, J., and Cardona, C.A. (2013). Techno‐economic analysis of
bioethanol production from lignocellulosic residues in Colombia: a process simulation
approach. Bioresource Technology 139: 300–307.
Rajendran, K. and Murthy, G.S. (2017). How does technology pathway choice influence
economic viability and environmental impacts of lignocellulosic biorefineries? Biotechnology
for Biofuels 10 (1): 1–19.
Shafiei, M., Karimi, K., and Taherzadeh, M.J. (2011). Techno‐economical study of ethanol and
biogas from spruce wood by NMMO‐pretreatment and rapid fermentation and digestion.
Shafiei, M., Kabir, M.M., Zilouei, H. et al. (2013). Techno‐economical study of biogas
production improved by steam explosion pretreatment. Bioresource Technology 148: 53–60.
Short, W., Packey, D.J., and Holt, T. (1995). A manual for the economic evaluation of energy
efficiency and renewable energy technologies. Technical Report, National Renewable
Energy Laboratory (NREL). www.nrel.gov/docs/legosti/old/5173.pdf.
Silva, J.F.L., Selicani, M.A., Junqueira, T.L. et al. (2017). Integrated furfural and first generation
bioethanol production: process simulation and technoeconomic analysis. Brazilian Journal
of Chemical Engineering 34 (3): 623–634.
Soleymani, M. and Rosentrater, K. (2017). Techno‐economic analysis of biofuel production
from macroalgae (seaweed). Bioengineering 4 (4): 92.
Taylor, G. (2008). Biofuels and the biorefinery concept. Energy Policy 36 (12): 4406–4409.
Turton, R., Bailie, R.C., Whiting, W.B. et al. (2012). Analysis, Synthesis and Design of Chemical
Processes, 4e. New York: Prentice Hall.
Vatavuk, W.M. (2002). Updating the CE plant cost index. www.chemengonline.com/Assets/
File/CEPCI_2002.pdf.
Vijayakumar, S. and Saravanan, V. (2015). Biosurfactants – types, sources and applications.
Research Journal of Microbiology 10 (2): 181–192.
Vlysidis, A., Binns, M., Webb, C., and Theodoropoulos, C. (2011). A techno‐economic analysis
of biodiesel biorefineries: assessment of integrated designs for the co‐production of fuels
and chemicals. Energy 36 (8): 4671–4683.
Wang, K., Brown, T., and Brown, R.C. (2014). Beyond ethanol: a techno‐economic analysis of
an integrated corn biorefinery for the production of hydrocarbon fuels and chemicals.
Biofuels Bioproducts and Biorefining 4: 190–200.
353
Index
a aspartic proteases 195

acetylation 138 aspen 319
acid detergent fiber (ADF) 32 assimilation 127
acid hydrolysis 34 autothermal gasifiers 54
acid impregnation 288
acid pretreatment 98 b
acrylic acid 125 β‐alanine pathway 222
acylation 138 β‐amylase 196
adipic acid 232 β‐glucosidases 193
adsorption 235 β‐mannanases 194
aerogels 256 β‐xylosidases 193
agricultural residues 50, 135, 316 bioactive compounds 18
air stripping 291 biocarbon 285
aldehydes and ketones 17 biocatalysis 206
alkaline and alkaline earth metals biocatalysts 206
(AAEMs) 287 biocatalyst technologies 205
alkaline pretreatment 98 biocatalyzed electrolysis 58
alkaline proteases 195 biochar 266, 273, 274, 299–300
alkali‐pretreated corncob (APC) 214 biochar in agriculture 275
alkoxylation 148 biochemical methods 269
allothermal gasifiers 54 biocomposites 151, 254
α‐amylase 196 biodegradation 125–126
ammonia fiber expansion (AFEX) 77 biodeterioration 127
amylases 196 biodiesel 2, 70
anaerobic bacteria 57 bioelectricity 2
anaerobic digestion 89 bioethanol 8, 13, 36, 318
arabinooligosaccharides 299 biofilms 172
arborform 254 biofragmentation 127
aromatic biopolymer 141 biofuels 7, 36, 47, 50
ash/ashes 271 biogas 55
aspartic acid 215 biohydrogen 2, 50, 55

354 Index
biological pretreatment 99, 203 cradle to gate 323

biomass 50, 88, 185, 268 cradle to grave 322
biomass integrated gasification combined crystallization 235
cycle (BIGCC) 114 cysteine proteases 195
biomass pyrolysis 52
biomass recalcitrance 128 d
bio‐oil 53, 104, 137, 285 dark fermentation 56
bioplastics 126 decision support system (DSS) 326
biopolymer 3, 125 degradable plastic 126
bioproducts 269 degree of polymerization (DP) 10
biorefinery/biorefineries 1, 8, 13, 333 dehydromucic acid 223
biorefinery technoeconomic evaluations 337 detoxification of levoglucosan 290–291
bioremediation 171 detoxification strategies 291
bioresources 313 dextran 134
biosurfactants 3, 15, 16 dimethyl furan (DMF) 317
2‐butenedioic acid 207, 212 direct carbon fuel cells (DCFC) 275
direct combustion 102
c direct liquefaction 52
capital cost 339 discounted profitability criteria 348
4‐carbon amino acid 215 distillation 235
carbon fibers 255 Dow‐DuPont 204
carboxylic acids 206 downstream processes 235
carboxymethylcellulose (CMC) 125, 150 dry distillation 271
catalytic fuel electrode 116
catalytic oxygen electrode 116 e
cellulase 193 elastic polymers 134
cellulose 10, 12, 30, 33–34, 50, 71, 91, 137, electricity 87
187, 203, 314 energy crops 51, 89, 135
cellulose microfibrils 186 energy‐driven biorefinery system 334
cellulosic polysaccharides 10 entrained flow reactors 54
char 273 environmental challenges 79
charcoal 277 enzymatic hydrolysis 13, 35
chemical pretreatment 98 enzymes 186
chromatography 235 esterification 17, 138, 148
circular economy 170 etherification 138
clean energy 69 european biochar certificate (EBC) 277
climate change 36 exocellobiohydrolase 193
co‐combustion 112 exoglucohydrolase 193
co‐firing 112 exopolysaccharide 134
co‐generation systems 285 extracellular polymers 146
combined pretreatment 99
combustion 89, 267–268 f
consolidated bioprocessing (CBP) 38 fast pyrolysis 104
conventional plastics 152 fatty acid methyl esters (FAMEs) 290
corn straw hydrolyzates 214 fibrillar phase 9
cradle to cradle 323 first‐generation bioethanol 38
Index 355
first‐generation biomass 50 higher alcohols 318

first‐generation biorefinery 313 higher heating value (HHV) 94
fixed capital investment (FCI) 334 high‐temperature fuel cell 116
fixed manufacturing costs 345 hybrid carbon fuel cells (HCFC) 275
flash pyrolysis 104 hydrogen 48
flowability 96 hydrophilic compounds 72
fluidized‐bed reactors 54 hydroxyethyl cellulose (HC) 150
forestry residues 135 hydroxymethylfurfural (HMF) 15
forest waste 315 hydroxyproline 12
fuel‐wood 50 3‐hydroxypropionic acid 222
fumaric acid 212, 213
furan 16 i
2,5‐furan‐dicarboxylic acid 223 I‐BIOREF 328
furfural 15 industrial waste 135
furfural resins 125 inhibitory compounds 291
internal combustion (IC) engine 112
g the international biochar initiative (IBI) 277
galactoglucomannans 189 intracellular polymers 146
galactomannan 188–189 inulinase 197
galacturonic acid (GalA) 11 invertase 197
gas turbine technology 114 ionic exchange 235
gasification 53, 89, 105. 206, 266, 267, 268 ionic liquid pretreatment 100
gasification reactors 54 itaconic acid (IA) 219
gasifier‐gas turbine combined technology 114
G‐lignin 249 k
global warming 21 kraft lignin 15, 255
glucaric acid (C6H10O8) 220
glucoamylases 196 l
glucomannan 188–189 laccase 194
gluconic acid 230 l‐aspartic acid 215
glutamic acid 227 levoglucosan 287
glutaminase 197 levulinic acid (LA) 210
glycolipids 161 life cycle analysis/assessment 4, 257, 314,
grasses 91 320, 322
greenhouse effect 29 life cycle inventory analysis 320
greenhouse gas emission 36 lignin 12, 13, 31, 50, 72, 91, 189, 203, 247,
greenhouse gases (GHG) 2, 29, 36, 47 249, 315
grindability 96 lignin‐based polyurethane 256
G‐S‐H‐lignin 249 lignin extraction 251
G‐S‐lignin 249 lignin‐modified graphene aerogels 256
gums and hydrocolloids 19 lignin peroxidases 194
lignin valorization 4, 248
h lignocellulose 185
hardwood 90 lignocellulose‐derived products 186
hemicellulose 11–12, 18, 31, 33, 50, 72, 91, lignocellulosic biomass 7, 13, 33, 51, 71,
188, 203, 314 89–91, 135, 269
356 Index
lignosulfonates 12, 254 organic acids 3, 16

lipases 195 organic dicarboxylic acid 218
liquefaction 102, 103, 267 organosolv pretreatment 100
liquid biofuels 69 overliming 291
liquid–liquid extraction 235 oxidative dehydrogenation 17
l‐malic acid 218 oxo‐degradable polymers 126
low‐carbon economy 48 4‐oxopentanoic acid 210
m p
malic acid (C4H6O5) 218 panel on climate change 87
malonyl‐coa pathway 222 pectin 11, 12, 19
manganese peroxidase 195 pectinase 196
mannan 188 perennial grasses 91
mannooligosaccharides 299 pervaporation 235
mannosylerythritol lipids 161 phenolic compounds 17
materia‐driven biorefinery system 334 phenolic resins 125
matrix phase 9 photofermentation 56, 58
membrane separation technologies 235 photosynthesis 267
metabolic engineering 77, 204 physical pretreatments 97
metallocarboxy proteases 195 physicochemical pretreatment 99
metalloproteases 195 phytases 196
methylcellulose 125, 150 plant cell wall 9, 32
2‐methylenebutanedioic acid 219 plant‐derived biomass 185
methylene succinic acid 219 polyacrylonitrile 255
microbial digestion 291 polyesters 134
microbial electrolysis cell 58 polyethylene 125, 151
microcrystalline cellulose (MCC) 138 polyhydroxyal‐kanoates 134, 150
microfibrils 187 polyhydroxybutyrate 125
middle lamella 8, 186 polylactic acid 125, 151
molten oxide carbonate fuel cell 116 polymers 126
polyols 15
n polysaccharides 8, 11, 12
nanocellulose/nanocrystalline cellulose pretreatment 55, 73, 97
138, 141 primary cell wall 8
natural biopolymers 128 primary refining process 333
natural gas 48 process flow diagram (PFD) 338
net present value 334 process simulation 319
neutral detergent fiber (NDF) 32 protease 195
nitric oxidation 220 proximate analysis 93
noncellulosic polysaccharides 11 purple nonsulfur bacteria 56
noncondensable gases 285 pyrolysis 89, 103, 266, 267–268, 272
nondiscounted profitability criteria 348 pyrolytic sugars 286
o r
oligosaccharides 299 recalcitrant 50
operating costs estimation 343 renewable resources 269
Index 357
residual biomass 313 succinic acid 207

residues from forest trees 50 sugar alcohol 14
resorcinol formaldehyde‐based carbon sugarcane 265
aerogels 256 sulfation 138
rhamnolipids 161 surfactants 15
roasting 267 syngas 206
s t
saccharic acid 220 tecnoeconomic assessment (TEA) 324
Saccharomyces cerevisiae 38 thermal conductivity 97
saponins 16 thermochemical conversion processes 4, 51,
secondary cell wall 8 89, 106, 112, 267, 335
secondary refining process 333 thermochemical decomposition 273
second‐generation biofuels 76 third‐generation biomass 50
second‐generation biomass 50 transesterification 70
second‐generation biorefinery 313
second‐generation ethanol/bioethanol 38, 285 u
separation techniques 237 ultimate analysis 94
serine carboxyl proteases 195 urban and domestic wastes 316
serine proteases 195
v
short‐chain carboxylic compounds 206
valeric biofuels 317
simultaneous saccharification and
valorization of lignin 258
fermentation (SSF) 38
value‐added products 333
slow pyrolysis 104
vanillin 17, 18
social challenges 79
softwood 90 w
solid oxide ceramic electrolytes 117 wastes from food processing 50
solid oxide fuel cell (SOFC) 116, 318 well to wheel 323
solid substrate/state fermentation 73, white‐rot fungi 195
168, 192 working capital estimation 342
solvent extraction 291
sophorolipids 161 x
steam cycle 111 xanthan 134
steam explosion 100 xylan 188
steam methane reforming (SMR) 60 xylanase 193
stirred tank reactors (STR) 168 xylooligosaccharides 299
submerged/liquid fermentation 73, 192 xylose 39, 78, 91

Lignocellulosic Biorefining Technologies Book

Uploaded by

Document Information

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Lignocellulosic Biorefining Technologies Book

Uploaded by

Copyright:

Available Formats

Lignocellulosic Biorefining Technologies

Lignocellulosic Biorefining Technologies

Anuj Kumar Chandel

Silvio Silvério da Silva

Limit of Liability/Disclaimer of Warranty

Library of Congress Cataloging‐in‐Publication Data

Set in 9.5/12.5pt STIXTwoText by SPi Global, Pondicherry, India

List of Contributors vii

1 Biorefining of Lignocellulose into Valuable Products 1

2 Bulk and Specialty Chemicals from Plant Cell Wall Chemistry 7

4 Production of Biohydrogen from Lignocellulosic Feedstocks 47

5 Recent Advances in the Production of Biodiesel Using Lignocellulosic Biomass 69

6 Bioelectricity Production from Lignocellulosic Biomass 87

8 Sustainable Production of Biosurfactants and Their Applications 159

11 Valorization of Lignin Into Value-Added Chemicals and Materials 247

13 I ntegrated Process of Biomass Thermochemical Conversion to Obtain Pyrolytic

Peyman Abdeshahian Shayaram Basumatary

Guadalupe Bustos Vázquez

Jeon Woon Choi

Bruna Curry Carneiro Douglas Faria

Luís António da Cruz Tarelho

Samar Das Geraldo Ferreira David

Diego Cunha Rocha Manuel Garcia-Perez

Paulo Eichler Euripedes Garcia Silveira Junior

Regina Célia Espinosa Modolo Fernanda Gonçalves

Pranab Goswami Sandeep Kumar

Indarchand Gupta Anuj Kumar Chandel

Victor Haber Perez Pankaj Kumar Rai

Avinash P. Ingle Valdemar Lacerda Jr

Genyr Kappler Paulo Ricardo Franco Marcelino

Vinayak Kulkarni Jaqueline Mattia

Mahdi Mazuchi Nathalia Ribeiro Ferreira da Silva

Carlos Alberto Mendes Moraes Luis V. Rodríguez-Duran

Paulo Roberto Wander Lourdes Zumalacárregui de Cárdenas

Beatriz Zumalacárregui de Cárdenas

Biorefining of Lignocellulose into Valuable Products

Lignocellulosic biomass is a major stakeholder in biorefineries. The transformation of

Lignocellulosic Biorefining Technologies, First Edition. Edited by Avinash P. Ingle,

As mentioned earlier, biohydrogen, biodiesel, and bioelectricity are imporant high‐value

Chandel, A.K. and Silveira, M.H.L. (2017). Sugarcane Bio‐Refinery: Technologies,

Bulk and Specialty Chemicals from Plant

Lignocellulosic Biorefining Technologies, First Edition. Edited by Avinash P. Ingle,

2.2 ­Plant Cell Wall Structure and Function

Figure 2.1 Plant cell wall structure and composition.

“tracheids in seedless vascular plants and gymnosperms and vessels in angiosperms”

2.3 ­Plant Cell Wall Chemical Composition

2.3.1 Cellulosic Polysaccharides

2.3.2 Noncellulosic Polysaccharides

2.3.2.2 Pectic Polysaccharides

2.3.3 Proteins and Glycoproteins

2.3.4 Phenolic Compounds

2.4 ­Valuable Chemicals Obtained from

Phenols and phenolics Gluconic acid

Arenes Organic acids

2.4.1 Sugar Alcohols

2.4.5 Organic Acids

other biomass transformations, such as hydrogenation or hydrogenolysis of polyols, which

2.4.6 Phenolic Compounds

2.4.7 Aldehydes and Ketones

2.4.8 Bioactive Compounds

2.4.9 Gums and Hydrocolloids

Chemical compounds Lignocellulosic source/ feedstock References

Furfural/hydroxy‐methylfurfural Hemicellulose/Pinus pinaster Peleteiro et al.

2.5 ­Conclusions and Final Remarks

13 I ntegrated Process of Biomass Thermochemical Conversion to Obtain Pyrolytic

2.2 Plant Cell Wall Structure and Function

2.3 Plant Cell Wall Chemical Composition

2.4 Valuable Chemicals Obtained from

2.5 Conclusions and Final Remarks

3.4 Lignocellulosic Biomass Utilization

3.5 Obtaining Fermentative Media

3.6 Fermentation of Second-Generation Sugars

4.2 Global Hydrogen Production and Use: Current Status

4.3 Biomass Importance and Contribution to Global

4.4 Hydrogen Production: Feedstock and Production

4.5 Biological Hydrogen Production Technologies

a ccumulate in the medium as it cannot be used by these anaerobes, thus lowering