New Biotechnology 39 (2017) 90–98

Contents lists available at ScienceDirect

New Biotechnology
journal homepage: www.elsevier.com/locate/nbt

Microalgae biorefineries: The Brazilian scenario in perspective

B.S.A.F Brasil* , F.C.P. Silva, F.G. Siqueira
Embrapa Agroenergy, Brasília/DF, Brazil

A R T I C L E I N F O A B S T R A C T

Article history:
Received 26 February 2015 Biorefineries have the potential to meet a significant part of the growing demand for energy, fuels,
Received in revised form 4 April 2016 chemicals and materials worldwide. Indeed, the bio-based industry is expected to play a major role in
Accepted 7 April 2016 energy security and climate change mitigation during the 21th century. Despite this, there are challenges
Available online 22 June 2016 related to resource consumption, processing optimization and waste minimization that still need to be
overcome. In this context, microalgae appear as a promising non-edible feedstock with advantages over
Keywords: traditional land crops, such as high productivity, continuous harvesting throughout the year and minimal
Microalgae strain development problems regarding land use. Importantly, both cultivation and microalgae processing can take place at
Microalgae cultivation
the same site, which increases the possibilities for process integration and a reduction in logistic costs at
Wastewater bioremediation
biorefinery facilities. This review describes the actual scenario for microalgae biorefineries integration to
Biofuels
Bio-based chemicals the biofuels and petrochemical industries in Brazil, while highlighting the major challenges and recent
Biomaterials advances in microalgae large-scale production.
ã 2016 Elsevier B.V. All rights reserved.

1. Introduction as bulk chemicals and materials [1]. Furthermore, the use of

The fast-growing human population and the consequent
growing demand for food, energy and water are and will continue
to be serious worldwide challenges for the coming decades [1]. In
addition, increasing concerns about anthropogenic climate change
and excessive dependency on fossil fuels are directing investment
towards more sustainable alternatives [2,3]. There is a consensus
that new technologies will be required to allow the conciliation of
economic growth and environmental sustainability in the long
term.
Biorefineries are facilities that integrate biomass conversion
processes into various marketable products and energy, while
optimizing the use of resources and minimizing waste, thereby
maximizing benefits and profitability [4,5]. The integration of
emerging biorefineries with other industries is a potential solution
to mitigate the threat of climate change and also provide means to
support the demand for energy, fuels, chemicals and materials [6].
Although countries like USA, Brazil and Germany already have
consolidated biorefineries, these are mainly based on the produc-
tion of ethanol and biodiesel from edible feedstocks such as corn,
sugarcane and soybean [5]. Thus, there is a need for innovative bio-
based products in order to substitute petroleum derivatives, such
* Corresponding author at: Av. W3 Norte (final), Asa Norte, Zipcode: 70770-901,
Brasília, DF, Brazil.
E-mail address: bruno.brasil@embrapa.br (B.S.A.F. Brasil).
alternative non-edible feedstocks is required to minimize the
competition for energy and food resources in the global market [7].
Recently, the world has experienced the birth of second-
generation ethanol commercial plants (using lignocellulosic
feedstocks), with the construction of the first facilities in Italy,
USA and Brazil. The use of lignocellulosic material resulting from
agro-industrial activities, such as sugarcane bagasse and straw, will
contribute to a more efficient use of land and proper environmen-
tal management [8]. However, there are concerns about logistics
costs and the indirect competition for the use of soil [8–10]. The
awareness of these constraints has driven attention and invest-
ment towards microalgae research and commercialization initia-
tives [11].
Microalgae are single-celled or colonial photosynthetic organ-
isms that are naturally present in different aquatic/humid
environments, including rivers, lakes, oceans and soils. They can
be used as raw material for the production of a plethora of
bioproducts, such as fuels, chemicals, materials, animal feed and
food supplements [12]. Algal biomass has considerable advantages
over traditional feedstocks: (i) High productivity—usually
10–100 times higher than land crops; (ii) Highly efficient carbon
capture; (iii) High lipid or starch content, which can be used for
biodiesel or ethanol production, respectively; (iv) Can be cultivated
in seawater, brackish water or even wastewater and (v) Can be
grown over non-arable land [10,13]. Microalgae can also be
harvested continuously throughout the year [14], which is the
reason for which they have the potential to form a continuous

http://dx.doi.org/10.1016/j.nbt.2016.04.007
1871-6784/ã 2016 Elsevier B.V. All rights reserved.
 B.S.A.F. Brasil et al. / New Biotechnology 39 (2017) 90–98 91

biofuel production chain such as in traditional oil refineries [9]. In
addition, both cultivation and microalgae processing can take
place at the same site, a characteristic that favors the integrated
and sequential production of various products and reduces logistic
costs at biorefinery facilities [1,3]. Nonetheless, there are signifi-
cant technological challenges to produce economically competi-
tive algal-derived biofuel, such as: (i) Microalgae strains
development (ii) Efficient methods for cultivation and crop
protection; (iii) Improvement of harvesting and lipid extraction
methods; (iv) Optimization of conversion/production processes of
fuels and coproducts [15–19].
2. Potential for microalgae production in Brazil
Brazil possesses a large tropical coastal area, 10,959 km in total,
has approximately 12% of the world’s freshwater supply, and
receives average insolation levels of 8–22 MJ/m2/day. The country
is also home to the world’s richest flora (40,989 species;
18,932 endemic) and possesses 3496 algae catalogued species
[20]. There are currently over forty laboratories and institutions
where algae cultures (microalgae, macroalgae and cyanobacteria)
are kept in Brazil. Five of these facilities hold a considerable
amount of strains (over 150 strains) [21,22].
The Brazilian companies Petrobras and Embrapa are leading
parallel initiatives aiming to characterize and domesticate
productive native algal strains. While Petrobras’ primary focus is
on marine microalgae for biodiesel production (See section
“Integration of microalgae production to the oil industry”),
Embrapa’s program is directed to continental microalgae genetic
resource characterization and the establishment of a long term
research program for biofuels and bioproducts production.
Furthermore, studies focused on native strains have reported
the isolation of Chlorella strains that present lipid productivity up
to 200 mg/L/day, which is comparable to the most promising
microalgae species isolated from other parts of the world, such as
Nannochloropsis gadinata and N. salina [23,24].
3. Microalgae production integration to the Brazilian fuel
industry
Considerable synergies do exist between microalgae produc-
tion/processing and a wide range of industries. There are
opportunities for the development of a sustainable microalgae-
based industry whose productivity is independent of soil fertility
and less dependent on water purity [19]. This section reviews the
current scenario and possibilities for microalgae cultivation/
processing in the fuel industry in Brazil. It is noteworthy, however,
that there is an array of initiatives in the context of aquaculture and
animal feed industries, rural and municipal wastewater treatment,
coal-fired power stations, among others not covered here, that are
currently being explored worldwide [5,13,19,25].
3.1. Integration of microalgae production to the sugarcane-ethanol
industry
There are currently 399 sugarcane-ethanol production facilities
in Brazil, mainly concentrated in the Northeastern and Southeast-
ern regions of the country. They were responsible for processing
658,820 Mt of sugarcane for the production of 27.96 billion liters of
ethanol, 37,880 Mt of sugar and 8870 MW of electricity in 2013.
Additionally, two second-generation facilities began operations in
2014. These plants have the capacity to produce 82 million and
40 million liters of cellulosic ethanol, respectively, using sugarcane
straw and bagasse as a feedstock (Fig. 1).
In 2014, the first industrial plant for the production microalgae
oils started commercial operations in Brazil [26]. The company
uses genetically modified microalgae cultivated in a closed
heterotrophic system to produce oil for renewable chemicals

Fig. 1. Microalgae production and processing in the context of sugarcane biorefinaries: Brazilian sugarcane biorefineries currently produce ethanol, sugar and electricity (gray
lines/icons). Microalgae production and processing can be coupled to sugarcane biorefinaries (black lines/icons), as shown in: (1) Renewable oils for industrial use and
personal care can be produced from sucrose by heterotrophic microalgae; (2) Vinasse and CO2 can be used to grow starch-rich algae, raw material for ethanol and electricity
production. In addition, these strains could be genetically modified to synthetize recombinant proteins such as cellulolytic enzymes. * The sugarcane straw processing for
cellulosic ethanol production was started in 2014 in Brazil** TRS: Total Recoverable Sugars. Based on information from references [4,25,31].
92 B.S.A.F. Brasil et al. / New Biotechnology 39 (2017) 90–98
(i.e.: Lubricants and cosmetics). The facilities, which have the
capacity to produce 100 Mt of renewable oils per year, use
sugarcane juice as carbon source for microalgae cultivation [26,34]
(Fig. 1).
Initiatives towards the use of sugarcane plant by-products, such
as vinasse and CO2, for microalgae mixotrophic cultivation are also
under way (Fig. 1). Vinasse is a liquid effluent generated at 10–15 L
per liter of ethanol produced, with a high organic load and acidic
pH. Carbon dioxide is produced in significant amounts during
sugarcane juice fermentation and bagasse burning. The proper
disposal and value aggregation of these effluents has been a
challenge for the sector. Indeed, there are technologies for
microalgae-based vinasse bioremediation and biomass production
currently at pre-commercialization stages in Brazil [28] (Fig. 1). In
addition, the genetic modification of microalgae strains for
recombinant cellulase production and secretion to the culture
medium might allow in house production of enzymes for cellulosic
ethanol production, thereby favoring biorefinery processes inte-
gration and costs reduction [29,30] (Fig. 1). Alternatively,
approaches focusing on the anaerobic treatment of vinasse prior
to algae cultivation have been shown to enhance Chlorella vulgaris
biomass productivity to 70 mg/L/d [31]. This approach can
integrate the production of methane, derived from the anaerobic
digestion process, to microalgae-based production of biofuels, such
as biodiesel and ethanol [31].
3.2. Integration of microalgae production to the biodiesel industry
Biodiesel production has been the most intensively studied area
of microalgae-based biotechnology worldwide, especially in the
last ten years [11,14,15–19]. In 2013, over 2.8 billion liters of
biodiesel were produced in Brazilian biodiesel plants, using mainly
soybean oil, animal tallow and cottonseed oil as feedstock [26].
Biodiesel production expansion in the last ten years has been
driven by an increase in the domestic demand for diesel coupled to
the gradual increment in the mandatory biodiesel blending.
Considerable attention has been given to promising alternative
feedstocks, such as oil palm, physic nut and microalgae, in order to
meet the demand forecasted for the next decades [27].
The use of biodiesel production by-products, such as glycerol
and POME (Palm Oil Mill Effluent), also offers attractive possibili-
ties to integrate oleaginous microalgae cultivation systems to
biodiesel plants. Cabanelas and collaborators [32] have shown that
native strains of Chlorella vulgaris and Botryococcus terribilis
presented considerable biomass productivities using domestic
wastewater supplemented with glycerol (50 mM) as cultivation
medium. In another study, Liang and collaborators [33] reported
that the glycerol supplementation at a concentration of 100 mM
increased C. vulgaris biomass productivity from 10 to 102 mg/L/day.
Furthermore, the use of POME, either crude or anaerobically
digested, for culturing microalgae is a promising alternative to
treat this effluent while producing energy-rich biomass [34–37].
3.3. Integration of microalgae production to the oil industry
Brazil’s proven oil reserves, including large offshore fields
located deep under pre-salt layers at the country’s coast,
comprised 15.6 billion barrels in 2014 (around 0.9% of the world’s
proven oil reserves). The exploration of these reserves might place
the Brazilian oil industry as one of the ten top producers
worldwide in the near future [38].
Produced water (PW) is the largest waste stream generated in
oil and gas industries. This geologic water has varying concen-
trations of impurities such as inorganic salts, hydrocarbons and
added elements used in the separation of water and oil in the
production line [39]. The effect of discharging produced water on
the environment is an issue that causes significant environmental
concern [40]. The Brazilian oil company, Petrobras, has been
making investments focused on microalgae-based treatment of
produced water and CO2 emissions capture at oil exploration
platforms (Fig. 2). In 2012, the company’s first microalgae
cultivation plant started pre-commercial operations at the
Northeastern coastal region, taking advantage of its favorable
climate and high number of sunny days. An native Nannochloropsis
oculata strain has been adapted for growth in PW leading to a
higher growth rate (m = 0,22/d) in PW diluted at 50% in f/2 medium
compared to the control (m = 0,08/d) grown in 100% f/2 medium
[41].
Recent advances in hydrothermal liquefaction (HTL) processes
for whole microalgae biomass conversion into biocrude oil have
opened new perspectives regarding the production of microalgae-
based energy, chemicals, materials and biofuels (Fig. 2) [16]. This
thermochemical technology can convert whole algal wet biomass
directly, thereby reducing the cost of biomass drying and lipid
extraction [16,42]. In the future, it might be possible to process
algae biocrude oil together with petroleum in order to provide raw
material for more sustainable alternatives to conventional oil
derivatives (Fig. 2).
4. Challenges for microalgae large-scale production
In spite of the promising opportunities envisioned for micro-
algae-integrated biorefineries, the economic viability of large-scale
algal biomass cultivation for low-value products (e.g.: Biofuels,
bulk chemicals and biomaterials) has not been achieved yet.
Indeed, even considering recent technologic advances, the costs for
microalgae biofuel production are still at least 2-fold higher than
for its fossil-based counterparts (Table 1). Therefore, continued
enhancements are needed in algal biology, as well as in the areas of
algal cultivation, harvesting and processing. In addition, after
identifying economically viable biorefinery combinations, envi-
ronmental risks and resources management analysis should be
considered in order to determine the feasibility of the technologies
and their combinations in a proposed area [46,52]. In this section
the recent advances related to microalgae biorefineries are
critically discussed in the light of their large-scale techno-
economic feasibility.
4.1. Microalgae biomass cultivation
Microalgae are usually cultivated under photoautotrophic
conditions, using the photosynthetic metabolic pathway to capture
energy from light while fixing inorganic carbon (e.g.: CO2) [53,54].
However, there are several microalgae species that can also grow in
the absence of light using organic carbon compounds as both
energy and carbon sources in a condition known as heterotrophic
cultivation [53,55]. Additionally, some microalgae can grow using
both photoautotrophic and heterotrophic modes combined
(mixotrophic cultivation), where organic and inorganic carbon
sources are metabolized in the presence of light [54]. All three
growth modes have been reported for microalgae cultivation using
oil/biofuel industries by-products as culture media substrates
(Table 2).
Heterotrophic microalgae cultivation is usually conducted
inside opaque fermenters [53] and tends to achieve higher
productivities than photoautotrophic and mixotrophic modes
(Table 2). Furthermore, since light independent growth is not
diminished by the increase in the medium turbidity during
microalgal cultivation, higher cell densities can be obtained
leading to reduced harvesting costs [66]. However, issues related
to the high substrate costs and high risks of contamination
preclude large-scale heterotrophic microalgae cultivation for
 B.S.A.F. Brasil et al. / New Biotechnology 39 (2017) 90–98 93

Fig. 2. Microalgae production and processing in the petrochemical sector: Brazil currently explores oil in both off shore and on shore platforms, giving rise to significant
amounts of effluents, such as produced water and CO2 (gray icons). Microalgae can be cultivated using both of these effluents to generate oil-rich biomass (1). Algae extracted
oil or algae biocrude, which is generated by the hydrothermal treatment of the whole algae biomass, can be used as feedstock for the production of electricity, biofuels, bio-
based chemicals and biomaterials (black icons). In a future scenario, oil refineries might be enabled to receive algae feedstock becoming actual biorefineries. Based on
information from references [38–40].

biofuels [53,67,68]. Instead, heterotrophic microalgae have been
commercially used only for the production of higher value
products (i.e.: cosmetics and lubricants) (Fig. 1).
On the other hand, photoautotrophic cultivation is the most
widely mode used for microalgae biomass cultivation [53,69–72].
It is performed in photobioreactors (PBRs), which can be of either
open or closed types [53]. It is still not clear which production
system is the best for commercial applications. However, due to
their low capital and operational costs, open PBRs (e.g.: Open
ponds and raceways) are often reported as less costly (Table 1).
Indeed, open PBRs are cheaper and easier to build and operate, but
present lower biomass productivity [14,53]. Furthermore, they are
dependent on the climatic conditions and are continuously
threatened by invading species, such as non-target algae, grazers
and bacteria, indicating that further improvements will be
required for large-scale biomass production [53]. Recently, a novel
open cultivation system, named Algae Raceway Integrated Design
(ARID), has been demonstrated to provide superior temperature
and energy management than conventional raceways, which leads
to significant cost reduction, especially at subtropical/temperate
latitudes [46,73].
Closed PBRs (e.g.: Tubular reactors and flat panels) present
relatively higher biomass productivities, better temperature,
better contamination control and more efficient CO2 capture than
open systems [15,18]. However, scalability limitations due to gas
exchange requirements and the high installation and operation
costs leads to a lower economic feasibility of closed systems
[15,18,74,75]. Indeed, as demonstrated in Table 1, closed systems
are comparatively more expensive than open pond/raceways. A
detailed explanation of differences between these two modes has
been extensively reviewed elsewhere [76–78] and is beyond the
scope of this article. Hybrid systems, on the other hand, have
presented promising results in large-scale production of marine
microalgae as well as in economic viability analysis [79] (Table 1).
Such systems take advantage of using closed PBRs to produce cell-
dense microalgae inoculum in a contamination protected environ-
ment followed by large-scale mass production in low cost/easily
operated open systems [79].
Significant cost reductions (>50%) can be achieved if CO2,
nutrients and water for microalgae cultivation are obtained at low
cost [51] (Table 1), which makes the biorefinery strategies
described in here (see section “Microalgae production integration
to the Brazilian fuel industry”) particularly attractive. Photoauto-
trophic microalgae cultivation using residues or wastewater is
usually achieved by reducing the substrate organic load and
increasing nutrient availability through previous substrate pre-
treatment (e.g.: Anaerobic digestion) [31]. This strategy is being
currently explored worldwide, especially through the combination
of urban/agriculture wastewater treatment and microalgae bio-
mass production [reviewed in 80,81].
Regarding the use fuel industry by-products as media for
microalgae growth, though, the most commonly reported cultiva-
tion mode is mixotrophy (Table 2). Mixotrophic microalgae have the
potential to capture CO2 emissions while lowering the organic load of
the medium during growth [54]. Although theoretically all PBRs
systems could be used for mixotrophic cultivation, the high organic
load associated with the culture medium will probably hamper the
use of large-scale systems based solely in open ponds/raceway. Thus,
the use of a two-stage heterotrophic/mixotrophic and photoauto-
trophic culture strategy, where the microalgae seed is produced in
fermenters (heterotrophic) or closed PBRs (mixotrophic) and
subsequently inoculated in open photoautotrophic systems, appears
to be the most promising approach to manage contaminants when
using media with high organic load [68].
94 B.S.A.F. Brasil et al. / New Biotechnology 39 (2017) 90–98

Table 1
Minimum sale prices for microalgae intermediate products and drop-in biofuels according to the technologic processes used. “N.A.”: Not Available.

Cultivation System Harvesting Extraction/Conversion Production cost Reference

Open Ponds Filter press dewatering
Hybrid (Closed PBR/Open ponds) Flotation
+Centrifugation
Hybrid (Closed PBR/Open ponds) Filter press dewatering
Closed PBR Flotation
+Centrifugation
Hybrid (Closed PBR/Open ponds) Filter press dewatering
Algae Raceway Integrated Design Electrocoagulation
+ GMO strain
Hybrid (Closed PBR/Open ponds) Filter press dewatering
Open Ponds Dissolved air flotation
+ Centrifugation
Open Ponds Flocculation
+ Centrifugation
Open Ponds Dissolved air flotation
+ Centrifugation
Closed PBR Dissolved air flotation
+ Centrifugation
Closed PBR Dissolved air flotation
+ Centrifugation
Closed PBR Dissolved air flotation
+ Centrifugation
Closed PBR Dissolved air flotation
+ Centrifugation
Open Ponds Flocculation
+ High productivity + Centrifugation
+ Free nutrients/CO2
Open Ponds Flocculation
+ Centrifugation
Closed PBR Flocculation
+ High productivity + Centrifugation
+ Free nutrients/CO2
Closed PBR Flocculation
+ Centrifugation
Wet solvent lipid extraction 8,79 US$/gallon of algal biodiesel [43]
+ Oil hydrotreating
+ Anaerobic digestion of residues
Hydrolysis 13,94 US$/gallon of algal biodiesel [44]
+ In situ transesterification
+ Flash separation
Wet solvent lipid extraction 14,12 US$/gallon of algal biodiesel [43]
+ Oil hydrotreating
+ Anaerobic digestion of residues
Hydrolysis 87,14 US$/gallon of algal biodiesel [44]
+ In situ transesterification
+ Distillation
Hydrotermal liquefaction 7,30 US$/gallon of algal biocrude [45]
+ Catalytic hydrothermal gasification
+ Solids to animal feed
Hydrotermal liquefaction 7,50 US$/gallon of algal biocrude [46]
+ Catalytic hydrothermal gasification
Hydrotermal liquefaction 9,04 US$/gallon of algal biocrude [45]
+ Catalytic hydrothermal gasification
Wet solvent lipid extraction 8,52 US$/gallon of algal TAG [47]
+ Oil hydrotreating
+ Anaerobic digestion of residues
Hot algal lipid stream extraction 10,87 US$/gallon of algal TAG [48]
+ Anaerobic digestion of residues
Wet solvent lipid extraction 12,73 US$/gallon of algal TAG [49]
+ Oil hydrotreating
+ Anaerobic digestion of residues
Wet solvent lipid extraction 18,10 US$/gallon of algal TAG [47]
+ Oil hydrotreating
+ Anaerobic digestion of residues
Wet solvent lipid extraction 31,61 US$/gallon of algal TAG [49]
+ Oil hydrotreating
+ Anaerobic digestion of residues
Hydrothermal treatment 77,00 US$/gallon of algal TAG [50]
+Wet solvent lipid extraction
Hydrothermal treatment 109,00 US$/gallon of algal TAG [50]
+Wet solvent lipid extraction
N. A. 0,44 US$/kg of algal biomass [51]
N. A. 1,98 US$/kg of algal biomass [51]
N. A. 4,19 US$/kg of algal biomass [51]
N. A. 11,02 US$/kg of algal biomass [51]

Indeed, contamination control and crop protection pose a major
challenge for microalgae large-scale cultivation, especially if the
use of waste streams rich in organic compounds is considered
[54,82–84]. In line with this assumption, industrial residues/
effluents are often pretreated in order to favor microalgae growth
over contaminants. The pretreatment processes used can be simple
dilutions, pH corrections and suspended solids settling as well as
clarification, hydrolysis, ultraviolet irradiation, anaerobic diges-
tion, filtration, heavy metals removal, among others [31,32,56–
65,84] (Table 2). In addition, a number of strategies can be
deployed to mitigate contamination during the microalgae
cultivating course [82,85,86]. For example, cyanobacteria and

Table 2
Productivities of microalgae strains cultivated using biofuel and petrochemical industries by-products as culture media substrates. “N.A.”: Not Available.

Microalgae growth mode Substrate/Medium Microalgae Species Biomass Productivity Lipid productivity Reference
Heterotrophic Pretrated sugarcane molasse Chlorella zofingiensis 1,55 g L 1 d 1 0.710 g L 1 d 1
[56]
Mixotrophic Corn ethanol thin stillage supernatant Chlorella vulgaris 2,5 g L 1 d 1 1,1 g L 1 d 1 [57]
Mixotrophic Diluted crude glycerol Botryococcus braunii 0,271 g L 1 d 1 0,044 g L 1 d 1
[58]
Mixotrophic Diluted technical glycerol Scenedesmus sp. 0.229 g L 1 d 1 0,031 g L 1 d 1
[59]
Mixotrophic Diluted technical glycerol Nannochloropsis sp. 0,074 g L 1 d 1 0,015 g L 1 d 1
[60]
Mixotrophic Diluted sugarcane molasse Botryococcus braunii 0.184 g L 1 d 1 0.068 g L 1 d 1
[61]
Mixotrophic Diluted sugarcane vinasse Scenedesmus sp. 0,064 g L 1 d 1 N.A. [62]
Mixotrophic Diluted beet vinasse Spirulina maxima 0,240 g L 1 d 1 N.A. [63]
Mixotrophic Glycerol diluted in domestic wastewater Botryococcus terribilis 0,282 g L 1 d 1 0,035 g L 1 d 1
[32]
Mixotrophic Produced Water Cyanobacteria (Oscillatoriales) 4,8 g m2 d 1 N.A. [64]
Mixotrophic Treated produced water Microalgae polyculture 11 g m2 d 1 N.A. [65]
Photoautotrophic Anaerobically digested vinasse Chlorella vulgaris 0,070 g L 1 d 1 0,017 g L 1 d 1
[31]
 B.S.A.F. Brasil et al. / New Biotechnology 39 (2017) 90–98
rotifers invaders can be reduced in Nannochloropsis salina cultures
by setting salinity to 22 ppt and using ammonium chloride as a
nitrogen source [65]. Pesticides such as parathion and dichlor-
odiphenyltrichloroethane (DDT) have also been successfully used
in Chlorella cultures to control infestation by Copepoda crustaceans
with no significant toxicity for the algae [87]. Furthermore, the
management of contaminants in large-scale production systems
using microscopic and molecular methods for real-time tracking of
pests combined with specific crop protection strategies (e.g.:
pesticide application) have been recently demonstrated [85,88].
It is important to highlight though, that maintaining axenic
conditions (algal monocultures without other microorganism’s) in
algal culturing systems would neither be practical nor economic
viable, since a large diversity of microbial organisms will inevitably
be present, even in treated cultures/substrates [83]. Therefore, the
selection of robust microalgae strains capable of achieving and
maintaining high growth rates in exposed cultivation systems even
in the presence of a small percentage of contaminants will be
required. These strains will probably be specific for each substrate/
location as indicated by the higher productivities obtained with
indigenous microalgae grown in produced water [64,65] shown in
Table 2. In addition, in order to maximize microalgae culture
resilience and productivity, the use of engineered polycultures
where diverse organisms are grown together in a synergistic
manner should be considered [89]. Also promising is the use of
breeding and genetic engineering (GMO) techniques to develop
strains with traits that provide crop protection and fast efficient
growth [reviewed in 15,90]. Indeed, a two-fold increase in the
biomass productivity has been reported in transgenic Chlamydo-
monas reinhardtii strains capable of self-adjusting the ratios of
their chlorophyll binding proteins in response to changing light
levels or culture densities [65]. Financial modeling based on these
strains productivities indicated that 240% increase in receipts and
85% reduction in the cost algal biocrude would be achieved
compared to the non-GMO baseline case [46].
Finally, since spills of cultured microalgae are expected to
eventually occur from biomass production facilities, the risk of
releases into natural ecosystems must be considered for large-
scale cultivation. In addition, as massive spills might also happen
due to extreme climatic events (e.g.: Storms, floods and earth-
quakes), strategies to mitigate cultured strains leakage using
physical containment and genetic modifications that preclude
cultured microalgae from competing in nature should be applied
either to transgenic or nontransgenic algae [52,91]. The mitigation
strategy employed should be carefully evaluated case-by-case
considering the microalgae strain chances of surviving and
reproducing in nature as well as its capability of causing
irreversible, or slowly reversible, impacts in the environment
[52]. For example, a strain carrying traits that enhance competitive
fitness both in artificial systems and in the wild, such as improved
nutrient uptake or prevention of contamination by toxin produc-
tion [92], should be classified as high risk. On the other hand, traits
that enhance capture of light [93,94], which is usually not a
limiting factor in nature, or transgenic herbicide resistance, as long
as the herbicide is not generally used in the environment, could be
classified as low risk [52]. Additionally, assessing the environmen-
tal risks of uncontrolled non-native strains releases can be
especially challenge, since large numbers of species with varying
degrees of fitness co-exist in natural ecosystems in an unpredict-
able balance [95,96]. Thus the most stringent mitigation systems
are needed both for non-native species and for selected/
engineered strains that possess enhanced fitness into the wild.
In such cases, genetic modifications that allow cultivation only in
artificial systems and are lethal to the algae in nature (e.g.:
disruption/reduction of carbon capturing or nitrate utilization) will
probably be required [52].
95
4.2. Microalgae biomass harvesting and conversion
After cultivation, the algal biomass must be harvested and
converted into biofuels and bioproducts. The choice of the
downstream processes used greatly influences the economic
viability, efficiency, operating costs and energy requirements of
microalgae biorefineries [97] (Table 1). Harvesting is generally a
multistage process resulting in the concentration of algae in water,
being responsible for up to 30% of the total production costs. It
generally involves one or more steps of solid-liquid phase
separation, such as flocculation, filtration, flotation, settling and/
or centrifugation [98]. Centrifuges are a proven technology that has
been used as a means for harvesting algae for many years [74].
However, the high energy requirements and the costs to centrifuge
large amounts of water reduce the economic viability of this
process [97]. Therefore, harvesting methods with low-costs and
high processivity are required to dewater algal biomass prior to
lipid extraction or biomass conversion.
Harvesting techniques based on the use of chemical flocculants
and dissolved air flotation are low-cost methods that have been
successfully applied either to wastewater treatment or to cultured
microalgae [99–101]. Especially promising are methods based on
the use of non-toxic flocculants such as mixed calcium and
magnesium hydroxides [102]. In addition, recent economic
assessments indicate that methods based on membrane filtration,
ultrasound and electrocoagulation harvesting have the potential to
achieve relatively low costs. Among them, electrocoagulation is the
most mature technology with commercial applications existing in
the wastewater treatment industry to remove a variety of charged
ionic contaminants from solution [97,103].
Electrocoagulation uses metallic electrodes to produce posi-
tively charged ions that induce coagulation of microalgal cells,
which are negatively charged. The settled coagulated cells can then
be removed by conventional sedimentation methods [97]. Electro-
coagulation systems can achieve 95% recovery of algae using only
25% of the energy compared to centrifuge technology [16,97,103].
However, the after harvesting microalgae solution generated
possesses up to 6% total solids when starting with an algae
solution of approximately 0.1% total solids [97,103]. Consequently,
electrocoagulation may serve as a primary harvesting methodolo-
gy that can be used in combination with further concentrating
methods (e.g.: Membrane filtration or centrifugation) to achieve
higher concentration factors or higher throughput.
The technological pathway traditionally envisioned for down-
stream processing of microalgal biomass involves biomass
harvesting and drying followed by the extraction of the lipidic
fraction and conversion to biofuels (i.e.: biodiesel). The remaining
lipid extracted algae (LEA) fraction would be directed to the
production of fertilizers, animal feed or other bioproducts [16,19].
This model, however, has been challenged by recent advances in
extraction/conversion processes capable of handling wet feed-
stocks, as well as, converting whole algal biomass into oil
[16,46,97,104] (Table 1).
The prohibitive costs and energy requirements of algal biomass
drying have led to the development of processes capable of
extracting lipids from wet biomass, such as the wet solvent
extraction process. This process pretreats harvested algae biomass
in water with conditioning chemicals to enable non-polar solvent
to remove lipids (Valicor Renewables, LLC). The extracted algal
mass is then phase separated and lipids are recovered from the
nonpolar solvent by distillation [97]. The economic viability of
using only the lipidic fraction for the production of biofuels while
converting the LEA fraction into other products (i.e: fertilizers and
animal feed) is controversial though [19,45,46]. Furthermore, the
use of wastewater for cultivating microalgae might render algal
96 B.S.A.F. Brasil et al. / New Biotechnology 39 (2017) 90–98
biomass unsuitable for the production of animal feed or food
supplements.
Recent studies indicate that whole wet algal biomass conver-
sion into oil through hydrothermal liquefaction and catalytic
hydrothermal gasification (HTL-CHG) might be the most promising
downstream processing pathway [16,46,97,104] (Table 1). Indeed,
Richardson and coworkers [97] reported that the revenues for a
scenario using HTL–CHG are 69% higher than using wet solvent
extraction. In addition, the combination of electrocoagulation
harvesting and HTL–CHG can lead to 90% lower lipid total cost
compared to centrifugation followed by wet solvent extraction
[46,97].
The HTL-CHG, which functions both as an extraction and as a
conversion technology, converts not only the algal lipids, but also
part of proteins and carbohydrates into oil that can be upgraded to
biofuels (e.g.: renewable diesel, jet fuel, gasoline) [104]. The total
oil yield from algae is generally in the range from 35% to 65% as
measured by weight of oil as a percent of feedstock dry weight,
which is considerably higher than the yields obtained with
methods that use only the lipidic fraction of algal biomass
[97,105]. HTL-CHG processing converts algal slurries with approxi-
mately 20% of solids in water into oil by using temperatures and
pressures just below the supercritical point of water. No acid
pretreatment, solvent, or separations of acid and solvent are
needed [105]. The wet feedstock is first processed via HTL to
produce oil leaving an aqueous effluent containing organic matter.
This effluent is then processed via CHG to gasify the remaining
organic matter producing methane gas and to recycle water and
nutrients, such as nitrogen, potassium, phosphorous and micro-
nutrients [97,105].
However, even though HTL-CHG technology looks promising,
future works focused on large scale demonstrations are still
required. Furthermore, for carbohydrate-rich algae biomass, it
might be more efficiently to biochemically convert feedstock into
ethanol and bulk chemicals instead. In such cases, processing of
algal biomass through hydrolysis followed by ethanol fermenta-
tion or mixed organic acid fermentation followed by ketonization
and catalytic upgrading to aromatics should be considered [16].
5. Conclusions
The biofuel and petrochemical refineries provide promising
possibilities for the exploration of microalgae feedstock integra-
tion to existing industrial facilities. Although still in its infancy,
microalgae large-scale production might provide valuable resour-
ces for future industrial ecology, bringing many environmental
deliverables such as reduction of fossil fuel usage, mitigation of
green-house gas emissions, better land use and improvement in
food security. However, in order to achieve commercial viability,
several innovative technological solutions are still needed to
overcome the issues related to microalgae strains development,
cultivation systems, crop protection, harvesting, processing,
environmental risks and resources management. Therefore, we
emphasize the need for large scale demonstrations to increase the
confidence in algae-based and integrated biorefineries. Hopefully,
the present and future R&D investments, policy initiatives and
public-private partnerships should push forward the development
of economically viable microalgae biorefineries setting the course
towards a future carbon-smart society.
Acknowledgements
The authors are grateful to the Empresa Brasileira de Pesquisa
Agropecuária (EMBRAPA),Coordenação de Aperfeiçoamento de
Pessoal de Nível Superior (CAPES), Financiadora de Estudos e
Projetos (FINEP) and Conselho Nacional de Pesquisa (CNPq) for
supporting this work. We also thank Maria Goreti Braga dos Santos
for assistance with the figures edition. The sponsors had no
involvement in study design; in the collection, analysis and
interpretation of data; in the writing of the report or in the decision
to submit the article for publication.
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