Biosurfactant production using 2 nd generation feedstocks

M. Guillén, E. Genescà, B. Lechuga, N. Alcover, A. Surribas

LEITAT Technological Center – C/ de la Innovació, 2 – 08225 –Terrassa – Barcelona – (Spain) www.leitat.org
mguillen@leitat.org

. INTRODUCTION
Surfactants are compounds that lower the surface tension either between liquids or liquids and solids, allowing easier spreading. Surfactants are widely used in
detergents or as wetting agents, emulsifiers, foaming agents, and dispersants. The most common surfactants are molecules from petrochemical origin which after
application are discharged into sewage systems or directly into surface waters causing a negative environmental impact. Biosurfactants are surface-active compounds
produced either intra- or extracellularly by a wide range of microorganisms. These biomolecules have properties that allow their use in a great number of industrial
applications and the replacement of chemically synthesized surfactants. However, the production cost of these biocompounds is one of the main facts that impair the
replacement of chemical surfactants by these natural molecules.

OBJECTIVE
The objective of the present work is to produce biosurfactants out of biomass which is not competing with the food chain. Different non-edible fats & oils and
different lignocellulosic sources will be the feedstock (“ingredients”) of the 2nd generation biomass process. Additionally, the present work aims to obtain a process up
to 30% cheaper and 30% more sustainable than the corresponding chemical routes or the biotechnology processes starting from 1st generation edible feedstocks.

RESULTS
•Screening of hydrophilic and hydrophobic carbon sources from lignocellulosic compounds and non-edible fats and oils.
The screening of the hydrophilic and the hydrophobic carbon sources using the available feedstocks was carried out in shake flasks at 30°C and 150 rpm during
144h. The producing microorganism was precultured in a commercial rich media . The obtained results (table 1) show that the best productivity is achieved using
the lignocellulosic feedstock 1 and the fat&oil feedstock 1, obtaining 2.11 and 1.33-fold less productivity compared to the control culture (Table 1).
Table 1. Productivity obtained using different hydrophilic and hydrophobic carbon sources as substrates
HydrophIlic carbon source Hydrophobic carbon source Productivity (g biosurfactant/L·h)
Glucose Oleic acid 0.211 ± 0.032
Lignocellulosic 1 Oleic acid 0.10 ± 0.015
Lignocellulosic 2 Oleic acid 0.034± 0.008
Glucose Fat&oil 1 0.158± 0.023
Glucose Fat&oil 2 0.048± 0.021
Glucose Fat&oil 3 0.066± 0.012
Glucose Fat&oil 4 0.114± 0.022
Glucose Fat&oil 5 0.110± 0.007

•Fed-batch strategy for biosurfactant production

Fed-batch strategy was applied for the study of the biosurfactant production at bioreactor scale. The fed-batch process was carried out using glucose and
oleic acid as control substrates (Fig. 1). The lignocellulosic 1 and the fat and oil 1 were used as alternative hydrophilic and hydrophobic carbon sources,
respectively (Fig. 2-3). The main components of the media were 100g/L of the hydrophilic carbon source, 10g/L of the hydrophobic carbon source and 5g/L of
the nitrogen carbon source. The hydrophobic carbon source was maintained at a maximum value of 10g/L during the culture. The fed-batch phase was started
when the hydrophilic carbon source reach a value of 30g/L. Hydrophilic substrate additions were carried out to maintain its concentration at 30g/L during the
fed-batch phase. The temperature, aeration and shaking were maintained at 30°C, 1vvm and 500 rpm, respectively.

120 120 120 90 120 80

80 70
100 100 100 100
70
60
Hydrophobic 1 (g/L)
Biosurfactant (g/L)

Biosurfactant (g/L)

Biosurfactant (g/L)
Hydrophilic 1 (g/L)

80 80 80 60 80
oleic acid (g/L)

Oleic acid (g/L)

Glucose (g/L)

Biomas (g/L)

Glucose (g/L)

Biomas (g/L)

50

Biomas (g/L)
50
60 60 60 60 40
40
40 40 30
40 30 40
20 20
20 20 20 20
10 10
0 0 0 0 0 0
0 20 40 60 80 100 120 0 20 40 60 80 100 120 0,00 20,00 40,00 60,00 80,00 100,00 120,00
time (h) time (h) Time (h)
Oleic acid (g/L) Glucose (g/L) Biosurfactant (g/L) Biomas (g/L) Hydrophobic 1 (g/L) Glucose (g/L) Biosurfactant (g/L) Biomas (g/L) Hydrophilic 1 (g/L) Oleic acid (g/L) Biosurfactant (g/L) Biomas (g/L)

Fig1. Fed-batch control using glucose and oleic acid Fig2. Fed-batch control using glucose and hydrophobic 1 Fig3. Fed-batch control using hydrophilic 1 and oleic acid

Table 2. Productivity obtained using different hydrophilic and hydrophobic carbon sources
The replacement of the commercial hydrophilic carbon source by the lignocellulosic 1 affects as substrates
more negatively the productivity than the replacement of the commercial hydrophobic carbon Fed-batch Biosurfactant [g/L] Productivity (g biosurfactant/L·h) % Productivity
source by the fat&oil 1 (Table 2). Control 98,00 0,83 -
The productivity decreased 16% using the alternative hydrophobic carbon source. On the other Fat&oil 1 82,87 0,70 84,31
hand, the productivity decreased 29% using the alternative hydrophilic carbon source (Table 2). Lignocellulosic 1 66,90 0,59 71,08

MAIN CONCLUSION
•Among the different lignocellulosic and fat&oil feedstocks tested in shake flasks cultures, the lignocellulosic 1 and the fat and oil 1 were the best carbon sources for the
biosurfactant production.
•During the fed-batch bioprocesses carried out, the use of the fat and oil 1 feedstock instead of the commercial hydrophobic carbon source led to higher productivity
compared to the used of the lignocellulosic 1 as an alternative for the commercial hydrophilic carbon source.
This project has received funding from the European Union’s Seventh Framework Program for research, technological development and demonstration under grant agreement 289194 www.bioconsept.eu