Production of Biobutanol from Jatropha Seed Cake...23© Reliance Industries Ltd., 2013 Butanol Titers Using Biomass Biomass Hydrolysate Additives in medium Butanol titer (gL-1) Productivity

Jasmine Isar

Production of Biobutanol from

Jatropha Seed Cake

2 © Reliance Industries Ltd., 2013

The Reliance Group, founded by Dhirubhai H. Ambani (1932-2002), is India’s largest private

sector enterprise with businesses in the energy and materials value chain, and having annual

revenues in excess of US$ 68 billion. The flagship company, Reliance Industries Limited (“RIL”),

is a Fortune Global 500 Company and enjoys global leadership in its key product lines. RIL’s

Jamnagar refinery complex has the world’s largest refining capacity (1.3 million bpsd) at a single

location and has an impeccable track record of safe and reliable operations. RIL is continuing its

efforts to be a creator of intellectual property and is driving a “technology transformation” agenda

that promotes scientific and technical excellence by fostering an eco-system for creativity and

innovation.

RIL Overview

3

“Any statement, opinion, prediction, comment, or observation made in this presentation/publication are

those of the presenter/author only and in no condition should be construed necessarily representing the

policy or intent of Reliance Industries Ltd. (RIL).

The information presented herein is of the presenter/author’s own and in no way RIL shall attract any

liability for any inconsistency or irregularity in terms of accuracy completeness, veracity, or truth of the

content of the presentation/publication. In addition, RIL shall not be liable for any copyright infringement

and misrepresentation for the presented content as the content, in good faith, is presumed to be a

creation of presenter’s/author’s own mind.

The scope of this presentation/publication is strictly for knowledge sharing purposes and not

necessarily to provide any advice or recommendation to the audience/readers. Any endorsement,

recommendation, suggestion, or advice made by the presenter/author shall be in his personal capacity

and not in professional capacity as an employee of RIL. Any person acting on such endorsement,

recommendation, suggestion, or advice will himself/herself be responsible for any injury/damages.”

Disclaimer


Energy Crises and Alternative Fuels

Dismal scenario of fossil fuel availability

Perils of petroleum resource exhaustion

Stringent environmental legislation


Alternative Fuels

Properties Methanol Ethanol Butanol Gasoline

Molecular

formula

CH3OH C2H5OH C4H9OH Many

Energy content

(per gallon)

63k Btu 78k Btu 110 k Btu 115k Btu

Motor octane

number

91 92 94 96

Ethanol may have the early mover’s advantage, but the race is not yet over , butanol is an

emerging biofuel


Butanol has higher energy content

Six times less evaporative

Non-corrosive

100% substitutable for gasoline

No engine modification

Butanol is a ‘higher grade’ fuel in many respects, the only challenge is to produce it enough

to meet the world’s insatiable demand

Butanol v/s Other Fuels


Source : Ramey and Yang, DOE Report, 2004

Biological Route to n-Butanol


Limitations of Butanol Fermentation

Low yields of butanol

Product toxicity to the bacterium

Poor recovery

Culturing and handling of anaerobes

Butanol is toxic to the bacterial system at 1.3% concentration


Source: http://www.biobutanol.com/The -Players.html

Global Players


Economic Viability

The economic viability of n-butanol fermentation is governed

mainly by three factors

High product titer

(achievable by overcoming solvent toxicity)

Raw material cost

(using lignocellulosic biomass)

Solvent recovery costs

(if titers can be raised from 12 to 19 gL-1, cost of recovery can

be cut down to half)


Butanol toxicity

Butanol toxicity is the biggest

challenge in butanol fermentation

Butanol tolerance can be key to

high butanol titers


Strain Improvement

Chemical mutagenesis was carried out to generate butanol tolerant

strains

Mutants were screened for solvent tolerance in medium with

varying concentrations of n-butanol


Butanol Tolerance

Mutant strain was tolerant upto 3.0% (v/v) of butanol

Wild Type Strain Mutant Strain


Mechanism of Butanol Tolerance

1. Altering the lipid composition

Myristic (C14:0) and Palmitic acid

(C16:0) were found to be present

in solvent tolerant mutant strain

Oleic acid (C18:1) conc. is high in

the solvent non-tolerant wild type

strain

Standard FAMEs

Wild type strain

Mutant strain


2. Over expression of the heat shock proteins, HSP 60


Solvent tolerant mutant strain showed higher levels of GroEL

Western Blot showing GroEL expression in mutant (A) and wild-type (B)

Lanes 2–5 protein

Lane 1, standard GroEL

Lane 2, 120 h

Lane 3, 96 h

Lane 4, 72 h

Lane 5, 48 h

kDa

A B



Rhodamine G accumulation in

wild-type & mutant

Rhodamine 6G

P-glycoprotein binds a large number of

lipophilic compounds, such as antibiotics,

dyes, organic solvents and mediates

the energy dependent efflux of these

toxic compounds from the bacterial cells

Rhodamine 6G (dye) is a P- glycoprotein

substrate

P-glycoprotein is an efflux pump (ABC transporter) of multidrug resistance family


Lignicellulosic Biomass For Biofuels

Corn stover Switch Grass

Jatropha Seeds

Another challenge is the cost of raw material which can be mitigated

by using ligno-cellulosic biomass as the feedstock

18

© Reliance Industries Ltd., 2013

Company Microbe Molecule Biomass

Butamax

(DuPont/BP)

Clostridum sp.,

E.coli

Iso-butanol,

n- butanol

Corn

Green Biologics Clostridium sp. n - butanol Lignocelluloses

Butalco Yeast Unclear Lignocelluloses

Gevo Yeast , E.coli Iso-butanol Corn

Cobalt Biofuels Clostridium sp. n- butanol Wood pulp,

sugarbeet

Tetra Vitae Clostridium sp. n- butanol variable

Lignocellulosic Biomass used for Biofuels

Source: http://www.biobutanol.com/The -Players.html


Why Jatropha?

Hardy nature

Short gestation time of about 3 years

Productive life of 50 - 100 years

Not browsed by animals

Adaptability to varied agro-climatic

conditions and soil type

Drought resistance

Non competing with food crops for land and

water sources

Jatropha Is Better For Bio-fuel Production


Optimization of Jatropha Seed Cake (JSC)

Concentration

Acid pretreated JSC hydrolysates (2%, 5%, 7% &10% w/v )

examined for butanol production

Mutant grown in JSC (7 %w/v) with 2 % glucose yielded 13.2 g L−1

butanol


14.8 g L−1 of n-butanol in 96 h

Acetone : Butanol : Ethanol

2.6: 6.6: 0.8

Scale-up of Butanol Production using JSC (3L)


Scale-up of Butanol Production using JSC (15L)

A butanol titer of 18.6 g L-1 obtained in 72 h with a productivity of 0.26 g L-1 h-1


Butanol Titers Using Biomass

Biomass Hydrolysate Additives in

medium

Butanol titer

(gL-1)

Productivity

(gL-1h-1)

Yield

(g/g)

Wheat straw

(Pfromm et al. 2010)

Glucose, YE ,

Vitamin Sol

12.7 in 72h 0.176 0.26

Barley straw

(Qureshi et al. 2010a)

Glucose, YE ,

Vitamin Sol

18.01 in 68h 0.264 0.43

Corn Strover

(Qureshi et al. 2010b)

Glucose 10.4 in 96h 0.108 0.43

Switch grass

(Qureshi et al. 2010b)

Glucose 9.55 in 84h 0.114 0.37

Rice Straw

(Ranjan et al. 2013)

None 13.5 in 12 days 0.047 0.34

Corn Fiber

(Qureshi et al. 2008)

Glucose, YE ,

Vitamin Sol

6.4 in 88 h 0.072 0.27

Jatropha Seed cake

(This study)

Glucose, YE ,

Peptone

18.6 in 72 h 0.255 0.45


Solvent-tolerant mutant strain was developed

Increasing n-butanol productivity using cheap substrate JSC, a

potential lignocellulosic substrate

The process could be successfully scaled up at 15 L

Entire fermentation process was run in a single batch mode

Conclusion


Production of high yield of butanol (18.6 gL-1) in a single batch

process using jatropha seed cake as the biomass

Highly solvent tolerant mutant

No stripping of butanol produced during fermentation

Highlights

26

Forward Path

Optimization Strategies

Use of Flexible feedstocks

Efficient pre treatment method

Scale up of the process up to pilot size

Developing down stream processing of

butanol

Recovery of byproducts (CO2, H2)



Acknowledgements

Dr. Vidhya Rangaswamy

Harshvardhan Joshi

Dr. Sanjeev Katti

Reliance Life Sciences

Reliance Industries Ltd.

28

Thank You!


Documents

Production of Biobutanol from Jatropha Seed Cake...23© Reliance Industries Ltd., 2013 Butanol Titers Using Biomass Biomass Hydrolysate Additives in medium Butanol titer (gL-1) Productivity