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JMD DTA Aromatics Plant Production of Biobutanol from Jatropha Seed Cake Jasmine Isar RIL Overview 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. 2 © Reliance Industries Ltd., 2013 Disclaimer “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.” 3 Energy Crises and Alternative Fuels Dismal scenario of fossil fuel availability Perils of petroleum resource exhaustion Stringent environmental legislation 4 © Reliance Industries Ltd., 2013 Alternative Fuels Properties Methanol Ethanol Butanol Gasoline Molecular CH3OH C2H5OH C4H9OH Many formula Energy content 63k Btu 78k Btu 110 k Btu 115k Btu (per gallon) Motor octane 91 92 94 96 number Ethanol may have the early mover’s advantage, but the race is not yet over , butanol is an emerging biofuel 5 © Reliance Industries Ltd., 2013 Butanol v/s Other Fuels 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 6 © Reliance Industries Ltd., 2013 Biological Route to n-Butanol Source : Ramey and Yang, DOE Report, 2004 7 © Reliance Industries Ltd., 2013 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 8 © Reliance Industries Ltd., 2013 Global Players Source: http://www.biobutanol.com/The -Players.html 9 © Reliance Industries Ltd., 2013 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 -1 (if titers can be raised from 12 to 19 gL , cost of recovery can be cut down to half) 10 © Reliance Industries Ltd., 2013 Butanol toxicity Butanol toxicity is the biggest challenge in butanol fermentation Butanol tolerance can be key to high butanol titers 11 © Reliance Industries Ltd., 2013 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 12 © Reliance Industries Ltd., 2013 Butanol Tolerance Mutant strain was tolerant upto 3.0% (v/v) of butanol Wild Type Strain Mutant Strain 13 © Reliance Industries Ltd., 2013 Mechanism of Butanol Tolerance 1. Altering the lipid composition Wild type strain Myristic (C14:0) and Palmitic acid (C16:0) were found to be present in solvent tolerant mutant strain Mutant strain Oleic acid (C18:1) conc. is high in Standard FAMEs the solvent non-tolerant wild type strain 14 © Reliance Industries Ltd., 2013 Mechanism of Butanol Tolerance 2. Over expression of the heat shock proteins, HSP 60 Solvent tolerant mutant strain showed higher levels of GroEL kDa A B 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 15 © Reliance Industries Ltd., 2013 Mechanism of Butanol Tolerance 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 G accumulation in wild-type & mutant Rhodamine 6G (dye) is a P- glycoprotein substrate Rhodamine 6G P-glycoprotein is an efflux pump (ABC transporter) of multidrug resistance family 16 © Reliance Industries Ltd., 2013 Lignicellulosic Biomass For Biofuels Another challenge is the cost of raw material which can be mitigated by using ligno-cellulosic biomass as the feedstock Corn stover Switch Grass Jatropha Seeds 17 © Reliance Industries Ltd., 2013 Lignocellulosic Biomass used for Biofuels Company Microbe Molecule Biomass Butamax Clostridum sp., Iso-butanol, Corn (DuPont/BP) E.coli n- butanol 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 Source: http://www.biobutanol.com/The -Players.html 18 © Reliance Industries Ltd., 2013 Why Jatropha? Jatropha Is Better For Bio-fuel Production 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 19 © Reliance Industries Ltd., 2013 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 20 © Reliance Industries Ltd., 2013 Scale-up of Butanol Production using JSC (3L) 14.8 g L−1 of n-butanol in 96 h Acetone : Butanol : Ethanol 2.6: 6.6: 0.8 21 © Reliance Industries Ltd., 2013 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 22 © Reliance Industries Ltd., 2013 Butanol Titers Using Biomass Biomass Hydrolysate Additives in Butanol titer Productivity Yield medium (gL-1) (gL-1h-1) (g/g) Wheat straw Glucose, YE , 12.7 in 72h 0.176 0.26 (Pfromm et al. 2010) Vitamin Sol Barley straw Glucose, YE , 18.01 in 68h 0.264 0.43 (Qureshi et al. 2010a) Vitamin Sol Corn Strover Glucose 10.4 in 96h 0.108 0.43 (Qureshi et al. 2010b) Switch grass Glucose 9.55 in 84h 0.114 0.37 (Qureshi et al. 2010b) Rice Straw None 13.5 in 12 days 0.047 0.34 (Ranjan et al. 2013) Corn Fiber Glucose, YE , 6.4 in 88 h 0.072 0.27 (Qureshi et al. 2008) Vitamin Sol Jatropha Seed cake Glucose, YE , 18.6 in 72 h 0.255 0.45 (This study) Peptone © Reliance Industries Ltd., 2013 23 Conclusion 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 24 © Reliance Industries Ltd., 2013 Highlights 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 25 © Reliance Industries Ltd., 2013 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) 26 © Reliance Industries Ltd., 2013 Acknowledgements Dr. Vidhya Rangaswamy Harshvardhan Joshi Dr. Sanjeev Katti Reliance Life Sciences Reliance Industries Ltd. 27 © Reliance Industries Ltd., 2013 Thank You! 28 © Reliance Industries Ltd., 2013 .
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