yeast hardening for cellulosic ethanol production bianca a. brandt supervisor: prof j gorgens...
TRANSCRIPT
![Page 1: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/1.jpg)
Yeast Hardening for
Cellulosic Ethanol
productionBianca A. BrandtSupervisor: Prof J Gorgens
Co-Supervisor: Prof WH Van ZylDepartment of Process
Engineering University of Stellenbosch
Energy Postgraduate Conference 2013
![Page 2: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/2.jpg)
Introduction
• Growing global move towards sustainable green energy production– spurred by dependence on rapidly depleting finite fossil fuels – environmental and socio-economic concerns
• Studies into Alternative Clean, Renewable and Sustainable energy resources: – solar-electric/thermal, hydroelectric, geothermal, tidal, wave, wind and
ocean thermal power systems– furthermore, a great deal of work has gone into the development of
biofuels
![Page 3: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/3.jpg)
Introduction
• Why Biofuels?– vehicular transportation- energy stored easier in form of
combustible hydrocarbons then as electricity or heat– compatible with current distribution systems– supplement and replace fossil fuels
• A range of bio-fuels are currently being investigated
• Bioethanol - benchmark biofuel– production based on a proven low cost technological platform– Brazil and USA - cost effective 1st generation bioethanol– sugar and starch
• 2nd generation bioethanol from lignocelluloses
![Page 4: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/4.jpg)
Cellulosic Bioethanol
• Bioethanol from Lignocellulose– cheap, renewable, easily available, under utilized resource– energy/fuel and suitable molecules which can replace
petroleum products
• Lignocellulose bioethanol production process– degradation of lignocellulose to fermentable sugars– fermentation of sugars to bioethanol
• Optimum ethanol production bottle necked– suboptimal xylose utilization and release of microbial inhibitor
molecules during biomass degradation
Pretreatment FermentationHydrolysis
![Page 5: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/5.jpg)
Overcoming Inhibitor toxicity• Challenge – Release of inhibitor molecules during
lignocellulose degradation– furans, phenolics and weak acids – severely impact yeast fermentation efficiency
• Process Optimization – feedstock, pretreatment, hydrolysis conditions– fermentation strategies
• Detoxification of hydrolysate– physical (evaporation); chemical (over-liming)– biological: microbial and enzymatic approaches
• Shown detoxification costs can constitute 22% of total ethanol production cost (Ding et al., 2009)
– economically limited – inhibitor specific and loss of fermentable sugars
![Page 6: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/6.jpg)
Overcoming Inhibitor toxicity• Sustainable cost effective bioethanol fermentation
require “hardened” inhibitor resistant fermentation strains
• Rational engineering approach– Genetic modification – yeast oxido-reductase detoxification
genes– boost innate detoxification mechanisms of yeast– furfural, HMF, Formic acid– improved tolerance to specific inhibitor
• Evolutionary engineering techniques– mutation and long term continuous cultures– simulate natural selection under selective pressure
![Page 7: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/7.jpg)
Hardening yeast
• Despite on-going yeast hardening strategies
• Inhibitor resistant fermentation strains remain elusive and highly sought after!!
• Project aim : Generate “hardened” inhibitor resistant yeast strains
• Approach which combine Novel rational metabolic engineering and evolutionary engineering
![Page 8: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/8.jpg)
Hardening yeast
• Strain generation - Rational metabolic engineering– industrial xylose utilization base strains
• Identify and select yeast detoxification genes from literature– combine specific detoxification genes with cell membrane
stress response genes
• Express inhibitor resistance genes in Saccharomyces cerevisiae– novel gene combinations– elucidate synergistic /antagonistic combinations
![Page 9: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/9.jpg)
Hardening yeast
• Evolutionary engineering– long term continuous cultures - bioreactor– selective pressure – increasing concentrations of inhibitors– further enhance inhibitor resistance– evaluate fermentation efficiency in toxic hydrolysate
• Novel “HARDENED” inhibitor resistant strains
• Optimization of lignocellulosic bioethanol production
![Page 10: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/10.jpg)
Acknowledgements
Supervisors: Prof J Gorgens and Prof WH Van Zyl
Department of process engineering
NRF - Financial Support
![Page 11: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/11.jpg)
Yeast Hardening for Cellulosic Ethanol
production
Bianca A. BrandtSupervisor: Prof J Gorgens
Co-Supervisor: Prof WH Van ZylDepartment of Process Engineering
University of Stellenbosch
Energy Postgraduate Conference 2013
![Page 12: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/12.jpg)
Introduction• Growing global move towards sustainable green energy
production– Spurred by dependence on rapidly depleting Finite Fossil fuels – Various environmental and socio-economic concerns
• Studies into Alternative Clean, Renewable and Sustainable energy resources:
– solar-electric/thermal, hydroelectric, geothermal, tidal, wave, wind and ocean thermal power systems
– furthermore, a great deal of work has gone into the development of bio-fuels
![Page 13: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/13.jpg)
Introduction• Why Biofuels?
– Vehicular transportation- energy stored easier in form of combustible hydrocarbons then as electricity or heat
– compatible with current distribution systems– Supplement and replace fossil fuels
• A range of bio-fuels are currently being investigate
• Bioethanol - benchmark biofuel– production based on a proven low cost technological platform– Brazil and USA -cost effective 1st generation bioethanol– Sugar and starch
• 2nd generation bioethanol from lignocelluloses
![Page 14: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/14.jpg)
Cellulosic Bioethanal• Bioethanol from Lignocellulose
– cheap, renewable, easily available, under utilized resource– energy/fuel and suitable molecules which can replace petroleum
products
• Lignocellulose bioethanol production process– degradation of lignocellulose to fermentable sugars– fermentation of sugars to bioethanol
• Optimum ethanol production bottle necked– suboptimal xylose utilization and release of microbial inhibitor
molecules during biomass degradation
Pretreatment FermentationHydrolysis
![Page 15: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/15.jpg)
Overcoming inhibitor toxicity• Challenge – Release of inhibitor molecules during
lignocellulose degradation– furans, phenolics and weak acids – severely impact yeast fermentation efficiency
• Process Optimization – feedstock, pretreatment, hydrolysis conditions– fermentation strategies
• Detoxification of hydrolysate– physical (evaporation); chemical (over-liming)– biological: microbial and enzymatic approaches
• Shown detoxification costs can constitute 22% of total ethanol production cost (Ding et al., 2009)
– economically limited – inhibitor specific and loss of fermentable sugars
![Page 16: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/16.jpg)
Overcoming inhibitor toxicity• Sustainable cost effective bioethanol fermentation
require “hardened” inhibitor resistant fermentation strains
• Rational engineering approach– Genetic modification – yeast oxido-reductase detoxification genes– boost innate detoxification mechanisms of yeast– furfural, HMF, Formic acid– improved tolerance to specific inhibitor
• Evolutionary engineering techniques– mutation and long term continuous cultures– simulate natural selection under selective pressure
![Page 17: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/17.jpg)
Hardening yeast• Despite on-going yeast hardening strategies
• Inhibitor resistant fermentation strains remain elusive and highly sought after!!
• Project aim : Generate “hardened” inhibitor resistant yeast strains
• Approach which combine Novel rational metabolic engineering and evolutionary engineering
![Page 18: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/18.jpg)
Hardening yeast• Strain generation - Rational metabolic engineering
– Industrial xylose utilization base strains
• Identify and select yeast detoxification genes from literature
– Combine specific detoxification genes with cell membrane stress response genes
• Express inhibitor resistance genes in Saccharomyces cerevisiae
– novel gene combinations– elucidate synergistic /antagonistic combinations
![Page 19: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/19.jpg)
Hardening yeast• Evolutionary engineering
– long term continuous cultures - bioreactor– selective pressure – increasing concentrations of inhibitors– further enhance inhibitor resistance– evaluate fermentation efficiency in toxic hydrolysate
• Novel “HARDENED” inhibitor resistant strains
• Optimization of lignocellulosic bioethanol production
![Page 20: Yeast Hardening for Cellulosic Ethanol production Bianca A. Brandt Supervisor: Prof J Gorgens Co-Supervisor: Prof WH Van Zyl Department of Process Engineering](https://reader035.vdocuments.net/reader035/viewer/2022062716/56649e055503460f94af15f7/html5/thumbnails/20.jpg)
AcknowledgementsSupervisors: Prof J Gorgens and Prof WH Van Zyl
Department of process engineering
NRF - Financial Support