Aerobic Biorefining Study Of Biofuels And Chemicals

Lignocellulosic biomass is widespread and cheap renewable resources and could be converted into kinds of bio-based chemicals by pretreatment,detoxification,hydrolysis,fermentation,separation and purification.Current biorefinery research mainly focuses on anaerobic production of few bulk chemicals(ethanol or lactic acid).However,aerobic fermentation is essential for producing microbial lipid as feedstock of biodiesel and aircraft fuel and chemicals(sugar acids or citric acid).High solids loading is inevitable choice for achieving high product titer and low process cost in lignocellulose biorefinery.However,high solid content aerobic fermentation still exists many key issues.High solids loading can significantly decrease mass transfer rate,hence leading to oxygen transfer limit and mixing difficulty.Oxygen transfer limit and high inhibitors stress at high solids loading can impact strain fermentability.Tons of solid particles can cause operation difficulty of separation and purification and low product recovery yield.These issues hinder efficient implementation of aerobic fermentation,hence restricting application category of lignocellulose biorefinery for producing more chemicals.This study plans to break technical obstacles of aerobic biorefinery to achieve efficient production of various chemicals and biofuels by aerobic fermentation.The study firstly analyzes oxygen transfer characteristic of lignocellulose system at high solids loading for high product titer and proposes a solution for oxygen supply sufficiently in high solid content and high viscous lignocellulosic slurry,finally obtaining high titer of cellulosic sugar acids,citric acid and microbial lipid by technology optimization,process control and strain screening,which provides more application schemes for industrial biorefining technology.The first part of the thesis focused on investigation of oxygen transfer characteristic of lignocellulosic hydrolysate slurry at high solids loading to propose a solution for aerobic fermentation in high viscous lignocellulosic system.Oxygen transfer performance was evaluated in lignocellulosic hydrolysate at different solids loadings and a decrease tendence in oxygen transfer rate was uncovered with increasing solids loading.However,oxygen transfer rate could be significantly improved by adjusting operation conditions such as hydrolysate time and agitation rate.Gluconic acid production by Gluconobacter oxydans is ultimately hampered in oxygen transfer rate,but it still could be efficiently achieved in high solid content lignocellulosic slurry(30%,w/w)at moderate operation conditions.Computational fluid dynamics model was established based on rheology of lignocellulose slurry for optimally designing aerobic bioreactor and then achieved mature gluconic acid fermentation at low mixing energy consumption.The second part assessed aerobic fermentation for sugar acids production.G.oxydans could utilize all lignocellulose-derived sugars including glucose,xylose,arabinose,mannose and galactose,but cooperative utilization of these sugars only be enforced by whole cell catalysis.A long term adaptive evolution of G.oxydans was carried out in this part for intensifying utilization efficiency of non-glucose sugars and tolerance for high inhibitors stress and achieved the conversion of all sugars to the corresponding sugar acids by aerobic fermentation.Evolved strain obtained about 180 g/L of sugar acids using corn stover at extremely high solids loading of 35%.Cellulosic sugar acids reached to similar yield with starch sugar acids production and could compete partly with starch-based product at process cost based on techno-economic analysis.Cellulosic sugar acids presented excellent retarding efficiency as cement additive,showing well application prospect in construction industry.The third part attempted simultaneous saccharification and aerobic fermentation(SSF)for citric acid production.The mycelia of the filamentous Aspergillus niger fungus entangling with the rodlike corn stover fibers strongly intensified the viscosity of fermentation slurry,potentially worsening oxygen transfer performance.However,the SSF for citric acid production still could well be achieved by adjusting fermentation parameters.Finally,SSF produced 136 g/L of cellulosic citric acid,hence improved by 36%comparing with current level,possessing the potential for industrialized application.The fourth part investigated simultaneous saccharification and aerobic co-fermentation(SSCF)for microbial lipid production.Based on lower density of higher lipid content cell,this study proposed a new strain screening strategy for obtain high-yield oleaginous yeast Trichosporon cutaneum,ultra-centrifugal screening.This strategy is carried out by centrifugation for obtaining the lighter cells after culture as the seed for the next round culture.This study used this method improve by folds lipid content in cells,also induce cell morphology evolution.Screened strains(T.cutaneum MS28 and WL97)had more precursor Acetyl-CoA and coenzyme NADPH in lipid synthesis pathway than parental strain(T.cutaneum ACCC 20271)and better mutliple sugars utilization.These results demonstrated excellent lipid accumulation of screened strains in different aspects.Screened strains had larger cell volume and thinner cell wall than parental strain,and lower glucan content and mannan content in cell wall composition.These changes on cell morphology provided more space for lipid accumulation.A hydrostatic model was established to represent ultra-centrifugal screening mechanism and demonstrated screening efficiency and universality.This strategy could be suitable for screening light oleaginous cells,also heavy cells producing polyhydroxyalkanoate.Screened strain T.cutaneum WL97 could utilize all lignocellulose-derived sugars by SSCF to produce 41.6 g/L of lipid at solids loading of 30%,while parental strain only could partly consume glucose to generate 7.5 g/L of lipid.Finally,this study used screened strain by SSCF coupled with semi-continuous fermentation produce 46.7 g/L of lipid,which improved 2.9 folds than literature report,hence reaching to the leading level in the world.Almost 5%lipid content in fermentation slurry had exceeded the minimum threshold oil content(3%,w/w)of commercially recoverable oil field,with potential industrial implications.Special separation of oleaginous cells from fermented lignocellulose residual was achieved utilizing the hydrolysis of cellulase into fermentation slurry.Overall,this study solved key issues of aerobic fermentation in high solids loading and high viscous lignocellulose slurry,and implemented high performance production of biofuels and chemicals.A solution was proposed for sufficiently oxygen supply of aerobic fermentation in high solid content and high viscous lignocellulose slurry by analyzing oxygen transfer characteristic.Cellulosic sugar acids production by aerobic fermentation was very close to starch-based one at extremely high solids loading.High titer of citric acid production by SSF was firstly achieved by A.niger,and improved 36%of citric acid yield than previous reports.World-class cellulosic lipid was implemented using excellent strain obtained by an ultra-centrifugal screening.These high target biochemical productions substantially expanded industrialization path of biorefining technology,and strongly established basis for commercial application of renewable resources.