Effects Of Biological Modification And Surfactant Composite Strategy On Biomass Conversion Efficiency Of Corn Stover

With the increasing severity of the energy crisis,the development of material energy has become a necessary measure to alleviate the energy crisis.Due to the complex structure of cellulose encapsulated by the cross-linking of hemicellulose and lignin,the enzymatic hydrolysis recalcitrant of lignocellulose and the high enzyme cost have become the two main bottlenecks of the efficient biotransformation of lignocellulose.The improvement of biomass energy utilization through pretreatment to reduce the enzymatic hydrolysis recalcitrant of lignocellulose has attracted long-term attention,especially biological pretreatment with environment friendly and economic characteristics.Therefore,this subject conduct systematic research from two aspects:reducing cellulase enzymatic hydrolysis recalcitrant and improving cellulase utilization.Firstly,based on the co-expression of cellulase and xylanase in saccharomyces cerevisiae,the synergistic effect of xylanase and cellulase in the bioconversion of lignocellulose was studied.After that,the biotransformation efficiency of laccase modified corn stover(Lac-CS)was evaluated,and the key factors affecting the enzymatic hydrolysis efficiency of corn stover were revealed by the observation of biological modification on the surface microstructure of corn stover.The effect of surfactant on biotransformation was evaluated and its effect on non-productive adsorption of cellulase was investigated.Finally,the biotransformation effect of corn stover under the compound strategy of biological modification and surfactant addition was evaluated.It provides a new idea for biomass ethanol conversion strategy from the laboratory level,and obtains better results.The main conclusions are as follows:1)Cellulase and xylanase promote the conversion of lignocellulose to bioethanol.Cellulase-xylanase co-expression Saccharomyces cerevisiae(INVSc-CBH-TS/CA)biotransformed ethanol production is higher than the cellulase expression strain,and the synergistic effect of cellulase and xylanase can further improve the ethanol yield.At 120 h,the ethanol yield of INVSc-CBH-CA and INVSc-CBH-TS was 1.7 and 2.1times of INVSc-CBH.The ethanol yield of INVSc-CBH-CA/INVSc-CBH-TS at 120 h was 3.4 and 3.8 times higher than the sum of the ethanol yield of INVSc-CBH and INVSc-CA or INVSc-TS,indicating that the synergistic effect of cellulase and xylanase further affects the biotransformation efficiency.2)Laccase-based biological modification promotes enzymatic saccharification and ethanol conversion of corn stover.The GC-MS results show that lignin model compounds 4-hydroxybenzoic acid and p-coumaric acid or 3-hydroxycinnamic was found in corn stover bio-modified products.The FTIR results show that biological modification leads to changes in the functional groups related to lignin in the substrate,such as methyl,methylene,ester,methoxy,aromatic skeleton,etc.Those results confirm that laccase effect on the lignin group in corn stover.Lac-CS has better cellulase enzymatic hydrolysis effect and bio-transformed ethanol yield than original corn stover.The ethanol yield of INVSc-CBH-TS using Lac-CS as a substrate for simultaneous saccharification and fermentation(SSF)was 3.1 times than the control in4 th days.The ethanol yield of INVSc-CBH-CA with Lac-CS as the substrate in 6 days was 1.7 times than the control group.Through the study of the microstructure,it is found that the porosity,crystallinity and microstructure of Lac-CS are the key factors affect the effect of enzymatic hydrolysis.After enzymatic hydrolysis,the protein content in the supernatant of Lac-CS increased by 0.208 mg/m L compared to the control,but the glucose yield was 3.49 times than the control.This result showed that nonproductive adsorption of cellulase in Lac-CS decreased during enzymatic hydrolysis.3)Surfactants addition promote the enzymatic saccharification and ethanol conversion of corn stover.The addition of chemical surfactants(Tween-80)and biosurfactants(rhamnolipids,tea saponin)can effectively increase the ethanol yield and the enzymatic hydrolysis effect of corn stover,and the biological surfactant rhamnolipid has the best effect.In the lignocellulosic SSF process of INVSc-CBH-TS,the ethanol yield with 0.1% rhamnolipid added(0.5 g/L)was 1.61,1.08,2.3 times than the 0.5%Tween-80,0.01% tea saponin,and the control.During the enzymatic hydrolysis of cellulose,the protein content in the supernatant from high to low was 0.01% tea saponin(0.89 mg/m L),0.1% rhamnolipid(0.84 mg/m L),and 0.5% Tween-80(0.78mg/m L).After 0.01% tea saponin,0.1% rhamnolipid,and 0.5% Tween-80 added,the glucose yield of the enzymatic hydrolysis was 0.23 mg/m L,0.39 mg/m L,and 0.12mg/m L,respectively.It shows that rhamnolipids can increase the utilization rate of cellulase by reducing the non-productive adsorption of cellulase,and achieve the best enzymolysis effect.4)The combined effect of biological modification and surfactants can further increase the ethanol yield of corn stover bioconversion.The combined effect of laccase modification and biosurfactant rhamnolipid has the most obvious increase in biomass utilization.Using the combined strategy of biological modification and rhamnolipid,the SSF ethanol yield of INVSc-CBH-TS(0.69 g/L)on the 5th day increased 38%.compared to the control(0.5 g/L),and 30% higher than that of the control(0.56 g/L)in6th day.Comparing the ethanol production of the Tween80+biomodification compound strategy(0.58 g/L)and the tea saponin compound strategy(0.65 g/L)on the 6th day also increased significantly.In summary,this thesis uses cellulase-xylanase co-expression recombinant strains and laccase biological modification to remove the physical barrier of lignocellulose,and uses surfactants to reduce the non-productive adsorption of cellulase.This study systematically studied how to reduce the non-productive adsorption and realize the high efficiency conversion of biomass by removing the physical barrier of hemicellulose and lignin.The key factors affecting biomass conversion(crystallinity,porosity,surface microstructure)and the influence and principle of surfactant on the non-productive adsorption of cellulase were preliminarily analyzed,and laid a theoretical foundation for the development of efficient utilization of biomass energy.