Study On The “Structure-activity Relationship” Of The Interaction Between Lignin And Cellulase During The Process Of Hot Water And Green Liquor Pretreatment

Lignocellulose is a kind of abundant and renewable energy resources.Enzymatic hydrolysis is key step for lignocellulose converted into fermentable sugar and further transform sugar to biofuels.The dense structure and natural impedance of lignocellulose hinder the conversion efficiency of lignocellulose to biofuels.Pretreatment can effectively destroy cell wall structure of lignocellulose and improve the efficiency of enzymatic hydrolysis.However,lignin as main component of lignocellulose inhibited the enzymatic hydrolysis efficiency of lignocellulose.At present,the mechanism of the effect of lignin structure on the interaction between lignin and cellulase is still lacking.Therefore,exploring the interaction between lignin and cellulase will provide theoretical basis and application for improving the enzymatic hydrolysis and saccharification efficiency of lignocellulose.The purpose of this study was isolated lignin from hot water and green liquor pretreated sugarcane bagasse,eucalyptus and spruce,and to study the structure-activity relationship of the interaction between lignin and cellulase.The main results were presented as follows.(1)Green liquor pretreatment mainly removed lignin from lignocellulose,hot water pretreatment mainly removed hemicellulose from lignocellulose.After green liquor and hot water pretreatment,the surface pores of fibers increased,the surface of fibers became wrinkled and rough,exposed more internal microfibers,and the crystallinity and specific surface area of lignocellulose increased.After hot water pretreatment,the total sugar conversion of sugarcane bagasse,eucalyptus and spruce were 60.1%,21.8% and 20.4%,respectively.The total sugar conversion of sugarcane bagasse,eucalyptus and spruce were 77.3%,32.1% and 22.9%,respectively.Pearson correlation and multiple linear regression analysis indicated that the inhibition of lignin on enzymatic efficiency was greater than hemicellulose.(2)During the hot water pretreatment,the content of lignin basic structural monomer,methoxyl and aliphatic hydroxyl in MWL of sugarcane bagasse,eucalyptus and spruce decreased,the content of ?-?,?-5,phenolic hydroxyl and total hydroxyl increased,the content of molecular weight,carbon element and hydrogen element increased,and the content of oxygen element decreased.During green liquor pretreatment process,the content of aliphatic hydroxyl group,methoxyl group,phenolic hydroxyl group and total hydroxyl group in lignins were lower than that lignins during hot water pretreatment,the ?-?,?-5 and molecular weight in MWL increased,the content of carbon and hydrogen element increased,and the content of oxygen element was lower than hot water pretreated MWL.(3)QCM-D analysis indicated that the adsorption capacity of hot water and green liquor pretreatment lignins to cellulase was higher than that protolignins.Temperature and enzyme concentration increased the adsorption rate and amount of cellulase on lignins.The dynamic model of cellulase adsorption on lignin was built according to QCM-D analysis.A positive correlation was observed between ?-?,?-5,phenolic hydroxyl group and enzymes adsorption.SDS-PAGE analysis indicated that the order of cellulase components adsorbed by lignins was ?-glucanase>xylanase>endoglucanase>cellobilhydrolase.The electrostatic interaction increased the adsorption of ?-glucanase and xylanase on lignins,while the synergy of the hydrophobic interaction and hydrogen bond increased the adsorption of endoglucanase and cellobilhydrolase on lignins.Furthermore,lignin inhibited the enzymatic hydrolysis of cellulose,a negative correlation was observed between ?-?,?-5,phenolic hydroxyl group content and enzymatic hydrolysis of cellulose.(4)The amount of enzymes adsorption on G-type and S-type lignin monomers was higher than H-type lignin monomer,and the amount of enzymes adsorption on G-type lignin monomer was higher than S-type lignin monomer.The amount of enzymes adsorption on 5-5? dimer lignin model with aldehyde side chain was higher than that 5-5? dimer lignin model with propyl and hydroxyl side chain.The amount of enzymes adsorption on 5-5? dimer lignin model,pinoresinol and dehydro diconiferyl alcohol was higher than G-type lignin monomer,indicated that the 5-5?,?-? and ?-5 structure increased the adsorption of enzymes on lignin,which may be due to the condensed structure increase the hydrophobic interaction between lignin and enzymes.(5)AFM analysis showed that the interaction force between cellulase and different nanofibers was in the order of hot water pretreated-nanofibers > green liquor pretreated-nanofibers > raw materials nanofibers > bleached nanofibers.The lignin content on the surface of nanofibers increased the interaction force between cellulase and nanofibers.The interaction force between cellulase and lignins was at least 62% higher than that interaction force between cellulase and nanofibers.The interaction force between hot water pretreated lignins and cellulase was greater than protolignins and green liquor pretreated lignins.The condensation structure of lignin may increase the interaction between cellulase and lignin,which may be due to increase hydrophobic interaction force between lignin and cellulase.Conclusively,the hot water and green liquor pretreatment were used to explore the relationship between physicochemical properties of lignocellulose and the enzymatic hydrolysis of lignocellulose,clarified the inhibition mechanism of lignin on enzymatic hydrolysis of lignocellulose,revealed the “structure-activity relationship” of the interaction between lignin and cellulase,and provided theoretical and applied research basis for improving the enzymatic hydrolysis of lignocellulose.