Study On The Mechanism Of Organosolv Lignins On Enzymatic Hydrolysis Of Lignocellulosic Biomass

Bioenergy presents opportunities for improvement of climate and environmental,security of energy supply and economic development,and has in consequence been supported by the policy in many countries.Biorefining of lignocellulose to ethanol is one prospective form of bioenergy.However,the commercialization of bioethanol is restricted by the low efficiency and the high cost of cellulose saccharification technology.Furthermore,lignin is one of the important factors that hinder the cellulase hydrolysis,which can reduce the enzymatic hydrolysis yield mainly through the physical barrier action and its nonspecific adsorption with cellulase enzymes.Recently,related studies have shown that some modified lignin,such as lignosulfonates,?-carboxylated,and hydroxypropylated lignin,can promote enzymatic hydrolysis of cellulose or lignocelullose.The electrostatic interactions,hydrophobic interactions and hydrogen bonding interactions between lignin and cellulase play important roles in enzyme hydrolysis of lignocellulose.How does lignin inhibit or promote cellulase hydrolysis?It remains a problem for researchers in bioenergy field.The solution of this problem will help to remove the block effect of lignin in biorefining and promote the industrial development of bioethanol.This paper extracted different lignins from cottonwood,black willow,aspen,eucalyptus and loblolly pine with sulfuric acid and ethanol solvent.the influence of these ethanol organosolv lignins(EOL)on enzyme hydrolysis of cellulose was studied.The results showed that EOL samples from the cottonwood,black willow,aspen increased enzyme hydrolysis yield of microcrystalline cellulose by 5%-13%,while EOL samples from eucalyptus and loblolly pine reduced enzyme hydrolysis yield by 4%and 13%respectively.However,those EOL preparations had weak effect on enzyme hydrolysis rate(1.11 g/L/h).To further explore the mechanism of promotion or inhibition of EOL samples on cellulase hydrolysis,the physicochemical properties and structure characters of the lignins were examined.We found that adsorption bonding strength between lignin and cellulose enzyme,zeta potential,mean particle size of the lignins had good linear relationship with 72 h enzyme hydrolysis yield(R~2>0.90).Meanwhile,it showed a strong correlation between 72 h enzymatic hydrolysis yield and zeta potential and hydrophobicity.Hydrophobicity of lignin performed for a negative relationship and zeta potential showed a positive relationship with 72 h enzyme hydrolysis yield.On the other hand,PB structure might be associated with promoting effect of lignin on enzymatic hydrolysis.It needed to be proved by further research.Hydrochloric acid can be used as an alternative catalyst during the organosolv pretreatment to improve the lignin removal yield and the high value utilization of lignin.This article pretreated cottonwood,black willow,aspen,eucalyptus and loblolly pine with hydrochloric acid as the catalyst(HCl-EOL)and ethanol as solvent.The physicochemical properties and structure characteristics of HCl-EOL lignins were compared with EOL lignins.It showed that lignin removal yield(the yield of lignin)was obviously higher for HCl-EOL lignins.HCl-EOL samples from the cottonwood,black willow,and aspen increased the 72h enzyme hydrolysis yield of microcrystalline cellulose by 7%-10%.HCl-EOL lignins from eucalyptus and loblolly pine inhibited the enzymatic hydrolysis yield by 10%and 4%respectively.Compared to EOL preparations,the structures of HCl-EOL samples are similar,the combination strength between HCl-EOL preparations and enzyme decreased,surface negative charge deducted,and hydrophobicity increased.Similarly,it revealed that the effect of lignin on enzymatic hydrolysis is a function of both hydrophobic interactions and electrostatic repulsions.The lignin inhibition is controlled by lignin hydrophobicity and the lignin stimulation is governed by the negative zeta potential.The net effect of lignin depends on the combined influence of hydrophobicity and zeta potential.To further verify the hydrophobic effect and the influence of the electrostatic effect on the cellulase hydrolysis efficiency,this paper modified the lignins of aspen,eucalyptus and loblolly pine with 2-naphthol(2N-EOL),which is a carbenium ion clearing agent.The objective of this operation was to decrease the hydrophobicity,increase the surface negative charge of the lignins and alleviate or eliminate the inhibitory effect of them on cellulose enzyme hydrolysis.We found that the negative effect of EOL-EH and EOL-LP on enzymatic hydrolysis was converted to the positive effect after being modified by 2-naphthol.2N-EOL from eucalyptus and loblolly pine both increased enzymatic hydrolysis yield by 7%.Given 2D-HSQC spectra,the structure of 2N-EOL was changed during the organosolv pretreatment process.?-OH in?-O-4 structure was substituted with ethoxy,and C?in S unit was oxidized.In addition,?-position of 2N-EOL was replaced with 2-naphthol,which is the main reason to turn inhibition of EOL on the cellulose enzyme hydrolysis into promoting effect.To study the effect of EOL preparations on biorefinary process,we also studied the influence of the lignins on ethanol yield.Cellulose PH101 was incubated together with cellulase enzymes and saccharomyces cerevisiae(ATCC 26602)under the appropriate condition and fermented for 96 h.The variation trends of ethanol yield of simultaneous saccharification fermentation were quantificationally researched based on factors such as the dosage of enzyme,the type of lignin,lignin concentration and different substrates.The results showed that the ethanol yield increased with the increase of enzyme dosage,while the growth rate of ethanol yield slow down gradually when the dosage of cellulase higher than 10FPU/g.EOL preparations from the cottonwood,black willow,and aspen improved ethanol yield of SSF,but the EOL samples of eucalyptus and loblolly pine had inhibitory effect on SSF process.The higher concentration of lignin from cottonwood,black willow and aspen,the larger promoting effect on SSF process were.As EOL concentration from eucalyptus and loblolly pine increasing,the the inhibition of EOL samples on the SSF process were greater.Compared with the pure cellulose,as the substrate,the promotion/inhibition of EOL samples on SSF process pretreated aspen and loblolly pine was relatively weaker.