| With the depletion of petroleum resources and increasing concerns about environmental pollution, green and renewable biofuels as alterative of fossil energy sources have attracted extensive interest. Lignocellulosic biomass, which mainly consists of cellulose, hemicellulose and lignin is a promising renewable feedstock for production of biofuels, platform chemicals and advanced material due to its high availability, and low cost. However, it is highly recalcitrant to chemical and biological degradation due to its natural complicated structure. Therefore, pretreatment and fractionation of lignocellulosic biomass are generally necessary prior to its subsequent transformation and comprehensive utilization. Recently, ionic liquids(ILs) have emerged as a promising class of solvents for biomass pretreatment owing to their excellent physiochemical properties, especially the strong ability to dissolve cellulose and lignin. A series of easily biodegradable, renewable cholinium ionic liquids with low toxicity have been reported by our group. They can selectively extract lignin from lignocellulosic biomass, and they are a type of efficient solvents for pretreatment. However, the mechanisms of cholinium IL-based pretreatment and enzymatic hydrolysis of lignocellulosic biomass have not been reported so far. Thus, cholinium amino acids([Ch][AA]) and cholinium carboxylic acids([Ch][CA]) were tested as the pretreatment solvents based on the structure-function relationship. With Kraft lignin as the model lignin, changes in the structure and thermal properties were explored during IL pretreatment. Then, fractionation of rice straw and IL reuse were studied for the first time. In addition, the structure and properties of lignins obtained by different pretreatment methods(ball grinding, alkali, [Ch][AcO] and [Ch][Arg] pretreatment) were compared. The mechanism of cholinium IL-mediated selective lignin extraction was illustrated on the molecular level. Finally, the residue lignins(RLs) were isolated from the pretreated rice straw and then characterized. The effects of RLs on the cellulose and cellobiose hydrolysis, non-productive enzyme adsorption were examined. The mechanistic insight into the influence of RLs on enzyme hydrolysis of the pretreated rice straw was gained, based on the structures of RLs. A green, economic and efficient process of lignocellulose biomass pretreatment and fractionation was developed.It was shown that the skeleton structures of Kraft lignin were not damaged upon cholinium IL pretreatment, while the depolymerization and condensation reactions happened in side chain region. Both molecular weight and polydispersity index(PDI) were reduced, and the thermal properties changed. Infrared spectra(FTIR), 1H NMR spectra and two-dimension heteronuclear single quantum C-H correlation spectra(2D HSQC NMR) analysis indicated that Kraft lignin studied was isolated from softwood. FTIR and thermal analysis showed that aromatic skeleton structure of Kraft lignin was not damaged, except for the breakage of partial ester bonds. NMR analysis, gel permeation chromatography(GPC) and elemental analysis(EA) demonstrated that the depolymerization and condensation reactions occurred simultaneously during pretreatment; however, the depolymerization reaction was dominated. The β-O-4′ linkage was broken, while the β-β′ and β-5′ linkages were formed. In addition, the dehydration and demethoxylation reactions occurred. The molecular weight and PDI of the pretreated Kraft lignin decreased, and the degree of unsaturation increased significantly from 224 to 324-349. Thermal analysis(TGA and DSC) showed that pretreatment resulted in a higher maximal decomposition temperature and a higher glass transition temperature of Kraft lignin.It was feasibility to fractionate rice straw into carbohydrate-rich material(CRM) and lignin-rich material(LRM) through cholinium IL pretreatment, and these ILs showed excellent reusability. The order of the lignin extractability and the LRM recovery decreased as follows: [Ch][Arg] > [Ch][Lys] ≈ [Ch][Gly]> [Ch][AcO] > [Ch][Glc]. All ILs could be recovered with the yields of >91%. [Ch][Arg] was an effective pretreatment solvent for grass lignocelluloses such as wheat straw, sugarcane bagasse and corncob as well as eucalyptus. This IL could effectively remove lignin, thus resulting in significant improvements in the glucose yields(58-75%) in subsequent enzymatic hydrolysis. However, [Ch][Arg] pretreatment was inefficient to make pine susceptible to biodegradation. [Ch][Arg] showed excellent recyclability, and the total recovery was as high as 75% after reuse for 8 cycles. Besides, rice straw pretreated by the recycled IL remained highly digestible, and good glucose yields(63-75%) and xylose yields(25%) were achieved in the enzymatic hydrolysis.It was found that some similar reactions happened during cholinium IL- and alkali-mediated lignin extraction and some different reactions occurred also. FTIR, 1H NMR and 2D NMR analysis indicated that G, S, and H units were present in various isolated lignins, and the aromatic skeleton was not damaged during pretreatment. 2D NMR spectra showed that there were typical β-O-4′ signals in the side chain regions; the relative content of the β-O-4′ linkage decreased significantly from 78%(milled wood lignin, MWL) to 27.6%(alkali lignin, AL), 31.8%(cholinium acetate, AcOL) and 32.1%(cholinium argininate, ArgL). There were strong tricin signals in the aromatic regions. The S/G ratios of lignin were 0.50(MWL), 1.01(AL), 0.91(Ac OL) and 1.06(ArgL). The reults indicated that the β-O-4′ linkage was broken during alkali and cholinium IL pretreatment, and that both solvent systems preferred the S type lignin extraction. 1H NMR analysis showed that there was the β-βˊ linkage in AL, but the linkage was not detected in AcOL and ArgL, indicating occurrence of the condensation reactions during alkli pretreatment, possibly due to the stronger pretreatment severity. GPC analysis indicated that the molecular weight of AL was much higher than others, due to the formation of the condensation structure. EA showed that the degree of unsaturation of MWL(272) was the lowest, while the degree of unsaturation of AL(323) was the highest.It was found that the polysaccharide digestibility of CRM pretreated by various cholinium ILs was different significantly. In addition to the lignin content, specific surface area and pore volume, the composition and structures of the residual lignins(RLs) might exert effect on the enzymatic hydrolysis. Both sources and concentrations of RLs had the inhibitory effect on cellulose hydrolysis, while having no effect on cellobiose hydrolysis. The reason might be that the degradation of cellulose into cellobiose was a rate-limiting step, while conversion of cellobiose into glucose was fast. Cellulase adsorption on the RL from the same source was similar to β-glucosidase. The enzyme adsorption capacity of the RLs decreased in the following order: Glc-RL>Ac O-RL>Gly-RL. The loss of cellulase activity due to non-productive cellulase adsorption on RLs resulted in the reduced enzymatic hydrolysis efficiency. The higher the conent of the carboxyl group was, the weaker non-productive enzyme adsorption ability of RL was. However, the presence of the phenolic hydroxyl group would result in the inhibitory effect on the enzyme activity.The study not only preliminarily elucidates the mechanism of cholinium IL-based pretreatment and enzymatic hydrolysis of lignocellulosic biomass, but also establishs a green, economic and efficient process for pretreatment and fractionation of lignocellulose biomass. It lay a theoretical foundation for the value-added utilization of all fractions of biomass and it will promote the transformation of the fossil economy to the carbohydrate economy. |
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