Study On The Efficacy Of Lignin Separation And Purification Strategies Based On The Synergistic Effect Of Microbial Functions

Lignin is a renewable aromatic compound in nature,with plenty of reactive groups,such as phenolic hydroxyl,alcohol hydroxyl,methoxy and so on,which can be used wildly in industrial and agricultural production through modification to obtain high performance on adhesion,surface activity,and complexation.Current methods of lignin separation and purification applied in production generally suffer from high costs,high potential for secondary contamination and weakened properties of lignin materials.Comparatively,biological methods for separation and purification of lignin are potential on low cost,low energy consumption,mild reaction conditions and no secondary contamination,which are limited by efficiency of separation and purification due to the stable chemical bonding structure between lignin and carbohydrates(mainly hemicellulose)called‘lignin-carbohydrate complex’(LCC),so it is not possible to meet the requirements of industrial production.To address two key steps of lignin purification,namely LCC deconstruction and polysaccharides degradation,a new strategy named ‘mixed fungal fermentation combined with anaerobic digestion(FF+AD)’ was proposed in this study,which was based on functional synergistic potential between aerobic fungi and anaerobic microorganisms.In this strategy,a mixed fungal community with efficient destruction of LCC structure,and an anaerobic digestion system with deeply remove of cellulose and hemicellulose were connevted,which was attempt to achieve efficient lignin separation and purification through the functional synergy between microorganisms.The main results were as follows:(1)Lignin-carbohydrate complex(LCC)structure in substrates was loosened through fermentating by mixed fungal community.The community composed of Phanerochaete chrysosporium,Coriolus versicolor,Trichoderma viride and Penicillium sp.was the optimal combination,which obtained maximum values of 31.25 U/g on Li P activity and maximum values of 30.98 U/g on Mn P activity during the fermentation process.Furthermore,obvious degradation rate of 9.84% on cellulose and 10.65% on hemicelluloses were detected,which corresponded to 37.45 U/g of cellulase activity and 84.88 U/g of hemicellulase activity,respectively.Compared with control,the gene abundance of carbohydrate metabolism increased by 0.07% in this mixed fungal community,the total number of carbohydrate-active enzyme genes increased by 7118,and the gene abundances of CE1,GH5,GH9 and GH48 increased by 0.24%,0.20%,0.18% and 0.08% respectively,which were the significant factors for the strain combination’s ability to efficiently deconstruct LCC.(2)Optimising the fermentation process conditions of mixed fungal systems can effectively improve the degradation rate of substrates.The highest substrate degradation rates were achieved when the inoculum was 7%,the fermentation time was 18 d and the temperature was 25 °C.The degradation rates of cellulose and hemicellulose relative to this substance were 46.40% and 69.16%,respectively,which were 1.53 and 1.46 times higher than before the conditions were optimised.The effect of fermentation time on the degradation rate of the substrate was significant(P<0.05).Compared to the system before optimisation,the gene abundance of carbohydrate metabolism increased by 1.61% in the optimised system,the total number of carbohydrate-active enzyme genes increased by 245 and the gene abundance of CE1,GH5,GH9 and GH48 all increased by approximately 0.05%,which elucidated the microbiological mechanism of the enhanced substrate degradation rate after optimisation from a genetic perspective.(3)The anaerobic digestion process based on the efficient LCC deconstruction function of the mixed fungal system can effectively enhance the isolation and purification of lignin.After the substrate was treated by a mixed fungal fermentation combined with anaerobic digestion strategy(FF+AD group),its impurities were substantially removed,with cellulose and hemicellulose degradation rates of 87.65% and 96.34% in the substrate,respectively,and the purity of lignin in the final digestate obtained was 62.32%,approximately four times the purity of the raw material,with a recovery of 76.01%.Compared to the single anaerobic digestion process(AD group),after the substrate was fermented by mixed fungi,the cellulase activity and hemicellulase activity in the anaerobic system increased by 24.77% and 18.65%respectively,and Unclassified＿Bacteroidetes,Clostridium＿III,Petrimonas,MBA03＿norank,Caldicoprobacter,and Unclassified＿Ruminococcaceae,the sum of the abundance of the six microorganisms associated with lignocellulose hydrolysis increased by 4.89%,and the gene abundances of GH43,CE1 and GH13 increased by 0.11%,0.10% and 0.04%,respectively,which were the significant reasons why this strategy can efficiently disrupt the LCC structure while removing polysaccharides such as cellulose and hemicellulose,and ultimately achieve efficient lignin separation and purification.