Pretreatment by crude enzymatic liquid from Trichoderma harzianum and Aspergillus sp improving methane production performance during anaerobic digestion of straw

Abstract: In order to improve methane production from maize straw, crude enzymes from Trichoderma harzianum and Aspergillus sp. by solid-state fermentation were used to pretreat the maize straw before anaerobic fermentation. The experiment was divided into two parts: pretreatment and anaerobic digestion. Enzymatic treatments were done using enzymes (enzyme from Trichoderma harzianum, named as enzyme T; and enzyme from Aspergillus sp, named as enzyme A) inoculated into dry yellow maize straw at 50 ℃ for 6 days. The enzymes had the ability to hydrolyze cellulose and hemicellulose, and the CMCase and xylanase activity in enzyme T were (12.38 ± 0.69) and (230.53 ± 23.03) U/mL, from enzyme A were (3.32 ± 0.68) and (891.77±27.36) U/mL, respectively. Control samples (CK) were prepared using the same conditions as in enzymatic treatment, but with no enzyme added. Maize straw was pretreated for 6 days as the fermentation substrates and the sludge was inoculum. The working volume of each suction-flask reactor was 1 L, and 300 mL inoculum was loaded into each reactor. Batch tests were conducted in a walk-in incubation room for 20 days at (35±2) ℃. Triplicate reactors were run for each condition. Inoculum without any added feedstock was used as a blank. The biogas production and methane composition were measured every day and the pH value, volatile fatty acids (VFA), soluble chemical oxygen demand (sCOD) and alkalinity were measured during the fermentation. The relationship between VFA and sCOD or VFA and alkalinity during anaerobic fermentation was analyzed. High-throughput sequencing was used to determine the microbial community structure in 24 hours fermentation samples, in order to detect the effect of enzymatic pretreatment and its impact on the bacteria and archaea community in anaerobic fermentation system. The results showed that VFA concentration increased significantly after treated by enzyme T and enzyme A, and mainly reflected in the accumulation of acetic acid content at 0 d. After 2 days fermentation, propionic acid was the main component of VFA. At the 1st day, the ratio of alkalinity/VFA and sCOD/VFA of enzyme T group and enzyme A group were significantly increased, which were 18.69 and 8.69 in enzyme T group, and 10.89 and 7.18 in enzyme A group, respectively. These changes were mainly due to the decrease of VFA. However, CK group in both alkalinity/VFA and sCOD/VFA showed a gradual decline trend. The cumulative methane yield by enzyme T group and enzyme A group were increased by 7.79% and 10.06%, respectively, compared with the control. After 24 hours fermentation, the abundance of 9 genus of bacteria in enzyme T group were increased than CK group, and Clostridium, vadinBC27 and Ruminofilibacter were related to the degradation of cellulose. The bacteria in enzyme A group was similar with CK group, and only 5 genus were significant difference. The main archaea were Methanosaeta, Bathyarchaeota, Methanosarcina, Deep sea Euryarchaeotic Group-DSEG and Methanobacterium. The pretreatment could affect the microbial structure in the fermentation system. Effective pretreatment plays an important role in anaerobic digestion of lignocellulose. This study provides important theoretical and technical support for the use of enzymes in methane production of agricultural wastes.