Understanding The Mechanisms Behind Enhanced Anaerobic Digestion Of Lignocellulosic Waste By Micro-aeration

Anaerobic digestion（AD）of organic substrate such as waste activated sludge,food waste,manure and lignocellulosic waste and so on cannot only produce biogas,but also eliminate the pollution from these organic wastes,demonstrating economic and environmental profits.Basing on the virtues of high carbon content,renewable and rich resource,lignocellulosic waste is the ideal substrate for AD.The stable structure of lignocellulosic waste formed by the strong interlinkage of cellulose,hemicellulose and lignin limits the hydrolytic process and results in a long AD time and low methane yield.Several studies suggested that a little breathe of oxygen or air into AD system could facilitate the hydrolysis of lignocellulosic waste,leading to a shorter fermentation time and higher methane yield.Compared with other methods,micro-aeration enhanced AD of lignocellulosic waste possesses better economic and environmental benefit for it just requires limited oxygen or air.However,the mechanisms behind enhanced AD of lignocellulosic waste by micro-aeration were not fully unraveled.Basing on the above discussions,this thesis focused on the mechanisms behind the micro-aeration enhanced AD by using corn straw as a typical substrate of lignocellulosic waste,and the main contents and conclusions were as follows:（1）Different doses of oxygen were added into AD of corn straw to obtain optimum oxygen dose for the maximum cumulative methane yield,and the effect of soluble humic acids（SHAs）on AD process was also presumed.The results indicated that low dose of oxygen could raise the cumulative methane yield,while high dose of oxygen could reduce the cumulative methane yield.The highest cumulative methane yield,the maximum potential methane yield and the shortest fermentation lag time were achieved when oxygen dose was at 0.2 m L/g·VS/d.Micro-aeration improved the relative abundance of Firmicutes,which could be benefit to the excretion of extracellular enzyme（cellulase）in AD.Higher cellulase activities accelerated the degradation of corn straw,resulting in higher contents of VFAs and soluble humic acids（SHAs）concentrations.Under micro-aerobic conditions,the relative abundances of syntrophic VFAs-oxidizing bacteria（Synergistota and Spirochaetota）and oxygen-tolerant methanogens（Methanobacterium and Methanosarcina）were increased,which facilitated the conversion of VFAs and the methanogenesis.Besides,the higher SHAs concentration under micro-aeration might enrich electron shuttles,leading to an improving syntrophic relationship among VFAs-oxidizing bacteria and methanogens.（2）SHAs precipitated by Ca²⁺and addition of micro HAs（MHAs）and nano HAs（NHAs）were used to investigate the effects of humic acids concentrations on AD of corn straw.The results showed that humic acids concentrations affected AD of corn straw.The removal of SHAs reduced the number of electron shuttles,resulting in the reduction of maximum methane producing rate.Due to the large particle size,MHAs showed no effects on the AD of corn straw.On the contrary,the addition of NHAs increased the number of electron shuttles,and raised the maximum methane producing rate.With NHAs concentration at 1 g/L,the maximum methane producing rate,lag phase time and T₈₀ in NHAs1 were 120.6%,76.7%and 85%of them in Control,respectively.NHAs raised the relative abundance of hydrolytic and fermentative bacteria（Clostridia）,VFAs-oxidizing bacteria（Cloacimonadia,Synergistotia and Spirochaetia）and electron trophic methanogens（Methanosarcina）,which could accelerate the hydrolysis and fermentation of corn straw,benefit the oxidizing conversion of VFAs and utilize the electrons from VFAs oxidizing process to reduce CO₂for methane product.The average daily methane yield was promoted by 9.6%at 1 g/L NHAs during a semi-continuous operated AD,further suggesting that humic acids could be used as DIET mediators to enhance the syntrophic relationships among VFAs-oxidizing bacteria and methanogens.（3）Four total solid（TS）contents of 4.0%,4.5%,5.0%and 5.5%were set to create different VFAs concentrations to figure out the mechanisms behind the fast conversion of VFAs in micro-aeration enhanced AD of corn straw.The results showed that with the increase of organic loading rates（OLRs）the accumulation of VFAs and p H reduction occurred in AD of corn straw.Especially,at OLRs of5.5%TS content,the acidification by the quick accumulation of VFAs led to the failure of AD digesters.The addition of HAs accelerated the conversion of VFAs and raised p H,facilitating the methane producing rate and cumulative methane yield.The maximum methane producing rate and cumulative methane yield in HAs5.0 were 44.3%and 13.8%higher than those in C5.0.The addition of HAs imposed little effect on the core bacteria related to hydrolysis and fermentation.However,it increased the relative abundance of VFAs,especially acetate oxidizing bacteria,leading to a faster conversion of VFAs and the relief of inhibition on methanogenesis.Methanobacterium adapted better to high OLRs than Methanosarcina.However,the addition of HAs led to the faster conversion of VFAs produced at high OLRs,creating a suitable environment for Methanosarcina,and the summed up relative abundance of Methanobacterium and Methanosarcina was also increased in all HAs groups,ensuring the methangenesis by reduction of CO₂ with electrons on the rail.（4）0 m L/g·VS/d oxygen dose（control）,0.8 m L/g·VS/d oxygen dose and the coupling of0.8 m L/g·VS/d oxygen dose and 1g/L humic acids were used to study the effects of micro-aeration coupled with humic acids supplement on the anaerobic process during AD of corn straw.The results showed that during AD of corn straw the combination of micro-aeration and humic acids obtained the maximum potential methane products and the cumulative methane yield,which were 13.1%and 11.1%higher than those in control.The coupling of micro-aeration and humic acids could not only stimulated the growth of Bacteroidales to excrete more cellulases and hemicellulases for faster hydrolysis and acidification of substrate,but also accelerated the direct interspecies electron transfer between acetate-oxidizing bacteria Spirochaetaceae and hydrotrophic methanogen Methanobacterium for methane production by using humic acids as electron shuttles.Furthermore,the anaerobic digester were stably operated for the reasons that the antioxidases activity,metabolism（carbohydrate metabolism）and energy metabolism（oxidative phosphorylation and methane metabolism）were enhanced by the coupling of micro-aeration and humic acids.（5）A semi-continuous operated reactor was applied to study the effects of the optimum oxygen（0.2 m L/g·VS/d）on AD performance of corn straw at 1.1,1.5 and 2.1 g·VS/L/d OLRs.The results suggested that the median daily methane yield in M1.1,M1.5 and M2.1 were 6%,10%and 8%higher than those in C1.1,C1.5 and C2.1,respectively,which indicated that micro-aeration could enhance daily methane yield in AD of corn straw at different OLRs.Micro-aeration raised the relative abundance of Clostridia,which could excrete more cellulases（endoglucanase/EC.3.2.1.4 and beta-glucosidase/EC.3.2.1.21）and hemicellulase（xylanases/EC.3.2.1.8）,leading to a faster degradation of corn straw.With the increase of OLRs,the dominate methanogens were shifted from strictly acetate utilizing methanogens（Methanosaeta）to strictly hydrogen methanogens（Methanobacterium）,and micro-aeration increased the relative abundance of Methanobacterium in M1.1,M1.5 and MA2.1.Furthermore,higher SHAs concentration and relative abundance of VFAs-oxidizing bacteria（Cloacimonadia）improved the electron transfer capacity in MA2.1,which accelerated the electron transfer rate from Cloacimonadia to Methanobacterium,increased the methane producing rate and finally promoted methane yield.