Cultivation And Purification Of Ammonia-resistant Methanogens From Anaerobic Digestion Of Kitchen Waste

Garbage classification has gradually become a new trend in recent years.In this context,the utilization of food waste resources through the four-classification method is a focus of attention for different stack-holders.Kitchen waste is one of the three types of food waste,and its resource utilization is the most difficult.Anaerobic biogas production is the main technical path for the resource utilization of kitchen waste.The suppression of ammonia caused by high organic nitrogen is one of the key problems in anaerobic biogas production from kitchen waste.Therefore in this study,the cultivation and purification experiment of ammonia-resistant methanogens was carried out in a sequencing batch reactor for the anaerobic digestion of food waste,and the effects of two culture methods of domestication followed by purification and domestication-purification synchronous on the digestion performance and community structure of methanogens were analyzed.Current work also investigated the process conditions of freezedrying bacteria that were conducive to long-term storage.The comparative analysis of the digestion performance of methanogens showed that in terms of gas production in the system,the cumulative gas production and maximum methane production rate of the domestication-purification synchronous culture method were lower than that of the domestication followed by purification culture method,but the methane concentration was higher than that of the domestication followed by purification culture method.In terms of ammonia nitrogen concentration in the reactor,the change in ammonia nitrogen concentration of the domestication-purification synchronous culture method was generally smaller than that of the domestication followed by purification culture method.In terms of volatile fatty acids(VFAs)in the reactor,the organic acid consumption rate of the domestication-purification synchronous culture method was higher than that of the domestication followed by the purification culture method.And,the concentration of acetic acid and propionic acid of the domestication-purification synchronous culture method was in a low range.In terms of p H level in the reactor,the p H fluctuation of the domestication and purification synchronous culture method was lower than that of the domestication followed by purification culture method and the system stability was also better.In general,domesticationpurification synchronous culture method and smaller ammonia nitrogen concentration gradient was more suitable for cultivating ammonia-resistant methanogens.The comparative analysis of the community structure of methanogens showed that with the increase of ammonia nitrogen concentration,the OTU,Shannon and Chao indexes of archaea decreased,indicating that ammonia nitrogen was toxic to the methanogens.At the level of archaea,domestication should be done firstly.The dominant genus of methanogens under the domestication followed by purification culture method was from Methanobacterium of TAN of 0g/L condition to Methanoculleus of TAN of 4.0g/L,5.0g/L,6.0g/L condition.The dominant genus of methanogens under the domestication-purification synchronous culture was always Methanoculleus.Methanoculleus could tolerate higher concentrations of ammonia nitrogen and was the dominant flora in the high ammonia nitrogen system.The abundance of bacteria of the domestication-purification synchronous culture method was always higher than that of the domestication followed by purification culture method.Under the domestication followed by purification culture method,TAN had a significant effect on Methanosarcina(P<0.05),while under the domestication-purification synchronous culture method,TAN and other environmental factors had no significant effect on methanogens.This trend is indicating that the methanogens under the domestication-purification synchronous culture method were less affected by the concentration of ammonia nitrogen,the domestication-purification synchronous culture method was better.In general,it was better to cultivate ammonia-tolerant methanogens in a way that was synchronized with domestication-purification synchronous culture method and smaller ammonia nitrogen concentration gradientAccording to the water content and the number of viable bacteria,the freeze-drying process conditions of ammonia-resistant methanogens were determined as follows-80?,15 h,-50?,1.5h,-30?,1h,-20?,10 min,20?,2h,40?,2h,and the thickness of the bacterial liquid was 50 mm.The optimal storage temperature was 4°C and the number of viable methanogens was the highest and the water content was the lowest.The storage material was vials.The comparison experiment of freeze-dried bacterial powder and original bacterial liquid shows that under the same culture method,compared with adding freeze-dried bacterial powder.The system produces more gas after adding the bacterial liquid,but storage conditions of the bacterial liquid were more stringent.Domestication Purification of the methanogens obtained by the simultaneous culture method has higher system methane production and methane production efficiency,and better system stability.In general,the lyophilized powder of ammonia-resistant methanogens cultivated in a manner that was synchronized with domestication and purification and with a small ammonia nitrogen concentration gradient was more efficient.The research results provide a valuable information for alleviating and even overcoming the inhibition of anaerobic biogas ammonia production from kitchen waste.