The DGGE Analysis Of Microbial Communities In Anaerobic Degradation Of Petroleum Hydrocarbon And The Research On Methane Production Efficiency

The method translating petroleum hydrocarbon to methane gas with the methanogenic microbial flora in the strict anaerobic environment and mining methane gas directly or formation with gas reservoir as a strategic resource in situ reserves in the gas traps which can not only improve the level of exploitation and utilization of low grade oil reservoir,but also play the positive role in the treatment of anaerobic environment such as deep soil and marine oil pollution with oil.In this paper,the gas producing was studied on the same block range of different properties reservoir of different samples?different oil well produced water and oil?.In the same block well,microbial degradation of crude oil had similar degradation rules of gas production after the same kind of nutrition activating agent,although the depth and oil properties was different.And each station produced water of oilfield had also part of a similar microbial community structure.A group of mixed bacteria SL-302 and two strains of single bacteria SL302^-1 and SL302-2 were isolated by 240 days enrichment culture from polymer flooding reservoirs in Shengli 2-4NX302.The composition of mixed bacteria was studied by means of molecular biology,the microbial which was mainly composed of anaerobic bacillus,syntrophic bacteria and proteobacteria.Through the study of the effects of temperature,salinity,initial pH value and other environmental factors,the optimal conditions of mixed bacterial SL-302: temperature was 55?,pH value was 7,salinity was 2%,inoculation amount was 2%,polyacrylamide concentration was below 1% and the safety concentration of inhibiting sulfate reduction was 0.6%.The activation effect of carbon source,nitrogen source,phosphorus source,trace elements and other nutritional elements was studied.The best activator formula of mixed bacteria SL-302: with the foundation of inorganic salt culture medium,adjusted the trace elements of Fe²⁺,Co²⁺,Ni²⁺ concentration to 1mg·L^-1,0.8mg·L^-1,0.12mg·L^-1,respectively.Addition of carbon source,nitrogen source and phosphorus source,the glucose concentration was 0.6%,sodium nitrate concentration was 0.6%,and the two potassium dihydrogen phosphate and potassium phosphate hydrogen concentration of 1:1 was 0.15g·L^-1.In order to verify the effect of mixed bacteria SL-302 and its activation conditions,anaerobic test was carried out by selecting oil-water mixture from six wells of Liaohe,Shengli and Gudao oilfield.After 300 days activation experiment,the experimental group shengli 2-4NX302 produced 30 m L gas in 120 days,the efficiency was doubled comparing to group which only add activation experiments for 240 days culture.Two experimental groups of Gudao oilfield produced little gas.The results showed that SL-302 with its activator had a positive effect on the different nature of the crude oil,and the activation effect became worse when the crude oil quality worse.The thin oil activation effect was better than heavy oil.The sequence of the activation effect was super heavy oil < heavy oil< thin oil.According to DGGE?Deformation Gradient Gel Electrophoresis?profile,mixed bacteria SL-302 had significantly affected the composition of the microbial community structure in all stations,parts of the original microorganism of produced water disappeared in adding mixed bacteria SL-302 with the culture time increasing,and the mixed bacteria of various microbial members can survive and provide a new bacteria source to the process of gas production in the new environment.The mixed strain SL-302 with its activator can meet the needs of different oil gas production efficiency improvement.The paper finally researched the metabolism mechanism of oil degradation producing gas by the mixed strain SL-302 and analyzed the degradation rule of crude oil.The results showed that components of crude oil degradation ability: n-alkanes > aromatichydrocarbons > asphaltene.Intermediate metabolites and gas composition was studied.Methane,propane,n-pentane and other odd carbon organic gas had high content,and the proportion of acetic acid,butyric acid,twelve acid and other small molecules organic acid with even carbon increased relatively.It illustrated that reservoir anaerobic microorganism had powerful ability to degraded even carbon hydrocarbon in gas production process of anaerobic microbial degradation,and the reservoir existed terminal oxidation through the course of degradation.At the same time,the intermediate metabolite acetic acid changes with culture time increased,initial concentration of acetic acid were raised with the culture time increased and reached the highest concentration after 150 days,since then the concentration had maintained a high concentration,but the gas production had not significant inhibition under high concentration of acetic acid.The results of the paper enriched the diversity of anaerobic hydrocarbons biodegradation,and enhanced the understanding of an microbial community structure change by exogenous microbes with its activator and expanded the recognition of block range of reservoir activation strategy.Simultaneously,this study contained methanogenic mechanism on anaerobic hydrocarbon degrading research,screening and evaluation of functional microbes and had important value in process of methanogenesis by anaerobic microbial degradation enhanced in reservoir oil.