Study On Directional Culture Of Petroleum Hydrocarbon Degrading Bacteria And Mechanism Of Oil-recovery In Low Permeability Reservoirs

Crude oil continues to play a crucial role in the development of world economy with continuous increase in the demand of energy.The difficulty to enhance oil recovery from low-permeability reservoirs requires to develop an alternative and cost-effective crude oil recovery process.Compared with traditional tertiary oil recovery technologies,microbial enhanced oil recovery(MEOR)is considered an economical and environmentally-friendly tertiary oil recovery technology and has gained increasing attention over recent years.In this paper,High-throughput Sequencing and Metagenomics technology were used to study characteristics of microbial community composition in low-permeability reservoir and the mechanism of microbial degradation of crude oil.And the system of directional activation of Bacillus genus,which activated by specific nutrients,was established and applied to the field of low-permeability reservoir.The main research results are as follows:(1)Two strains of highly efficient petroleum degrading bacteria(Pseudomonas aeruginosa,HF;Bacillus subtilis,XH)were screened from the production water of low permeability reservoir.They metabolize a large number of biosurfactants under facultative anaerobic conditions,which reduce the surface tension of the medium to34.00 m N/m and 34.63 m N/m respectively.The optimal growth p H(6?9),salinity(10000?50000 mg/L),temperature(30?40?).(2)The combination of nitrogen sources 0.8%Na NO₃:(NH₄)₂SO₄=2:1 and phosphorus sources 1.4%KH₂PO₄:Na H₂PO₄=5:2 was considered the optimal nutrients for the emulsification of crude oil by biodegradation of the crude oil and formation of biosurfactants.The optimal medium composition also include 2 wt%crude oil,0.06%yeast powder,trace element(Zn SO₄ 0.3%,Ca Cl₂ 0.25%,Cu SO₄ 0.25%,Mg SO₄·7H₂O0.5 0.15%).In the process of multistage subcultures with the optimal nutrients and simulated oil reservoir condition,Bacillus became the final genus(after sixth stage subculture)and showed activity to biodegrade n-alkanes and n-alkyl cycloalkanes in crude oil.The overall degradation rate of functional bacteria to crude oil is 33.5%,and n-alkane is 53?75%,and that of n-alkyl cycloalkanes was in the range of 67%to 75%.(3)The droplet size distribution shows that most of oil droplets have sizes between2 and 15?m.The crude oil droplets with sizes of 2?10?m counted to 69%in the first subculture broth,which decreased to 39%in the second subculture broth,and then gradually increased to 52%,65%,75%,and 80%,respectively,in the broths of the third to sixth subculture.The results of droplet size distribution show that the composition of bacteria community changed in a trend favoring enhanced emulsion of the crude oil.(4)Overall,there were not significant changes in the total abundances of O₁,N₁O₁and N₁O₂ classes of heteroatomic compounds which have relatively low abundances in crude oil before and after biodegradation.The abundance of heteroatomic class N₁decreased after biodegradation,as so for each corresponding DBE.Oppositely,the overall abundance as well as that for each DBE in the O₂ class increased after biodegradation,due to the oxidation reactions during biodegradation.(5)In the study of Metagenomics of functional bacteria for petroleum hydrocarbon degradation,There are six major classification units of petroleum hydrocarbon degradation proteins,i.e.energy production and conversion,amino acid transport and metabolism,carbohydrate transport and metabolism,transcription,inorganic ion transport and metabolism,functional location,etc.,which are C,E,G,K,P,S,accounting for 5.03%,7.52%,5.78%,6.23%,6.33%and 25.5%,respectively.In metabolism,there are 5625 genes for hydrocarbon metabolism,accounting for 21.98%of metabolism,and 4897 genes for amino acid metabolism,accounting for 19.13%of metabolism.There are alkane degrading genes 1-monooxygenase(alkm,EC:1.14.15.3),alcohol dehydrogenase(ADH,EC:1.1.1.1),aldehyde dehydrogenase(ALDH,EC:1.2.1.3),which can form a complete alkane degradation path.The corresponding gene coding numbers in the bacteria are 3,56 and 3,respectively.The results show that there are many genes involved in material metabolism and energy metabolism,which may be the reason why the petroleum hydrocarbon degrading bacteria can be activated by inorganic salt ions and degrade hydrocarbons in crude oil.(6)Through microscopic simulation visual model,observation and analysis of distribution and flow of the remaining oil in the process of water flooding and microbial oil displacement were conducted.Microbes attach to and surround oil droplets.Droplets are deformed by the action of microbes and break up into smaller droplets.Smaller droplets are in turn broken up into micro droplets and are able to escape pore spaces.and the recovery of crude oil is close to 91.4%.The emulsification of remaining crude oil can be further improved by microbial composite surfactant,and the recovery of crude oil can be further improved by 2.6%.The microbial composite silica further split the crude oil into smaller oil drops,which can drive nearly 99%of the crude oil.(7)The field test of microbial flooding for 2 injection and 10 production wells has been carried out in Erdong block of Karamay Oilfield in Xinjiang.The oil production of well T20248 increased from 14 t to 276.8 t,that of T20371 increased from the 17 t to187.86 t,that of T20427 increased from 47 t to 224.12 t,that of T20421 increased from48 t to 179.27 t,that of two-way well of T20422 increased from 34 t to 119.21 t,that of T20247 increased from 14 t increases to 101.5 t.The oil production of the whole experimental well group increased about 1500 t in five months(15%of the decline was not deducted).