油藏微生物的代谢特征与提高原油采收率技术

油藏是一个高温、高压、少氧、寡营养和封闭的极端环境,油田经过多年注水开发后,在油藏内部形成了相对稳定的微生物群落体系,这些微生物以石油烃分解为起始,形成了一个复杂的食物链,对油藏碳、硫和金属离子的元素地球化学循环起着非常重要的作用。微生物提高原油采收率技术(MEOR)是利用微生物及其代谢产物与油藏和原油发生作用来提高原油采收率的一种新技术,具有成本低、适应性强和环境友好等特点,因此有望成为未来化学驱后油藏和高含水油藏进一步提高采收率的重要手段。对油藏内源微生物及其介导的生化反应,微生物采油原理、发展历程和现场试验进行综述,并提出了未来的发展方向。     

油藏微生物的代谢特征和生态结构调控

油藏环境中孕育着多种多样的微生物,这些微生物代谢类型多、变异性大,在微生物生态系统中占有重要位置。研究油藏中微生物的代谢特征和相互之间的生态关系,有助于提升对微生物提高采收率机理的认识。本文对油藏环境中常见微生物类群的代谢特征与功能、生态结构与调控等进行了简要综述。     

Effects of oil reservoir conditions on the production of water‐insoluble Levan by Bacillus licheniformis

Bacillus licheniformis produced a water‐insoluble levan which has potential application as a selective plugging agent in microbial enhanced oil recovery (MEOR). The microorganism grew on sucrose, glucose, and fructose but produced levan only on sucrose. Plugging may thus be selectively controlled in the reservoir by substrate manipulation. B. licheniformis and a crude preparation of its extracellular enzymes were evaluated for their ability to produce levan under reservoir conditions. Oil reservoirs which have a temperature of less than 55°C, a pH between 6 and 9, a pressure less than 500 atm, and a salt concentration of 4% or less are potentially suitable. Examples of such reservoir conditions are found in Lloydminster on the Alberta‐Saskatchewan border, one of the largest Canadian oil reserves.     

Microbial processes in the Athabasca Oil Sands and their potential applications in microbial enhanced oil recovery

The Athabasca Oil Sands are located within the Western Canadian Sedimentary Basin, which covers over 140,200 km² of land in Alberta, Canada. The oil sands provide a unique environment for bacteria as a result of the stressors of low water availability and high hydrocarbon concentrations. Understanding the mechanisms bacteria use to tolerate these stresses may aid in our understanding of how hydrocarbon degradation has occurred over geological time, and how these processes and related tolerance mechanisms may be used in biotechnology applications such as microbial enhanced oil recovery (MEOR). The majority of research has focused on microbiology processes in oil reservoirs and oilfields; as such there is a paucity of information specific to oil sands. By studying microbial processes in oil sands there is the potential to use microbes in MEOR applications. This article reviews the microbiology of the Athabasca Oil Sands and the mechanisms bacteria use to tolerate low water and high hydrocarbon availability in oil reservoirs and oilfields, and potential applications in MEOR.     

Exopolysaccharide production by a genetically engineered Enterobacter cloacae strain for microbial enhanced oil recovery

Microbial enhanced oil recovery (MEOR) is a petroleum biotechnology for manipulating function and/or structure of microbial environments existing in oil reservoirs for prolonged exploitation of the largest source of energy. In this study, an Enterobacter cloacae which is capable of producing water-insoluble biopolymers at 37 °C and a thermophilic Geobacillus strain were used to construct an engineered strain for exopolysaccharide production at higher temperature. The resultant transformants, GW3-3.0, could produce exopolysaccharide up to 8.83 g l⁻¹ in molasses medium at 54 °C. This elevated temperature was within the same temperature range as that for many oil reservoirs. The transformants had stable genetic phenotype which was genetically fingerprinted by RAPD analysis. Core flooding experiments were carried out to ensure effective controlled profile for the simulation of oil recovery. The results have demonstrated that this approach has a promising application potential in MEOR.     

细菌在油水界面粘附的理论、特性及应用

细菌在油水界面粘附主要应用于烷烃污染的环境修复、石油开采与加工、食品加工等方面,其研究已经引起各界的广泛关注。本文基于XDLVO理论阐述细菌在油水界面的粘附机理;总结细菌表面特性、油相及水相性质对细菌粘附的影响;介绍细菌粘附作用在微生物驱油、生物乳化与破乳以及烷烃降解方面的应用,并对今后的研究方向提出展望。     

油藏铁还原微生物的研究进展

地下深部油藏通常为高温、高压以及高盐的极端环境,含有非常丰富的本源嗜热厌氧微生物,按代谢类群可分为发酵细菌、硫酸盐还原菌、产甲烷古菌和铁还原菌。从油田环境已经分离出90株铁还原微生物,如热袍菌目、热厌氧杆菌目、脱铁杆菌目、δ-变形菌纲脱硫单胞菌目、γ-变形菌纲希瓦氏菌属和广古菌门栖热球菌属等,这些菌株生长温度范围为4-85°C,生长盐度范围为0.1%-10.0%NaCl,还未见到文献报道油藏铁还原菌的耐压性研究。在油藏环境中存在微生物、矿物和流体(油/水)三者之间的相互作用,油藏中的粘土矿物能够作为微生物生命活动的载体,也能为微生物代谢作用提供电子受体。本文综述了油藏铁还原菌分离和表征的研究进展,简述了油藏铁还原菌的环境适用性,并展望了铁还原菌在提高原油采收率方面的应用前景。     

一株可乳化降解原油的Compostibacillus的分离及特性

【背景】通过实施多轮次微生物采油,华北油藏产出液菌浓达到了106个/mL以上,油藏内部已经形成了较稳定的微生物发酵场,从其中筛选出能够乳化降解原油的微生物,并在地面对其进行扩大培养,然后再应用到微驱油藏,以进一步提高微生物采油实施效果。【目的】筛选乳化降解原油性能良好的菌株,对其进行多相分类学鉴定和性能评价。【方法】利用原油为底物筛选乳化降解性能良好的菌株,通过形态特征观察、生理生化测定、16S rRNA基因序列分析等确定菌株的分类地位。通过乳化能力、降解率等方法确定菌株的原油乳化降解特性。【结果】从华北油田采集的地层水样品中分离得到一株乳化原油的菌株BLG74,经多相分类鉴定表明其是土壤堆肥芽孢杆菌(Compostibacillus humi)的新菌株,亲源性99.6%。该菌株的生长温度为30-60℃ (最适温度45℃),pH6.5-9.5(最适pH7.0),NaCl浓度0%-7%(质量体积比)。菌株BLG74在玉米浆培养基中培养,其发酵液的表面张力为56.3 mN/m,乳化力约95%,在初始原油质量浓度0.5%、温度45℃的条件下培养20d,对原油的降解率可达40.8%。【结论】菌...     

Production of biosurfactant and antifungal compound by fermented food isolate Bacillus subtilis 20B

A biosurfactant producing strain, Bacillus subtilis 20B, was isolated from fermented food in India. The strain also showed inhibition of various fungi in in-vitro experiments on Potato Dextrose Agar medium. It was capable of growth at temperature 55 degrees C and salts up to 7%. It utilized different sugars, alcohols, hydrocarbons and oil as a carbon source, with preference for sugars. In glucose based minimal medium it produced biosurfactant which reduced surface tension to 29.5 mN/m, interfacial tension to 4.5 mN/m and gave stable emulsion with crude oil and n-hexadecane. The biosurfactant activity was stable at high temperature, a wide range of pH and salt concentrations for five days. Oil displacement experiments using biosurfactant containing broth in sand pack columns with crude oil showed 30.22% recovery. The possible application of organism as biocontrol agent and use of biosurfactant in microbial enhanced oil recovery (MEOR) is discussed.     

厌氧产表面活性剂微生物提高原油采收率的研究进展

微生物强化采油(microbial enhanced oil recovery,MEOR)是近年来在国内外发展迅速的一项提高原油采收率技术。微生物在油藏中高效生产表面活性剂等驱油物质是微生物采油技术成功实施的关键之一。然而,油藏的缺/厌氧环境严重影响好氧表面活性剂产生菌在油藏原位的生存与代谢活性;油藏注空气会增加开采成本,且注入空气的作用时效和范围难以确定。因此,开发厌氧产表面活性剂菌种资源并强化其驱油效率对于提高原油采收率具有重要意义。本文综述了国内外近年来利用厌氧产表面活性剂微生物提高原油采收率的研究进展,简述了微生物厌氧产表面活性剂的相关驱油机理、菌种资源开发现状以及油藏原位驱油应用进展,并对当前的研究提出了一些思考。     

Response of microbial community structure to microbial plugging in a mesothermic petroleum reservoir in China

Microbial plugging, a microbial enhancement of oil recovery (MEOR) technique, has been applied in a candidate oil reservoir of Daqing Oil Field (China). The goal of this study is to monitor the survival of injected bacteria and reveal the response of microbial communities in field trial of microbial plugging through injection of selected microbial culture broth and nutrients. Culture-dependent enrichment and culture-independent 16S rDNA clone library methods were used. The results show that it was easy to activate targeted biopolymer-producing bacteria in a laboratory environment, and it was difficult for injected exogenous bacteria to survive. In addition, microbial communities in the oil reservoir also changed before and after the field trial. However, microbial communities, activated by fermentative medium for biopolymer-producing bacteria, appeared to show greater differences in the laboratory than in the natural reservoir. It was concluded that microbial populations monitoring was important to MEOR; results of response of microbial communities could provide a guide for the future field trials.     

Core flooding tests to investigate the effects of IFT reduction and wettability alteration on oil recovery during MEOR process in an Iranian oil reservoir

Microbial enhanced oil recovery (MEOR) refers to the process of using bacterial activities for more oil recovery from oil reservoirs mainly by interfacial tension reduction and wettability alteration mechanisms. Investigating the impact of these two mechanisms on enhanced oil recovery during MEOR process is the main objective of this work. Different analytical methods such as oil spreading and surface activity measurements were utilized to screen the biosurfactant-producing bacteria isolated from the brine of a specific oil reservoir located in the southwest of Iran. The isolates identified by 16S rDNA and biochemical analysis as Enterobacter cloacae (Persian Type Culture Collection (PTCC) 1798) and Enterobacter hormaechei (PTCC 1799) produce 1.53 g/l of biosurfactant. The produced biosurfactant caused substantial surface tension reduction of the growth medium and interfacial tension reduction between oil and brine to 31 and 3.2 mN/m from the original value of 72 and 29 mN/m, respectively. A novel set of core flooding tests, including in situ and ex situ scenarios, was designed to explore the potential of the isolated consortium as an agent for MEOR process. Besides, the individual effects of wettability alteration and IFT reduction on oil recovery efficiency by this process were investigated. The results show that the wettability alteration of the reservoir rock toward neutrally wet condition in the course of the adsorption of bacteria cells and biofilm formation are the dominant mechanisms on the improvement of oil recovery efficiency.     

Isolation and characterization of Halomonas sp. strain C2SS100, a hydrocarbon-degrading bacterium under hypersaline conditions

Aims:  To isolate and characterize an efficient hydrocarbon-degrading bacterium under hypersaline conditions, from a Tunisian off-shore oil field.
Methods and Results:  Production water collected from 'Sercina' petroleum reservoir, located near the Kerkennah island, Tunisia, was used for the screening of halotolerant or halophilic bacteria able to degrade crude oil. Bacterial strain C2SS100 was isolated after enrichment on crude oil, in the presence of 100 g l⁻¹ NaCl and at 37°C. This strain was aerobic, Gram-negative, rod-shaped, motile, oxidase + and catalase +. Phenotypic characters and phylogenetic analysis based on the 16S rRNA gene of the isolate C2SS100 showed that it was related to members of the Halomonas genus. The degradation of several compounds present in crude oil was confirmed by GC–MS analysis. The use of refined petroleum products such as diesel fuel and lubricating oil as sole carbon source, under the same conditions of temperature and salinity, showed that significant amounts of these heterogenic compounds could be degraded. Strain C2SS100 was able to degrade hexadecane (C₁₆). During growth on hexadecane, cells surface hydrophobicity and emulsifying activity increased indicating the production of biosurfactant by strain C2SS100.
Conclusions:  A halotolerant bacterial strain Halomonas sp. C2SS100 was isolated from production water of an oil field, after enrichment on crude oil. This strain is able to degrade hydrocarbons efficiently. The mode of hydrocarbon uptake is realized by the production of a biosurfactant which enhances the solubility of hydrocarbons and renders them more accessible for biodegradation.
Significance and Impact of the Study:  The biodegradation potential of the Halomonas sp. strain C2SS100 gives it an advantage for possibly application on bioremediation of water, hydrocarbon-contaminated sites under high-salinity level.     

Microbially enhanced oil recovery from carbonate reservoirs

About half of the world's oil production is from carbonate formations. However, most of the research in microbially enhanced oil recovery (MEOR), a potentially important tertiary recovery technology, has focused on sandstone reservoirs because, in general, they are geologically simpler than carbonate reservoirs and easier to model in the laboratory. Carbonate formations have a wide range of pore geometries and distributions, resulting in complex flow dynamics. The low matrix permeabilities and the dual porosity characteristics of most carbonate formations, coupled with the chemistry of carbonates, have slowed implementation of enhanced oil recovery methods. A review of the data on carbonate reservoirs in Dwight's Energydata TOTL System indicated that 40% of the oil‐producing carbonate reservoirs surveyed in the United States have environmental, geological, and petrophysical conditions that would make them candidates for MEOR. A review of a number of MEOR field trials showed that rates of oil production could be increased by as much as 200%. Microbial activity in these trials was probably due to that of indigenous populations rather than the microorganisms injected for the trials. Detrimental effects such as loss of injectivity and increased souring were not reported. Based on analysis of the geology and petrophysical characteristics of carbonates, two common mechanisms of MEOR, microbial acid production and microbial gas production, are especially suited for application in carbonate reservoirs.     

Growth and Starvation of a Strain of Klebsiella pneumoniae Isolated from a Brazilian Oil Formation

Summary The changes that occur in cell size and shape during experimental starvation and resuscitation of bacteria may find practical application in the biotechnological processes used in microbial enhanced oil-recovery (MEOR), in in situ bioremediation and in the creation of biobarriers. In the work described here, the aim was to observe certain aspects of the response of a strain of Klebsiella pneumoniae, isolated from oil producer wells belonging to PETROBRAS (National Oil Company of Brazil), to starvation and growth under various culture conditions. Cells of K. pneumoniae were taken from an exponentially-growing culture, washed and suspended in phosphate-buffered saline solution, without nutrients, where they remained for 91 days. Aliquots were withdrawn periodically, to perform viable and total cell counts and measure the optical density and cell dimensions. The starved cells were resuspended in sodium citrate medium, where they recuperated and exhibited a typical growth curve. The results indicate that this strain is a viable option for future trials on the transport and growth of microorganisms inside porous media, aimed at possible applications in MEOR.     

从大庆原油样品中分离和初步筛选菌株

目的:根据目标菌株的特性,应用选择培养基筛选出提高石油三次采油采收率的菌株,并对其进行相关性质的鉴定。方法:从大庆原油样品中经过富集培养,平板分离。结果:获得2株细菌,1号菌株属于假单胞菌属,2号菌株属于芽孢杆菌属。对两株细菌进行了定性分析,结果表明两株细菌均能产酸、产气、产表面活性剂。证明其具有降低原油黏度的作用。     

Biosurfactant production using molasses and whey under thermophilic conditions

Biosurfactant production was studied by Bacillus licheniformis K51, B. subtilis 20B, B. subtilis R1 and Bacillus strain HS3 using molasses or cheese whey as a sole source of nutrition at 45 degrees C. The isolates were able to grow and produce biosurfactant under shaking as well as static conditions. Maximum biosurfactant production was achieved with molasses at 5.0-7.0% (w/v). The biosurfactant retained its surface-active properties after incubation at 80 degrees C at a wide range of pH values and salt concentrations for nine days. Oil displacement experiments in sand pack columns with crude oil showed 25-33% recovery of residual oil.     

油藏微生物群落研究的方法学

油藏微生物群落的解析和认知是开发和应用微生物采油技术的基础。利用各种提高油藏微生物可培养性的方法和非培养技术解析不同油藏微生物的群落结构、功能和多样性,对定向调控油藏微生物群落、开发和应用有效微生物驱油技术具有重要的指导意义。通过调查新近发展的提高微生物可培养性的方法和措施以及不依赖于培养的分子微生物生态学技术,总结了油藏微生物群落研究方法学的最新进展。提高微生物可培养性的方法和措施主要通过模拟微生物的生存环境,减少富营养的毒害作用、添加信号分子维持微生物细胞间的作用和提供新型电子供体和受体等手段采用稀释法、高通量培养法等方法得以实现;不依赖于培养的分子微生物生态学技术主要包括荧光原位杂交、末端限制性片断长度多态性分析、变性梯度凝胶电泳和构建克隆文库等技术。这些方法学的进展为更有效的获得各种油藏微生物资源、调控油藏微生物群落以提高石油采收率提供理论指导。     

Production and characterization of a glycolipid biosurfactant from Bacillus megaterium using economically cheaper sources

Criteria selected for screening of biosurfactant production by Bacillus megaterium were hemolytic assay, bacterial cell hydrophobicity and the drop-collapse test. The data on hemolytic activity, bacterial cell adherence with crude oil and the drop-collapse test confirmed the biosurfactant-producing ability of the strain. Accordingly, the strain was cultured at different temperatures, pH values, salinity and substrate (crude oil) concentration in mineral salt medium to establish the optimum culture conditions, and it was shown that 38°C, 2.0% of substrate concentration, pH 8.0 and 30‰ of salt concentration were optimal for maximum growth and biosurfactant production. Laboratory scale biosurfactant production in a fermentor was done with crude oil and cheaper carbon sources like waste motor lubricant oil and peanut oil cake, and the highest biosurfactant production was found with peanut oil cake. Characterization of partially purified biosurfactant inferred that it was a glycolipid with emulsification potential of waste motor lubricant oil, crude oil, peanut oil, diesel, kerosene, naphthalene, anthracene and xylene.     

油藏微生物及其压力适应机制

目前我国油田开发主要处于高含水后期,微生物驱提高石油采收率技术(MEOR)以低成本、环境友好等独特的优势引起了石油工业界的重视。实际上,经过半个多世纪的发展,MEOR已经成为提高采收率的重要前沿技术。高压是油藏的主要环境特征,在影响油藏微生物生存与活性等方面具有重要作用。本文从油藏及其微生物的主要特征、微生物对高压环境的适应机制以及高压下微生物降解烃的代谢特征等方面进行了综述。介绍了对油藏微生物资源、群落结构、微生物在油水相中分布的认识,微生物乳化原油机制,以及微生物在油藏厌氧环境中协同代谢、受温度和压力影响的特点,并列举了MEOR的矿场应用。在高压适应机制上,微生物主要通过改变和调整细胞膜结构、增加胞内脂质组分和表达胞内特殊酶等作用来实现对压力的适应;在高压下烃降解微生物代谢速率低于常压,而且耐压菌和嗜压菌具有不同的烃降解效率。