黄土地区石油污染土壤生物修复的强化技术初探

以石油为典型污染物,在本次试验前期工作筛选保藏的众多优势石油降解细菌中选择4株降解能力突出的菌,对该4菌株(分别编号为A、B、C、D)进行随机混合构建优势降解菌群。结果表明：菌群A-C-D降解石油能力最强,3 d内原油的降解率达到了39.67%,比单菌除油率提高了13.21%;对该菌群的最佳投加配比进行确定,菌群的最佳接种配比为A∶C∶D=1∶2∶0.5,3 d内菌群A-C-D在不同接种配比情况下降解率变化范围为27.8%~44.2%,最高值与最低值相差16.4%,因此菌群间各菌必须维持在一定的数量配比的情况下才能达到理想的降解效果。对影响生物修复效果的环境因素,如营养物质(C、N、P)、表面活性剂、通氧量、电子受体等进行综合考虑,通过正交试验确定菌群A-C-D的最佳修复条件为：营养物质配比C∶N∶P为75∶8∶3,表面活性剂为0.5%,通气条件为六层纱布,电子受体H₂O₂的加入量为1.5%。在最佳修复条件下,3 d内原油的降解率达到6146%,比自然条件修复下的除油率4.7%提高了56.76%,较只进行菌种强化时的最高除油率44.2%提高了约17%。     

污染土壤石油生物降解与调控效应研究

为考察修复过程中土壤石油降解效应,选用中国北方某油田区现场的原油污染土壤,进行了投加除油菌、调节氮磷营养含量和水分含量等强化措施的修复试验。经过180d修复,结果表明,土壤石油污染物去除率达70·6%,去除速率达0·15g·kg-1·d-1,与自然条件相比,石油污染物半衰期由929d减少为103d。饱和烃去除率占总石油去除的75%以上,主要为十八烷、二十七烷、二十九烷、三十一烷与三十四烷。初期投菌对石油污染土壤的生物修复具有快速启动作用,但后期多次投菌对生物修复的促进作用不明显。对氮磷营养水平的适当调节有利于生物修复进行,本试验中对除油菌生长和石油去除最有利的有效态碳氮比为C∶N=100∶1~50∶1。在石油去除过程中,土壤石油去除速率的变化与除油菌数量变化趋势基本一致,表明微生态环境调控对于污染土壤中的石油降解具有显著的促进作用。     

微生物修复石油污染地下水的实验研究

为修复陕北黄土区石油污染地下水,采用优化土著微生物菌群的生物技术,进行了地下水中石油的降解与修复实验研究。在实验装置内加入了1.5%的优化菌群制剂,优化出的菌群初步鉴定主要有:假单胞菌属、微球菌属、放线菌属、真菌类的青霉属和曲霉属等。实验结果显示,在实验装置中人为添加石油含量为182.5 mg/l、862.5 mg/l、1695.0 mg/l时,经过28~37 d的微生物修复,地下水中石油的降解率为27.47%~92.46%,而无菌对照中的石油含量变化在5%以内,说明在无菌条件下地下水中石油降解缓慢。该实验过程验证了微生物修复技术在地下水石油污染修复中的有效性,探索了应用的可行性。     

石油污染场地浅层地下水MNA原位修复潜能及微生物降解效益评估

监测式自然衰减（MNA）能够高效低耗地原位修复石油污染地下水的场地,微生物对污染物的降解对MNA过程起到了重要作用。在分析东北石油污染场地地下水中总石油烃（TPH）、电子受体的质量浓度分布和变化规律基础上,划分了微生物功能区。采用溶质通量计算法,对MNA原位修复的潜能及其微生物降解效果进行了评估。结果显示,场地微生物降解正在发生,利用的电子受体不同,划分为Mn、Fe和SO^2-₄还原区。污染通量模型计算显示：上游地区微生物降解强度不断增强,下游地区微生物降解强度不断降低。监测期内石油烃总量降低了394 kg,微生物降解为自然衰减过程中的主要作用,其贡献率为64%~93%,每个通量断面内微生物降解率为0.18~0.73 kg/d。由此可以证明,MNA可以有效地修复地下水中的石油污染。     

高效耐盐碱石油烃降解菌筛选及降解特性研究

为探究长庆油田污染土壤中微生物对石油的降解特性,在该油田多个油井附近采集了10处含油污染土壤进行石油烃降解菌的筛选、分离及降解实验。通过对筛选出的四株石油烃降解菌株5-5、5-X、9-2、10-3进行革兰氏染色、菌落形态观察、生化理化试验及16S rDNA测序,鉴定出这四株菌株分别为醋酸钙不动杆菌（Acinetobacter calcoaceticus）、不动杆菌（Acinetobacter sp.）、蒙氏假单胞菌（Pseudomonas monteilii）和乳酸不动杆菌（Acinetobacter lactucae）。14 d降解实验结果显示,这四种菌株对总石油烃的降解率分别为50.92%、51.27%、78.30%和44.39%;尤其菌株Pseudomonas sp.9-2表现出优异的降解性能,且对不同组分石油烃（正构烷烃、异构烷烃及芳烃）的降解率分别达到了94.65%、69.73%和59.07%,对长链正构烷烃也体现出了较好的降解性能。另外,抗逆性试验结果表明菌株Pseudomonas sp.9-2对pH和盐度的耐受范围分别为5.0~10.0、0.5%~6.0%,表明该菌株对盐碱环境具有较好的适应性,可用于盐碱石油污染土壤的微生物修复研究。     

土壤石油污染物微生物降解机理与修复技术研究

较为系统地概述了土壤石油污染物的微生物修复技术和近年来石油烃代谢机理以及微生物细胞膜转运石油烃机理研究等方面的进展。文中着重讨论烷烃代谢途径、影响烷烃降解酶合成的阻遏物和激活物、芳香烃代谢过程和基因方面的研究,以及细胞膜结构的改变和石油烃的迁移等重要特性,最后从宏观系统的修复技术和微观分子学方面探讨微生物降解石油污染土壤的发展方向。     

土壤残油生物降解性与微生物活性

利用色谱－质谱联机（GC／MS）、柱层析分析方法、浊度法和微生物脱氢酶法，确定土壤残油污染物的构成、油污土壤优势降解菌、残油降解过程中微生物活性及其动态变化。研究结果表明，长期残留在淄河滩土层中的石油 烃类主要由沥表肝质、胶质构成，占总烃类的60％－80％，经过90d生物降解试验，残油降解率仅为20％，污染土层中的主要优势菌为球菌和杆菌，菌群鉴定为黄假单胞菌（Xanthonmonas)、芽孢杆菌（Bacillus )等。据微生物脱氢酶动态变化的初步研究结果，微生物脱氢酶活性变化可作为衡量微生物活性变化的 重要指标。     

当地下水邂逅DNA:石油类有机污染及其生物降解

地下水科学与工程研究发展到今日,已经成为一门涉及多个领域的综合性学科。地下水污染的控制和修复研究更需要跨学科的技术和知识支持,而生物修复作为一种高效低耗修复的技术成为环境领域的研究热点。微生物因其自身特性及其对污染的降解主导特征对确定有机物污染场地的永续修复具有重要意义。简要地综述了地下水有机污染及其原位修复、有机污染物和地下环境微生物的交互作用,进一步聚焦生物降解机制、生物修复和细菌研究。在此基础上以某石油污染场地地下水为例,进行了地下水中分离微生物菌株及其降解特征的实验研究。结果表明:放线菌降解效果最好,细菌和真菌次之;两两组合降解效果好于单菌,表明存在协同作用;不同菌株混合降解率较低,表明具有拮抗作用。通过动力学实验得出对TPH的降解符合一级反应动力学方程及其降解速度和降解半衰期。就微生物对有机组分降解而言,烷烃和总石油烃降解规律相似;难降解组分降解率低,后期因烷烃转化使其浓度升高;苯浓度变化不大。微生物活性实验表明:活菌总数和脱氢酶活性与降解率呈正相关变化。运用生理生化及分子生物学方法鉴定得出了具体的菌种。     

季节性低温胁迫下陇东黄土高原油污土壤环境因子对耐冷混合菌场地生态修复的响应

为了揭示季节性低温胁迫下陇东黄土高原油污土壤环境因子对耐冷混合菌场地生态修复的响应机制,利用自主筛选构建的耐冷石油降解混合菌在甘肃省庆阳市庆城县马岭镇长庆油田陇东油泥处理站开展了为期7个月的场地修复实验,采用常规方法测定了不同季节土壤理化特性、酶活性和微生物群落特性等环境指标。结果表明：（1）在季节性低温胁迫下（9-11月）M2组（耐冷混合菌处理组）月平均降解率明显增加（P<0.01）,JZJ+M1组（金盏菊联合常温混合菌处理组）和M2组累计TPH降解率分别为15.37%±3.51%和28.64%±4.12%。（2）M2组土壤脱氢酶和多酚氧化酶在低温季节（LT）活性最高,且温度和处理存在显著交互作用（P<0.01）。在土壤营养元素方面,无论何种处理方式（JZJ+M1和M2）二者含量均为RT（常温季节）高于LT（P<0.05）,同时明显高于CK组（P<0.05）。（3）M2组土壤微生物群落Shannon-Wiener指数和均匀度指数在LT高于RT（P<0.05）。（4）NMDS和Pearson相关性分析结果显示,M2组在低温季节具有较高TPH降解率主要与土壤多酚氧化酶、脱氢酶、单月TPH降解率（μ2）和Shannon-Wiener指数有关,且均呈极显著正相关关系。通过分析环境因子的季节响应,优化场地修复在低温环境的降解条件、加速低温期石油烃降解速率,以期为陇东地区低温耐冷混合菌场地生态修复技术的推广应用提供参考和基础数据资料。     

覆膜态Fe（OH）3在厌氧条件下生物降解苯和甲苯的初探

天然条件下氯代烃的污染经常会与石油烃的污染共存,对于这种混合污染羽的治理,第一步采用粒状铁还原氯代烃,后续利用微生物和第一步产生的副产物生物降解石油烃。苯系物是石油烃中毒性较大、存留时间较长的污染物,本文利用批实验的方法研究了厌氧条件下用Fe(OH)3覆膜于石英砂表面的Fe(Ⅲ)作为电子受体降解苯和甲苯。结果表明,Fe(Ⅲ)作为电子受体时苯和甲苯能够发生厌氧生物降解,经过驯化后苯和甲苯的降解速度明显加快。降解实验表明甲苯的降解速度比苯的降解速度快,苯降解的半衰期是4.02d,甲苯降解的半衰期是3.81d。     

Isolation,biodegradation ability and molecular detection of hydrocarbon degrading bacteria in petroleum samples from a Brazilian offshore basin

The detection of microorganisms with potential for biodeterioration and biodegradation in petroleum fields is of great relevance, since these organisms may be related to a decrease in petroleum quality in the reservoirs or damage in the production facilities. In this sense, petroleum formation water and oil samples were collected from the Campos Basin, Brazil, with the aim of isolating microorganisms and evaluating their ability to degrade distinct classes of hydrocarbon biomarkers (9,10-dihydrophenanthrene, phytane, nonadecanoic acid and 5α-cholestane). Twenty eight bacterial isolates were recovered and identified by sequencing their 16S rRNA genes. Biodegradation assays revealed that bacterial metabolism of hydrocarbons occurred through reactions based on oxidation, carbon–carbon bond cleavage and generation of new bonds or by the physical incorporation of hydrocarbons into microbial cell walls. Based on the biodegradation results, selective PCR-based systems were developed for direct detection in petroleum samples of bacterial groups of interest, namely Bacillus spp., Micrococcus spp., Achromobacter xylosoxidans, Dietzia spp. and Bacillus pumilus. Primer sets targeting 16S rRNA genes were designed and their specificity was confirmed in silico (i.e. computational analysis) and in PCR reactions using DNA from reference strains as positive and negative controls. Total DNA from oil was purified and the amplification tests revealed the presence of the target bacteria in the samples, unraveling a significant potential for petroleum deterioration in the reservoirs sampled, once proper conditions are present for hydrocarbon degradation. The application of molecular methods for rapid detection of specific microorganisms in environmental samples would be valuable as a supporting tool for the evaluation of oil quality in production reservoirs.     

稠油微生物开采在新疆油田的现场应用

为了提高原油采收率,提高稠油油藏开发效益,与新疆油田合作开展了稠油微生物开采的现场试验.试验优选了混源采油菌组合,采用单井吞吐的生产方式,分两批对21口稠油开发井进行了微生物开采现场试验.经采油菌作用,作业区的稠油粘度大幅度降低,在停止注蒸汽的情况下,大多数试验井都能达到经济产能,试验得到了较高的投资回报率.结果表明,所选用的采油菌组合对胶质、沥青质含量高的稠油作用效果显著,所以说稠油微生物开采技术值得在新疆油田的稠油开发中推广应用.     

Fate of petroleum hydrocarbons and toxic organics in Louisiana coastal environments

Numerous potentially toxic compounds are entering Louisiana’s inshore and nearshore coastal environments. To a large degree there is insufficient information for predicting the fate and effect of these materials in aquatic environments. Studies documenting the impact of petroleum hydrocarbons entering Louisiana coastal wetlands are summarized. Also included are research findings on factors affecting the persistence of petroleum hydrocarbons and other toxic organics (pentachlorophenol (PCP), 2,4-dichlorophenoxyacetic acid (2,4-D), creosote, etc.) in sediment-water systems. Sediment pH and redox conditions have been found to play an important role in the microbial degradation of toxic organics. Most of the hydrocarbons investigated degrade more rapidly under high redox (aerobic) conditions although there are exceptions (e.g., 1,1,1-trichloro-2,2-bis(4-chlorophenyl) (DDT) and polychlorobiphenyls (PCBs)). Some of these compounds, due to their slow degradation in anaerobic sediment, may persist in the system for decades.     

本源微生物降解辽河原油的非烃分子结构变化

利用高效本源降解菌在不同时间范围内对辽河油样进行了微生物降解模拟实验,以研究不同降解程度原油分子结构的变化规律。微生物作用前后原油样品做族组分分离及进一步的中性氮化合物和酸性化合物分离,然后对其不同组分进行色谱质谱分析和红外光谱分析。结果表明,辽河油样中的非烃化合物经微生物作用以后,正构一元酸含量随降解时间的增加而增加,且变化幅度逐渐增大;五环三萜类羧酸中C28组分和莫烷酸系列随着降解时间的增加含量上升;轻度的微生物降解作用对咔唑、甲基咔唑、二甲基咔唑等化合物没有明显影响,中等强度降解具有一定的影响;从胶质、沥青质红外光谱图看出,微生物能够降解该原油胶质、沥青质。侧链长度随降解程度增加而变短;芳香环结构抗微生物降解能力强,缩合程度变化不大;而取代基上的烷烃易被微生物氧化,氧化程度随降解程度增加而增加,产生酸类、醚类物质。探讨原油微生物降解中非烃化合物结构及其地球化学特征对石油勘探开发都有重要指导作用。     

Population structures shift during the biodegradation of crude and fuel oil by an indigenous consortium

Petroleum and fuel oil are complex mixtures of recalcitrant hydrocarbons. The biodegradation of these hydrocarbons needs the action of a vast variety of enzymatic capacities. A microbial consortium offers the capability to degrade complex substrates through the assembly of different biochemical reactions, providing a metabolic versatility superior to axenic cultures. In this work, the microbial population dynamics, taxonomy, and the catabolic capacity of a stabilized consortium exposed to fuel and crude oil was analyzed through metagenomics. The stabilized consortium degraded 59% of crude oil components after 8 days, and 34% of fuel oil components after 130 days. Population dynamics analysis indicates that in fuel oil the biodiversity richness was higher; however, denaturing gradient gel electrophoresis similarity dendrogram shows significant changes in the microbial population during crude oil degradation. Taxonomy studies indicate a great genera divergence; only eight microbial genera were common in both samples. In crude oil, the Limnobacter sp. was the most abundant specie (15.6%), while Sphingomonas wittichii (7.9%) and Novosphingobium aromaticivorans (7.6%) were abundant in fuel oil. These microorganisms have been reported to participate in the degradation of aliphatic and aromatic hydrocarbons. Functional analysis suggests that fuel and crude oil components changed the interactions between the consortium members affecting the collective metabolic functionality.     

Degradation Kinetics of Petroleum Contaminants in Soil-Water Systems

On the basis of site investigation and sample collection of petroleum contaminants in the soil-water-crop system in the Shenyang-Fushun sewage irrigation area, the physical-chemical-biological compositions of the unsaturated zone is analyzed systematically in this paper. At the same time, the degradation kinetics of residual and aqueous oils is determined through biodegradation tests. The studies show that dominant microorganisms have been formed in the soils after long-term sewage irrigation. The microorganisms mainly include bacteria, and a few of fungus and actinomycetes. After a 110-days' biodegradation test, the degradation rate of residual oil is 9.74%-10.63%, while the degradation rate of aqueous oil reaches 62.43%. This indicates that the degradation rate of low-carbon aqueous oil     

Geochemical insights into contribution of petroleum hydrocarbons to the formation of hydrates in the Taixinan Basin,the South China Sea

Methane hydrate in the South China Sea(SCS)has extensively been considered to be biogenic on the basis of itsδ13C and δD values.Although previous efforts have greatly been made,the contribution of thermogenic oil/gas has still been underestimated.In this study,biomarkers and porewater geochemical parameters in hydrate-free and hydrate-bearing sediments in the Taixinan Basin,the SCS have been measured for evaluating the contribu-tion of petroleum hydrocarbons to the formation of hydrate deposits via a comparative study of their source inputs of organic matters,environmental conditions,and microbial activities.The results reveal the occurrence of C14-C16 branched saturated fatty acids(bSFAs)with relatively high concentrations from sulfate-reducing bacteria(SRBs)in hydrate-bearing sediments in comparison with hydrate-free sediments,which is in accord with the positive δ13C values of dissolved inorganic carbon(DIC),increasing methane concentrations,decreasing alka-linity,and concentration fluctuation of ions(Cl-,Br,SO2-,Ca2+,and Mg2+).These data indicate the relatively active microbial activities in hydrate-bearing sediments and coincident variations of environmental conditions.Carbon isotope compositions of bSFAs(-34.0％o to-21.2％o),n-alkanes(-34.5％o to-29.3％o),and methane(-70.7％o to-69.9％o)jointly demonstrate that SRBs might thrive on a different type of organic carbon rather than methane.Combining with numerous gas/oil reservoirs and hydrocarbon migration channels in the SCS,the occurrence of unresolved complex mixtures(UCMs),odd-even predominance(OEP)values(about 1.0),and biomarker patterns suggest that petroleum hydrocarbons from deep oil/gas reservoirs are the most probable carbon source.Our new results provide significant evidence that the deep oil/gas reservoirs may make a contribution to the formation of methane hydrate deposits in the SCS.