生物表面活性剂合成条件的优化及提取方法

利用油平板筛选法和排油圈法从辽河某油田的土壤样品中分离筛选高效的生物表面活性剂产生菌,通过16S rRNA基因序列分析对所筛选菌株进行鉴定。采用L16(45)正交实验对所筛选菌株合成生物表面活性剂的条件进行优化。比较5种提取方法对发酵液中生物表面活性剂的提取效果。共分离得到17株菌,其中菌株A3、As和Y产表面活性剂的能力最强。16S rRNA基因序列分析表明,这3株菌均为假单胞菌属(Pseudomonas)。菌株A3、As、Y和A3-As混合菌的生物表面活性剂提取量较优化前有明显的提高,其产量分别是优化前的8.4倍、6.4倍、5.6倍和5.7倍,其中,菌株As的生物表面活性剂产量最高,可达20.55 g/L。对于3种纯菌和A3-As混合菌而言,5种提取方法中均以CHCl_3萃取的量最高;而对其他混合菌,5种提取方法的差别并不显著。菌株A3、As、Y均属于高效的生物表面活性剂产生菌,具有很好的应用前景。     

石油降解菌XD-1产表面活性剂的性能

从含油废水中筛选分离到1株原油降解菌XD-1,鉴定为假单胞菌(Pseuomonas sp.).初步实验表明菌XD-1具有较强的产表面活性剂乳化原油的作用,对该菌的产表面活性剂性能进行了研究.实验证明,菌XD-1所产表面活性剂为脂肽类物质,菌在生长对数期产表面活性剂,表面活性剂的产生为生长相关型;充足的碳源是产表面活性剂的必需条件,菌利用原油为碳源时能持续大量地产表面活性剂;原油和尿素为产表面活性剂的最适碳源和氮源,菌XD-1产表面活性剂的最佳营养培养基组成为葡萄糖10 g,尿素4 g,磷酸二氢钾1 g,微量元素液4 mL,水1 L,pH 8.0.     

石油降解菌的筛选优化及其对油污土壤的修复特性

分别以牛肉膏蛋白胨-布氏哈斯培养基、蓝色凝胶培养基作为初筛和复筛培养基,从石油污染土壤中筛选出2株可产生微生物表面活性剂的石油烃降解菌。并将菌株投加到油污土壤中进行修复研究,考查了不同影响因素对修复效果的影响。研究结果表明,(1)2株菌对中度石油污染土壤有较好的修复效果,向油污土壤中直接投加菌株修复70 d时对石油烃的去除率为52%;(2)向油污土壤中投加降解菌并同时补充氮营养液,修复70 d时对土壤中总石油烃的去除率可达到75%;对土壤中正构烷烃的去除率为66%;(3)与土壤的含水率及土著菌的降解效果相比,向油污土壤中投加降解菌以及补充氮磷营养液是影响石油污染土壤修复效果的关键因素。     

一株产生物表面活性剂耐盐碱菌株的分离及鉴定

从黄河三角洲石油污染盐渍土中分离获得产生物表面活性剂的耐盐碱菌株,用于治理该地区的土壤污染问题。采用富集培养方法,分离得到4株优势菌,其中菌株BG降表面张力的能力最强。经生理生化实验和16SrRNA序列分析,鉴定菌株BG属于芽孢杆菌属(Bacillus sp.)。对该菌株产生物表面活性剂的性能进行研究,结果表明:菌株BG在37℃、180r/min下振荡培养48h,可将发酵液的表面张力由初始的58.4mN/m降低至25.1mN/m;温度对菌株BG产生物表面活性剂的影响较大,最适温度是37℃,此温度下菌株BG有良好的耐盐碱特性,显著降低发酵液表面张力。采用酸沉法提取了菌株BG产生的生物表面活性剂,鉴定为一种脂蛋白类物质,其中脂类和蛋白质质量分数分别为64.7%、35.3%。     

石油降解菌XD-1产表面活性剂的性能

从含油废水中筛选分离到1株原油降解菌XD-1,鉴定为假单胞菌（Pseuomonas sp.）.初步实验表明菌XD-1具有较强的产表面活性剂乳化原油的作用,对该菌的产表面活性剂性能进行了研究.实验证明,菌XD-1所产表面活性剂为脂肽类物质,菌在生长对数期产表面活性剂,表面活性剂的产生为生长相关型;充足的碳源是产表面活性剂的必需条件,菌利用原油为碳源时能持续大量地产表面活性剂;原油和尿素为产表面活性剂的最适碳源和氮源,菌XD-1产表面活性剂的最佳营养培养基组成为葡萄糖10 g,尿素4 g,磷酸二氢钾1 g,微量元素液4 mL,水1 L,pH 8.0.     

一株生物表面活性剂产生菌的鉴定及发酵条件优化

从新疆克拉玛依油田含油污水中筛选获得一株生物表面活性剂产生菌BM-9,经过16S r DNA鉴定,该菌株为铜绿假单胞菌(Pseudomonas aeruginosa)。紫外光谱和红外光谱分析结果表明,菌株BM-9所产的生物表面活性剂为一种糖脂类生物表面活性剂。以表面张力(ST)和乳化指数(E24)为指标对菌株BM-9的发酵条件进行优化,得到最佳发酵碳源为2%葡萄糖、初始p H值为7.0、发酵温度为35℃。在最佳发酵条件下,提纯获得生物表面活性剂为淡黄色固体粉末,产量为108.7 mg/L,临界胶束浓度为85.82 mg/L,能将水溶液的表面张力降至26.6 m N/m,对柴油的乳化指数为72.6%,具有较大的工业应用价值。     

铜绿假单胞菌NY3所产表面活性剂对原油降解的影响

铜绿假单胞菌NY3是从石油污染土壤中分离出的一株能快速代谢疏水性化合物的菌种。研究了该菌产表面活性剂及对原油降解的作用。实验表明,在敞开体系中,投加82 mg/L NY3菌产的鼠李糖脂,240 h能使NY3对原油的降解率提高50%。投加甘油使NY3产鼠李糖脂与降解原油同步进行,与投加鼠李糖脂对原油降解的促进作用相近。投加9‰甘油使NY3在168 h对原油的降解率提高43%。NY3菌能同时降解原油中的直链烷烃及菲(Pr)和芘(Ph)等多环芳烃。在敞开体系中用少量甘油使产鼠李糖脂和降解原油同步进行,节约处理费用。研究结果为NY3菌株在露天石油污染环境修复中的应用奠定了基础。     

脂肽生物表面活性剂作用下多菌种石油生物降解影响因素优化

探索多种菌种降解石油过程中菌种和脂肽生物表面活性剂的作用,筛选石油降解的主要因素及最佳合,并为石油污染物的降解机理研究和石油污染修复提供指导。基于正交实验筛选主要影响因素,采用Box-Behnken实验探讨各因素最佳水平。正交实验中脂肽生物表面活性剂是多菌种降解石油过程中最主要的影响因素,在Box-Behnken实验中,其能显著地影响石油降解率。菌种降解能力是石油饱和烃组分生物降解的最主要影响因素,但脂肽生物表面活性剂是芳烃、胶质和沥青质组分降解的最主要的影响因素。研究所用菌种中,解淀粉芽孢杆菌(Bacillus amyloliquefaciens)和假单胞菌(Pseudomonas aeruginosa)在石油降解过程最重要,是本实验的石油降解最优菌。菌种和脂肽生物表面活性剂的添加浓度配比对于石油降解具有重要的影响。解淀粉芽孢杆菌和假单胞菌添加量5%,脂肽生物表面活性剂粗品添加量200 mg·L~(-1)的降解效果最优,理论上,最高降解率可达63.78%,验证降解率达到了53.89%,相对于多菌种正交实验最高降解率提高了5.54%。利用正交实验和Box-Behnken实验筛选最优降解菌和最优菌种组合的方法,具有分析因素多、实验量少等优点,具有较好的应用前景。     

石油降解菌产表面活性剂的条件优化

为了了解陕北石油降解菌产表面活性剂的情况,对实验菌株CT-6以原油为唯一碳源时产表面活性剂的排油活性、表面张力和乳化性能进行了研究,并对实验菌株产表面活性剂的条件进行了优化.结果表明,排油圈直径达到8.0cm,表面张力降低到33 mN/m,乳化指数达89.9％,石油降解率达到68.0％;优化后的条件为:接种量10％、pH为8、温度28℃、转速220 r/min,该条件下,排油圈直径增加了13.1％,7d时乳化指数达到93.0％.     

一株产表面活性剂石油降解菌筛选及其特性

为了从含有原油的样品中筛选出降解性能良好的产生表面活性剂菌株。利用富集、驯化培养、不同配方培养基筛选和排油活性测定,筛选出产高效生物表面活性剂的菌株。在形态特征和生理生化鉴定的基础上,对其16S rDNA序列进行了分析,并分别采用称重法和薄层层析法(TLC)测定其原油降解率和鉴别其表面活性剂的类型。进一步考察了不同种类的碳源和氮源、不同梯度的温度、盐度、酸碱度对其生长和排油活性的影响。筛选出一株高效脂肽类生物表面活性剂的菌株A-08,对其进行形态学、生理生化特性及16S rDNA序列同源分析,此菌株为荧光假单胞菌(Pseudomonas fluorescens)。A-08最适条件为碳源是食用油和液体石蜡,氮源为(NH_4)_2SO_4,温度40℃,盐度20%,pH 7.0,培养7 d后原油降解率为40.37%。实验筛选的菌株能适应较广泛的环境条件,可以对石油污染进行有效生物修复,具有较好的意义和应用价值。     

油污土中降解柴油细菌的分离鉴定及降解能力研究

从所采集柴油污染土壤样品中富集、分离得到柴油降解优势菌株,命名为B-3和B-4.根据其生理生化性质及16S rDNA序列比对分析,确定2株菌分别属于Tetrathiobacter kashmirensis、假单胞菌属(Pseudomonas sp.).由于实验中,B-3的生长曲线较特殊,故以B-3和典型石油烃降解菌假单...     

Evaluation of B. subtilis SPB1 biosurfactants' potency for diesel-contaminated soil washing: optimization of oil desorption using Taguchi design

Low solubility of certain hydrophobic soil contaminants limits remediation process. Surface-active compounds can improve the solubility and removal of hydrophobic compounds from contaminated soils and, consequently, their biodegradation. Hence, this paper aims to study desorption efficiency of oil from soil of SPB1 lipopeptide biosurfactant. The effect of different physicochemical parameters on desorption potency was assessed. Taguchi experimental design method was applied in order to enhance the desorption capacity and establish the best washing parameters. Mobilization potency was compared to those of chemical surfactants under the newly defined conditions. Better desorption capacity was obtained using 0.1 % biosurfacatnt solution and the mobilization potency shows great tolerance to acidic and alkaline pH values and salinity. Results show an optimum value of oil removal from diesel-contaminated soil of about 87 %. The optimum washing conditions for surfactant solution volume, biosurfactant concentration, agitation speed, temperature, and time were found to be 12 ml/g of soil, 0.1 % biosurfactant, 200 rpm, 30 °C, and 24 h, respectively. The obtained results were compared to those of SDS and Tween 80 at the optimal conditions described above, and the study reveals an effectiveness of SPB1 biosurfactant comparable to the reported chemical emulsifiers. (1) The obtained findings suggest (a) the competence of Bacillus subtilis biosurfactant in promoting diesel desorption from soil towards chemical surfactants and (b) the applicability of this method in decontaminating crude oil-contaminated soil and, therefore, improving bioavailability of hydrophobic compounds. (2) The obtained findings also suggest the adequacy of Taguchi design in promoting process efficiency. Our findings suggest that preoptimized desorption process using microbial-derived emulsifier can contribute significantly to enhancement of hydrophobic pollutants' bioavailability. This study can be complemented with the investigation of potential role in improving the biodegradation of the diesel adsorbed to the soil.     

Inhibitory properties of Enterococcus spp. isolated from faeces of healthy dogs

The aim of the present study was to evaluate the inhibitory effect by the cross-streak method of nine Enterococcus faecium strains isolated from faeces of healthy dogs and their treated and non-treated cell-free supernatant (CFS) by the well-diffusion test on the growth of potentially pathogenic bacteria isolated from clinical cases and aflatoxigenic Aspergillus section Flavi and the consequent aflatoxin B₁ (AFB₁) production. Results obtained from the cross-strake assay showed that E. faecium MF1, GJ18 and GJ40 presented the major inhibitory activity against all pathogenic strains assayed; E. faecium GJ40 produced the larger inhibitory zones (26–27 mm). Well-diffusion test results showed that the majority of the enterococci strains CFS had antimicrobial activity against the pathogenic microorganisms, especially on Gram negative indicators. Cell-free supernatant of E. faecium GJ40 was the one that produced the largest inhibition zones (14 to 21 mm) in the majority of the indicator microorganisms assayed. All supernatants treated with 10 N NaOH (pH6) showed no inhibitory effect on the indicator strain assayed. With respect to fungal inhibition, any of the CFS assayed significantly inhibited the Aspergillus strains growth. But, in general, all CFS reduced AFB₁ production from 8 to 87%. The results demonstrate that enterococci isolated from healthy dog feaces produce substances with the capacity to inhibit some potential pathogenic bacteria growth and the capacity of inhibiting or reducing the AFB₁ production in vitro.     

Factors affecting biosurfactant production by oil degrading Aeromonas spp. isolated from a tropical environment

An Aeromonas spp. was isolated from tropical estuarine water. The organism grew on crude oil and produced biosurfactant that could emulsify hydrocarbons. The peak growth and biosurfactant production was on the 8th day. The organism grew on a range of hydrocarbons that include crude oil and hexadecane while no growth was recorded on some hydrocarbons that include benzene. The biosurfactant produced by the organism emulsified a range of hydrocarbons with diesel (E24=65) as the best substrate and hexane (E24=22) as the poorest. After purification, the biosurfactant was found to contain about 38% carbohydrate and an unidentified lipid. No protein was present in the purified biosurfactant. Production of biosurfactant was highest in medium with glucose and lowest in the medium with diesel+acetate. Soybean was the best nitrogen source for biosurfactant production. The activity of the biosurfactant was enhanced optimally at NaCl concentration of 5%, pH of 8.0 and temperature of 40 degrees C. The biosurfactant retained 77% of its original activity after 120 min of exposure to heat at a temperature of 100 degrees C. Biosurfactant may be produced with this organism using non-hydrocarbon substrates such as glucose and soybean that are readily available and would not require extensive purification for use in food and pharmaceutical industries.     

红三叶草根际区石油降解菌的筛选及降解性能

从石油污染的土壤红三叶草（nifoliumrepensLinn）根际修复区中分离筛选得到4株以原油作为惟一碳源和能源进行生长繁殖的高效石油降解菌。通过菌落形态、显微镜个体形态观察、生理生化鉴定以及菌株16SrDNA序列分析,初步鉴定4株优势降解菌分别为动性杆菌、藤黄微球菌、蜡状芽孢杆菌和短小芽孢杆菌。采用气相色谱／质谱（GC／MS）法分析4株混合菌对石油烃的降解性能。结果表明：在摇床培养条件下,混合菌54d对总石油烃的生物降解率达到90．50％,较对照高67．72％。随着生物降解时间的延长,石油组分中的正构烷烃、异构烷烃及环烷烃相对总量均呈减小趋势,而芳香烃和其他醇类、醛和酸类的相对含量则有所增加。     

Utilization of a biosurfactant foam/nanoparticle mixture for treatment of oil pollutants in soil

Oil contamination has become a primary environmental concern due to increased exploration, production, and use. When oil enters the soil, it may attach or adsorb to soil particles and stay in the soil for an extended period, contaminating the soil and surrounding areas. Nanoparticles have been widely used for the treatment of organic pollutants in the soil. Surfactant foam has effectively been employed to remediate various soil contaminants or recover oil compounds. In this research, a mixture of biosurfactant foam/nanoparticle was utilized for remediation of oil-contaminated soil. The results demonstrated that the biosurfactant/nanoparticle mixture and nitrogen gas formed high-quality and stable foams. The foam stability depended on the foam quality, biosurfactant concentration, and nanoparticle dosage. The pressure gradient change in the soil column relied on the flowrate (N₂ gas + surfactant/nanoparticle mixture), foam quality, and biosurfactant concentration. The optimal conditions to obtain good quality and stable foams and high oil removal efficiency involved 1 vol% rhamnolipid, 1 wt% nanoparticle, and 1 mL/min flowrate. Biosurfactant foam/nanoparticle mixture was effectively used to remediate oil-contaminated soil, whereas the highest treatment efficiency was 67%, 59%, and 52% for rhamnolipid biosurfactant foam/nanoparticle, rhamnolipid biosurfactant/nanoparticle, and only rhamnolipid biosurfactant, respectively. The oil removal productivity decreased with the increase of flowrate due to the shorter contact time between the foam mixture and oil droplets. The breakthrough curves of oil pollutants in the soil column also suggested that the foam mixture’s maximum oil treatment efficiency was higher than biosurfactant/nanoparticle suspension and only biosurfactant.

     

Removal of hydrocarbon from refinery tank bottom sludge employing microbial culture

Accumulation of oily sludge is becoming a serious environmental threat, and there has not been much work reported for the removal of hydrocarbon from refinery tank bottom sludge. Effort has been made in this study to investigate the removal of hydrocarbon from refinery sludge by isolated biosurfactant-producing Pseudomonas aeruginosa RS29 strain and explore the biosurfactant for its composition and stability. Laboratory investigation was carried out with this strain to observe its efficacy of removing hydrocarbon from refinery sludge employing whole bacterial culture and culture supernatant to various concentrations of sand–sludge mixture. Removal of hydrocarbon was recorded after 20 days. Analysis of the produced biosurfactant was carried out to get the idea about its stability and composition. The strain could remove up to 85?±?3 and 55?±?4.5 % of hydrocarbon from refinery sludge when whole bacterial culture and culture supernatant were used, respectively. Maximum surface tension reduction (26.3 mN m^?1) was achieved with the strain in just 24 h of time. Emulsification index (E24) was recorded as 100 and 80 % with crude oil and n-hexadecane, respectively. The biosurfactant was confirmed as rhamnolipid containing C₈ and C₁₀ fatty acid components and having more mono-rhamnolipid congeners than the di-rhamnolipid ones. The biosurfactant was stable up to 121 °C, pH 2–10, and up to a salinity value of 2–10 % w/v. To our knowledge, this is the first report showing the potentiality of a native strain from the northeast region of India for the efficient removal of hydrocarbon from refinery sludge.     

Enhanced biodegradation of endosulfan and its major metabolite endosulfate by a biosurfactant producing bacterium

The present study was carried out to isolate bacteria capable of producing biosurfactant that solublize endosulfan (6,7,8,9,10,10-Hexachloro-1,5,5a,6,9,9a-hexahydro- 6,9-methano-2,4,3-benzodioxathiepine-3-oxide) and for enhanced degradation of endosulfan and its major metabolite endosulfate. The significance of the study is to enhance the bioavailability of soil-bound endosulfan residues as its degradation is limited due to its low solubility. A mixed bacterial culture capable of degrading endosulfan was enriched from pesticide-contaminated soil and was able to degrade about 80% of α-endosulfan and 75% of β-endosulfan in five days. Bacterial isolates were screened for biosurfactant production and endosulfan degradation. Among the isolates screened, four strains produced biosurfactant on endosulfan. ES-47 showed better emulsification of endosulfan and degraded 99% of endosulfan and 94% of endosulfate formed during endosulfan degradation. The strain reduced the surface tension up to 37 dynes/cm. The study reveals that the strain was capable of degrading endosulfan and endosulfate with simultaneous biosurfactant production.     

Effects of Fuel Variables on Diesel Emissions

The body of information presented in this paper is directed towards engineers in the field of environmental sciences involved in measuring and/or evaluating the emissions from a variety of diesel engines or vehicles. This paper summarizes recent data obtained by EPA on identification and quantification of different emissions (i.e. characterization) from a variety of diesel engines.

Extensive work has been done comparing emissions from some light duty diesel and gasoline passenger cars. The work on the diesel vehicles was expanded to include tests with five different diesel fuels to determine how fuel composition affects emissions. This work showed that use of a poorer quality fuel frequently made emissions worse. The investigation of fuel composition continued with a project in which specific fuel parameters were systematically varied to determine their effect on emissions. EPA is presently testing a variety of fuels derived from coal and oil shale to determine their effects on emissions.

EPA has also tested a heavy duty Volvo diesel bus engine designed to run on methanol and diesel fuel, each injected through its own injection system. The use of the dual fuel resulted in a reduction in particulates and NO _x but an increase in HC and CO compared to a baseline Volvo diesel engine running on pure diesel fuel.

Finally, some Ames bioassay tests have been performed on samples from the diesel passenger cars operated on various fuels and blends. An increase in Ames test response (mutagenicity) was seen when the higher aromatic blend was used and also when a commercial cetane improver was used. Samples from the Volvo diesel bus engine fueled with methanol and diesel fuel showed that use of a catalyst increased the Ames response.     

Hydrocarbon degradation, plant colonization and gene expression of alkane degradation genes by endophytic Enterobacter ludwigii strains

The genus Enterobacter comprises a range of beneficial plant-associated bacteria showing plant growth promotion. Enterobacter ludwigii belongs to the Enterobacter cloacae complex and has been reported to include human pathogens but also plant-associated strains with plant beneficial capacities. To assess the role of Enterobacter endophytes in hydrocarbon degradation, plant colonization, abundance and expression of CYP153 genes in different plant compartments, three plant species (Italian ryegrass, birdsfoot trefoil and alfalfa) were grown in sterile soil spiked with 1% diesel and inoculated with three endophytic E. ludwigii strains. Results showed that all strains were capable of hydrocarbon degradation and efficiently colonized the rhizosphere and plant interior. Two strains, ISI10-3 and BRI10-9, showed highest degradation rates of diesel fuel up to 68% and performed best in combination with Italian ryegrass and alfalfa. All strains expressed the CYP153 gene in all plant compartments, indicating an active role in degradation of diesel in association with plants.