高效生物表面活性剂产生菌筛选及其性质研究

目的:获得产高效生物表面活性剂的菌株并获得优化培养基。方法:通过从山东文登某加油站附近长期污染富含油质的土壤中逐步采用富集培养基和平板筛选培养基分离筛选菌株并进行优化培养寻找最优生长培养和高产生物表面活性剂的条件。结果:筛选出产表面活性剂的微生物12株,分别命名为BSF1#-BSF12#,从中筛选出1株高效表面活性剂产生菌BSF8#,优化培养结果表明BSF8#的最佳生长pH在7.5左右,最佳碳源为葡萄糖,最佳氮源为蛋白胨,BSF8#培养基中最佳NaCl浓度为2g/L。BSF8#菌株可将发酵液的表面张力由最初的48.29mN/m降到27.79 mN/m,上层乳化状发酵液的排油圈最大直径超过7.5cm,并经红外光谱分析确定其生物表面活性剂为1个糖肽类化合物。结论:BSF8#菌株产生的生物表面活性剂活性突出,有较大的开发潜力。     

一株石油烃降解菌的细胞疏水性及其乳化性质

【目的】从新疆油田石油污染土壤中分离到一株在25 °C条件下利用烃类产生生物表面活性剂的菌株红球菌(Rhodococcus sp.) HL-6, 对其菌体细胞疏水性及所产表面活性剂进行研究。【方法】通过细胞粘附性、表面张力及乳化活性测定对菌株所产表面活性剂进行性质研究。【结果】菌株HL-6在亲水性和疏水性基质中均能产生生物表面活性剂, 在疏水性基质中可以将培养液表面张力由初始的62.487 mN/m降到30.667 mN/m, 培养液在pH 6?9及NaCl浓度1%?5%范围内乳化效果良好, 在4 °C到55 °C范围内乳化效果均为100%, 菌株对柴油的耐受能力很高, 在30%柴油浓度下依然生长良好并且有44%的乳化活性。【结论】HL-6菌株的细胞表面具有很强的疏水性, 这有助于菌体细胞对烃类的摄取。该菌株能够利用烃类基质生产生物表面活性剂, 可以明显降低培养液表面张力并且对石油烃具有良好的乳化作用。说明菌株HL-6能够适应海洋滩涂石油污染的环境, 并可用于严重石油污染区域的生物修复。     

烟碱降解菌以烷烃为碳源产生物表面活性剂的研究

通过测定发酵过程中菌体浓度和发酵上清液的表面张力,研究了烷烃碳源和发酵条件对烟碱降解菌(Ochrobactrum sp.)产生物表面活性剂的影响。结果表明,菌株Ochrobactrum sp.以十三烷和十六烷为碳源生长较好,而利用液体石蜡可产生较多生物表面活性剂。以2%液体石蜡为碳源,装液量为40%(250 m L三角瓶),于30℃,120 r/min培养4 d时,发酵液表面张力能降低至42.1m N/m。     

一株产脂肽类表面活性剂的碱性Dietzia菌及特性研究

【目的】筛选降解性能良好的产生物表面活性剂的菌株,对其进行分类学鉴定,确定所产表面活性剂物质并对各影响因素进行评价。【方法】利用液体石蜡为底物筛选降解性能良好的产生物表面活性剂菌株,通过形态特征观察、生理生化测定、16S rRNA基因序列分析等实验确定菌株的分类地位。通过排油圈活性、表面张力值、薄层层析等方法确定生物表面活性剂的性质,分析碳、氮源和温度、pH、盐浓度各因素对菌株产生物表面活性剂的影响。【结果】从大连新港采集的样品中分离得到一株产表面活性剂的嗜碱菌株3372,经分类鉴定表明其是Dietzia cercidiphylli的新菌株。嗜碱菌3372发酵液粗提物的排油直径为6.1 cm,表面张力可从67.62 mN/m降到32.95 mN/m,经薄层层析分析,初步鉴定为脂肽类表面活性剂。综合各因素对发酵液表面活性的影响,菌株3372在pH为9.0、适盐浓度为3%的培养基中,经30°C培养可将发酵液表面张力值降到最低。【结论】嗜碱菌3372是脂肽类生物表面活性剂产生菌的新成员,其在高盐碱条件下产生表面活性剂的特性在工业应用上有一定的潜力。     

一株分离自海水的产生物表面活性剂菌株的鉴定及其发酵优化

为研究分离自海水的芽孢杆菌dhs-330产生物表面活性剂的培养条件和产物特性,采用16S r DNA基因序列分析鉴定菌种,对发酵培养基的碳源、氮源、p H值和培养温度进行优化,采用飞行时间质谱进行产物鉴定及抑菌圈法考察产物抑菌活性。结果显示,该菌株与Bacillus mojavensis菌株16S r DNA的序列相似性为99%。菌株发酵液表面张力可由70 m N·m-1降低至27 m N·m-1。发酵培养基的最适碳源、有机氮源和无机氮源分别为甘油、酵母膏和尿素;p H值为6.5~7.0、温度为30~35℃条件下,菌体生长和生物表面活性剂合成最为有利。发酵产物为脂肽-糖脂混合型生物表面活性剂,对海洋污损微生物Bacillus pumilus dhs04有显著的抑菌活性。菌株dhs-330是能合成脂肽-糖脂混合生物表面活性剂、具有海洋污损微生物防除潜力的优选菌株。     

一株糖脂表面活性剂产生菌的筛选及干酪根降解

【目的】从油页岩环境中筛选可降解油页岩干酪根的产生物表面活性剂菌株。【方法】从抚顺油页岩矿废水样品中用血平板法初筛,排油圈法、乳化法和表面张力法复筛,获得产生物表面活性剂菌株。对目标菌株进行生理生化鉴定、16S r RNA基因序列和系统发育分析,用薄层色谱鉴定其发酵液表面活性成分,优化产表面活性剂的培养条件,初步考察其对油页岩干酪根的降解能力。【结果】筛选到一株产糖脂表面活性剂菌株B-1,初步鉴定为Pseudomonas sp.,该菌株有良好的排油和乳化能力以及较低的表面张力,可利用烷烃、不饱和脂肪酸和糖类作为碳源。在30-34°C范围内添加0.3%Na Cl的葡萄糖培养基(p H 7.0)中该菌生长旺盛,发酵液表面张力最低为27 m N/m。菌株B-1在添加一定量葡萄糖的无机盐培养基中作用30 d后对干酪根的降解率为2.85%,高于不添加葡萄糖无机盐培养基对照组的降解率(1.04%)。【结论】菌株B-1是一株性能良好的产糖脂表面活性剂细菌,有降解干酪根的潜力。     

产生物表面活性剂芽孢杆菌培养条件的优化

为了提高生物表面活性剂的表面活性,通过单因素及正交试验对已筛选的产生物表面活性剂芽孢杆菌的培养基及培养条件进行了优化,优化后的培养基成分为可溶性淀粉20 g/L,氯化铵2 g/L,KH2PO46 g/L,K2HPO42 g/L,MgSO4.7H2O 0.3 g/L,NaCl 2 g/L,CaCl20.08 g/L,EDTA 0.4 g/L。培养条件为4%接种量,种龄16 h,初始pH7,培养温度37℃,摇床转速160 r/min,发酵48 h。优化发酵条件后,发酵液表面张力由初始67.5 mN/m降低至24.8 mN/m,生物表面活性剂产量达到1.08 g/L。     

一株产生物表面活性剂低温细菌的筛选与鉴定

采用血平板培养基及蓝色凝胶平板培养基初筛、排油圈法复筛,从低温环境下自然腐烂秸秆中分离筛选到4株产生物表面活性剂的低温细菌.其中菌株B-17发酵液排油圈达到最大,在5d内可使发酵液的表面张力由75.47 mN· m-1降至37.49 mN·m-1.通过形态特征、生理生化试验及16S rDNA序列分析,初步鉴定该菌为理研菌属(Petrimonas sp.).红外光谱分析表明,菌株B-17在代谢过程中能产生糖脂类表面活性物质.该菌发酵液的乳化能力在5d内仍能保持在75％,具有很好的增溶效果.研究表明,初始pH 7、盐浓度0.4％、温度20℃时,对菌株B-17生长和产生物表面活性剂最有利.本研究为低温环境下产生物表面活性剂细菌的开发奠定了基础.     

一株产表面活性剂细菌鉴定及发酵条件优化

本文利用16S rDNA序列并结合形态学特征,鉴定实验室前期从油污土壤中分离的产表面活性剂菌株1098-3为大肠埃希氏菌(Escherichia coli).通过单因子实验初步确定了其产生表面活性剂的最适条件:37℃,初始pH7.0,转速200 r/min,培养基配比为可溶性淀粉2.5％,胰蛋白胨1.5％,氯化钠0.3％,磷酸二氢钾0.5％,氯化钙0.004％,硫酸铵0.6％,硫酸镁0.07％,酵母粉0.06％,500 mL三角瓶装液量为200 mL.在此条件下,发酵液表面张力由69.3 mN/m降至34.3mN/m,此时发酵液中表面活性剂相对浓度(RBC)为500.     

产生物表面活性剂的地衣芽孢杆菌种子培养条件研究

本文对产生物表面活性剂的油藏地衣芽孢杆菌种子培养条件进行优化.通过测定种子培养液中菌体浓度和发酵液的菌浓及表面张力,研究温度、通气量、接种龄、接种量对种子生长和发酵产生物表面活性剂的影响.确定了种子适宜培养条件为装液量100 mL/250 mL,12层纱布封口,于50 ℃、180 r/min摇床培养14 h.以10％接种量接种发酵,发酵24 h的发酵液表面张力降至最低,为22.6 mN/m.在该条件下培养种子,可缩短种子培养时间,实现提前接种发酵并高产生物表面活性剂.     

无患子水提皂素液发酵精制联产生物表面活性剂槐糖脂

无患子水提皂素液,经纤维二糖酶水解,以无患子水提水解液为底物,接种丘陵假丝酵母,将水提液中糖组分发酵转化为槐糖脂,得到天然皂素及生物表面活性剂复合产物。在发酵过程中,2%的丘陵假丝酵母菌种接种量,溶液中葡萄糖消耗速率最快;在水提水解液中额外添加大豆油作为补充碳源能较大幅度降低溶液表面张力。经过发酵转化,溶液中表面活性物质浓度达到52.48 g/L,比发酵前提高了23.4%,溶液表面张力值明显降低。无患子精制发酵液中不含糖类成分,是理想的液体洗涤剂生产原料。     

Co-Utilization of Canola Oil and Glucose on the Production of a Surfactant by <Emphasis Type="Italic">Candida lipolytica</Emphasis>

Candida lipolytica synthesized a surfactant in a cultivation medium supplemented with canola oil and glucose as carbon sources. Measurements of biosurfactant production and surface tension indicated that the biosurfactant was produced at 48 h of fermentation. The surface-active species is constituted by the protein–lipid–polysaccharide complex in nature. The cell-free broth was particularly influenced by the addition of salt, the pH and temperature depending on the emulsified substrate (hexadecane or a vegetable oil). After comparison between ethyl acetate and mixtures of chloroform and methanol as solvent systems for surfactant recovery, it was found that ethyl acetate was able to extract crude surfactant material with high product recovery (8.0 g/L). The isolated biosurfactant decreased the surface tension to values of 30 mN/m at the critical micelle concentration. Emulsification properties of the biosurfactant produced were compared to those of commercial emulsifiers and other microbial surfactants.     

Enhancement of stability of biosurfactant produced by <Emphasis Type="Italic">Candida lipolytica</Emphasis> using industrial residue as substrate

This work describes experimental results carried out on the fermentation of Candida lipolytica, which produced a new biosurfactant when grown on a vegetable oil refinery residue as substrate. The cell-free culture broth containing the biosurfactant formed stable emulsions with hydrophobic natural compounds. Emulsification properties of the biosurfactant were not affected by salinity; however, treatment at a higher temperature decreased the emulsification activity, indicating applications in oil recovery. The isolated biosurfactant corresponds to a yield of 4.5 g/l, and the surface tension of water was reduced from 71 to 32 mN/m. Preliminary chemical characterizations showed that the biosurfactant consisted of protein (50%), lipid (20%), and carbohydrate (8%).     

高产生物表面活性剂菌株的紫外诱变选育

从新疆塔里木河边的土壤中筛选出1株产表面活性剂的菌株BIT-TLM1,该菌在以葡萄糖为碳源的无机盐培养基中,在150 r/min、40℃的条件下培养20 h,发酵液的表、界面张力分别降至29.29和0.61 mN/m。以菌株BIT-TLM1为原始菌株,对该菌进行紫外线诱变,选育出1株高产表面活性剂的诱变菌株UV-10,其产生表面活性剂的量达到0.309 7 g/g菌体,比原始菌株提高了9.22%,UV-10有较好的遗传稳定性,UV-10产表面活性剂的最佳培养条件:碳源为葡萄糖、氮源为NH4Cl、pH8.0和2%的盐度。     

废弃食用油脂生物合成鼠李糖脂研究进展

碳源的成本过高限制了鼠李糖脂的工业化生产及应用,废弃食用油脂作为一种廉价易得的碳源,越来越多的研究者开始关注用它发酵生产鼠李糖脂.废弃食用油脂的种类、投加量对鼠李糖脂的产量、结构、性质均会产生影响,目前研究中用废弃食用油脂作碳源,鼠李糖脂产量最高可达24.61g/L、表面张力最低达到24mN/m、产物CMC最低可达40.19mg/L.此外,本文还总结了菌株、氮源、微量元素、pH、溶氧及培养方式等因素对废弃食用油脂生产鼠李糖脂的影响,并展望了利用废弃食用油脂生产鼠李糖脂实现产业化的重点研究方向.     

Nutritional requirements and growth characteristics of a biosurfactant-producingRhodococcus bacterium

The nutritional requirements and growth characteristics of a biosurfactant-producingRhodococcus bacterium isolated from Kuwaiti soil were determined. Maximum cell yields (6.6 g/l) and biosurfactant production were achieved with a medium containing 2% (v/v)n-paraffin as a carbon and energy source, 0.2% lactose broth, optimal concentrations of nitrogen (nitrate), phosphorus, iron, magnesium and sodium sources, and minimal concentrations of potassium and trace element sources. The optimal pH was 6.8 for surfactant production and optimal temperature was 37°C. The biosurfactant produced after 16 to 33 h growth in a 7 I fermenter decreased both surface tension and interfacial tension of culture broth to below 27 and 1.8 mN/m, respectively, and was effective at critical micelle dilutions of 10^–3. Data on biosurfactant biosynthesis suggest that the product is produced as a primary metabolite and, therefore, could be produced effectively under continuous fermentation conditions.A.S. Abu-Ruwaida, S. Haditirto and A. Khamis are with the Kuwait Institute for Scientific Research, Biotechnology Department, P.O. Box 24885, 13109, Safat, Kuwait. I.M. Banat is now in Londonderry, Northern Ireland but was at the Kuwait Institute for Scientific Research at the time this paper was written. A.S. Abu-Ruwaida is the corresponding author.In view of the annexation of Kuwait by Iraq in August 1990, this paper has been accepted without return to the author for attention to minor details. The Editor-in-Chief therefore assumes full responsibility for any errors or omissions.     

Production of biosurfactant using different hydrocarbons by Pseudomonas aeruginosa EBN-8 mutant

The present investigation dealt with the use of previously isolated and studied gamma-ray mutant strain Pseudomonas aeruginosa EBN-8 for the production of biosurfactant by using different hydrocarbon substrates viz. n-hexadecane, paraffin oil and kerosene oil, provided in minimal medium, as the sole carbon and energy sources. The batch experiments were conducted in 250 mL Erlenmeyer flasks, containing 50 mL minimal salt media supplemented with 1% (w/v) hydrocarbon substrate, inoculated by EBN-8 and incubated at 37 degrees C and 100 rpm in an orbital shaker. The sampling was done on 24 h basis for 10 d. The surface tension of cell-free culture broth decreased from 53 to 29 mN/m after 3 and 4 d of incubation when the carbon sources were paraffin oil and n-hexadecane, respectively. The largest reduction in interfacial tension from 26 to 0.4 mN/m was observed with n-hexadecane, while critical micelle dilution was obtained as 50 x CMC for paraffin oil as carbon source. When grown on n-hexadecane and paraffin oil, the EBN-8 mutant strain gave 4.1 and 6.3 g of the rhamnolipids/L, respectively. These surface-active substances subsequently allowed the hydrocarbon substrates to disperse readily as emulsion in aqueous phase.     

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.