氨基甲酸乙酯水解酶在枯草芽孢杆菌中的表达及发酵优化

将来源于赖氨酸芽孢杆菌SC02的氨基甲酸乙酯水解酶(UH)基因在枯草芽孢杆菌Bacillus subtilis WB600中进行克隆和表达,在枯草芽孢杆菌中实现了UH活性表达,在摇瓶水平通过单因素考察和响应面分析实验对氨基甲酸乙酯水解酶发酵进行优化. 结表明,酶活最高可达到14.20 U/mL,产酶最佳培养基成分为：淀粉10 g/L、磷酸氢二钾9 g/L、麦芽浸膏25 g/L、硫酸镁1 g/L、胰蛋白胨55 g/L,最适发酵温度为37℃,最佳接种量4%. 在3 L发酵罐中采用最优发酵条件,酶活在16 h达到18.03 U/mL.     

响应面法优化纳豆激酶液态发酵条件

目的响应面法优化纳豆激酶液态发酵条件。方法以纳豆芽孢杆菌(Bacillus subtilis Natto)为出发菌株,液态发酵纳豆激酶。在单因素试验确定接种量、温度、初始pH及发酵时间对纳豆激酶活力产生影响的基础上,采用BoxBehnken软件进行响应面优化。结果最终确定纳豆激酶液态发酵的最佳条件为:接种量3%,温度40℃,pH 7.0,发酵时间84 h。在该条件下液态发酵生产纳豆激酶的活力可达749.41 U/ml,与模型预测的酶活力(752.35 U/ml)的相对误差为0.39%。结论成功优化了纳豆激酶液态发酵条件,为纳豆激酶的产业化生产奠定了基础。     

一株枯草芽孢杆菌的分离鉴定及其发酵条件优化

通过水浴处理,反复平板划线,从土壤中分离纯化出一株疑似枯草芽孢杆菌的菌株,命名为WZB。对该菌株进行形态学观察、生理生化实验和16S rDNA序列分析,最终鉴定该菌株为枯草芽孢杆菌。并对该菌株的发酵条件进行了初步优化,优化得到的发酵条件为:接种量3%、发酵温度34℃、初始pH值7.0、培养时间36 h,进一步为枯草芽孢杆菌菌剂开发应用奠定了基础。     

餐厨垃圾微生物发酵生产生物饲料的研究

以含水量60%的餐厨垃圾为原料,研究了对其用微生物（热带假丝酵母Candida tropicalis、啤酒酵母Sac-charomyces cerivisiae、枯草芽孢杆菌Bacillus subtilis）发酵生产生物饲料的工艺条件。确定最佳工艺条件为：餐厨垃圾添加1.5%的尿素,以热带假丝酵母∶啤酒酵母∶枯草芽孢杆菌=1∶1∶1混合菌作为种子液,接种量2%,30℃发酵48h后,50℃烘干。在此条件下所得发酵产物的粗蛋白含量达到28%,比原来增加了4.5%,枯草芽孢杆菌及总酵母菌含量分别在5.2×109 cfu.g-1和8.4×108 cfu.g-1以上,发酵产物有芳香气味,适口性强,具有蛋白饲料和微生态制剂的双重特性。     

D-核糖发酵培养基优化实验研究

对枯草芽孢杆菌(Bacillus subtilis db104)采用同源重组法构建得到的枯草芽孢杆菌转酮酶(tkt)缺失突变菌株B.sptn15-1的传代稳定性进行了检测,证明该工程菌在连续传代培养5代后仍具有92.9%的稳定性;并对该工程菌的最适发酵培养基进行了优化,使工程菌的D-核糖产量从31.70 g·L-1提高到37.44 g ·L-1,提高了18%.     

Surfactin生物表面活性剂及其驱油研究进展

表面活性素（surfactin）是一类由革兰氏阳性的枯草芽孢杆菌产生的脂肽（lipopeptide）型生物表面活性剂,因其具有优于化学合成表面活性剂的若干优点,如低毒性、高生物降解性、更好的环境相容性,且在极端环境下稳定性好,在提高石油采收率方面有较好的应用潜力,但是目前只有少数的生物表面活性剂可以大规模生产实现工业化应用。本文介绍了surfactin生物表面活性剂的化学结构和生物合成机制,并对其发酵生产过程的影响因素进行分析,为提高其生产经济性探索不同的策略,例如使用更便宜的原材料、优化培养基组分、优化反应器等,系统论述了surfactin生物表面活性剂的驱油机理和其与化学合成表面活性剂的复配研究,同时针对其应用时的不足之处提出研究新思路。     

玉米秸秆与豆粕混合发酵饲料工艺研究

研究了以玉米秸秆和豆粕为主要原料制备微生物发酵饲料的操作条件。开展了p H、配料含水量、发酵温度、乳酸菌接种量、枯草芽孢杆菌接种量等不同工艺参数的发酵优化。结果显示:采用p H为5.5、配料含水量为35%、发酵温度40℃、乳酸菌接种量为5‰,枯草芽孢杆菌接种量为1‰的工艺参数进行发酵,效果较好。     

一种去油污微生物洗涤剂的制备及性能研究

采用除油率和去污率为评价指标制备了一种去油污微生物洗涤剂。该洗涤剂包括枯草芽孢杆菌、铜绿假单胞菌和乳杆菌1发酵产生的表面活性剂(最佳条件是:培养基添加菊粉10 g/L或废弃食用油10 g/L;最佳发酵温度是30℃;最佳接种量OD600为0. 2;最佳通气量是0. 44 min(V/V);在发酵3天后加入3%的碳酸轻钙继续发酵2天);脂肪酶和蛋白酶按照等比例添加且为2%。     

纳豆激酶液体发酵条件的优化

以纳豆枯草芽孢杆菌(Bacillus subtilis natto)为出发菌,研究了其液体发酵产生纳豆激酶(Nattokinase,NK)的培养基组成(碳源、氮源、碳氮比和金属离子组成)和培养条件(温度、初始pH值、发酵时间、接种量和装液量)对产酶量的影响.结果表明,液体发酵培养基的最佳碳源为麦芽糖,浓度为1.0%;最...     

生物农药苏云金芽孢杆菌的研究进展

苏云金芽孢杆菌(Bacillus thuringiensis)制剂是目前应用广泛而有效的一种微生物杀虫剂.本文介绍了苏云金芽孢杆菌的菌种优选、发酵过程及剂型研究进展,具体阐述了发酵过程中培养基和发酵条件的优化、各种发酵方式和发酵设备等. 指出了目前发酵生产苏云金芽孢杆菌中存在的问题,提出了解决问题的建议并展望了其发展前景.     

Multifactorial Approach to Biosurfactant Production by Adaptive Strain Candida tropicalis MTCC 230 in the Presence of Hydrocarbons

In this study, Candida tropicalis MTCC 230 was used to adapted in hydrocarbon along with glucose for biosurfactant production, showing diauxic growth during the production. Biosurfactant was characterized through TLC and FTIR analysis as surfactin, a lipopeptide. Process parameters were optimized one factor at a time, showing the highest emulsification index (%E₂₄) at 54 %. The production of biosurfactant was enhanced by using biostatistically based experimental design with the interactive effect of different parameters. On the basis of Placket–Burman design, four factors, hydrocarbon, ammonium chloride, microelements and temperature are found to be significant (P < 0.05) for the production of biosurfactant. A second order polynomial regression model in central composite design estimated the maximum biosurfactant production in terms of the emulsification index (%E₂₄). The optimum combination of different parameters for the biosurfactant production, obtained for hydrocarbon, ammonium chloride, microelements and temperature are 81.41 %, 1.63 (g/l), 1.69 (g/l) and 35.25 °C, respectively. The biosurfactant production was increased twofold after optimization and selection of interactive parameters by response surface methodology.     

Economic production of Bacillus subtilis SPB1 biosurfactant using local agro‐industrial wastes and its application in enhancing solubility of diesel

BACKGROUND: The present work aimed to optimize a new economic medium for lipopeptide biosurfactant production by Bacillus subtilis SPB1 for application in the environmental field as an enhancer of diesel solubility. Statistical experimental designs and response surface methodology were employed to optimize the medium components. RESULTS: A central composite design was applied to increase the production yield and predict the optimal values of the selected factors. An optimal medium, for biosurfactant production of about 4.5 g L^?1, was found to be composed of sesame peel flour (33 g L^?1) and diluted tuna fish cooking residue (40%) with an inoculum size of 0.22. Increased inoculum size (final OD₆₀₀) significantly improved the production yield. The emulsifier produced was demonstrated to be an alternative to chemically synthesized surfactants since it shows high solubilization efficiency towards diesel oil in comparison with SDS and Tween 80. CONCLUSION: Optimization studies led to a strong improvement in production yield. The emulsifier produced, owing its high solubilization capacity and its large tolerance to acidic and alkaline pH values and salinity, shows great potential for use in bioremediation processes to enhance the solubility of hydrophobic compounds. © 2012 Society of Chemical Industry     

Ionic Behavior Assessment of Surface-Active Compounds from Corn Steep Liquor by Exchange Resins

Depending on their ionic nature, biosurfactants can be classified as nonionic, anionic, cationic, or amphoteric. The ionic behavior of biosurfactants is an important characteristic that dictates their use in industrial applications. In this work, a biosurfactant extract obtained from corn steep liquor was subjected to anionic or cationic resins, in order to study the ionic behavior under different operational conditions using response surface methodology. The independent variables included in the study are the dilution of biosurfactant solution, the amount of cationic or anionic resin, and the extraction time, whereas the dependent variables studied consisted of the surface tension of biosurfactant aqueous solution, after contacting with anionic or cationic resin. The results showed that biosurfactant extracted from corn steep liquor is amphoteric, since both resins were able to entrap this biosurfactant, making it particularly suited for use in personal care preparations for sensitive skin.     

Biosurfactant production from n-paraffins by an air isolate Pseudomonas aeruginosa OCD1

The potential production of biosurfactant was investigated with a strain of Pseudomonas aeruginosa OCD(1), which was isolated from air in our laboratory. The degradation of different hydrocarbons was studied with this microorganism. The values of surface tension and emulsification index of culture broth were very promising when n-octadecane was used as substrate. Characterization of biosurfactant revealed that the biosurfactant was rhamnolipid in nature. The surface tension of water was reduced to 31.5 mN/m from 72 mN/m with the critical micelle concentration of 35 mg/L. A low rhamnolipid concentration (< 5 mg/L) had a strong effect on reduction of surface tension.     

Effects of various nutritional supplements on biosurfactant production by a strain of <Emphasis Type="Italic">Bacillus subtilis</Emphasis> at 45°C

Effects of various factors on growth and biosurfactant production by Bacillus subtilis MTCC 2423 were studied. Sucrose (2%) and potassium nitrate (0.3%) were the best carbon and nitrogen sources. The addition of various metal supplements (magnesium, calcium, iron, and trace elements) greatly affected growth and biosurfactant production. The effect of the metal cations, used together, is greater than when they are used individually. The biosurfactant production increased considerably (almost double) by addition of metal supplements. Very high concentrations of metal supplements, however, inhibited biosurfactant production. Amino acids such as aspartic acid, asparagine, glutamic acid, valine, and lysine increased the final yield of biosurfactant by about 60%. The organism could produce biosurfactant at 45°C and within the pH range of 4.5–10.5. The biosurfactant was thermostable and pH stable (from 4.0 to 12.0). The capability of the organism to produce biosurfactant under thermophilic, alkaliphilic, and halophilic conditions makes it a suitable candidate for field applications. Infrared, nuclear magnetic resonance, and mass spectroscopy studies showed the surfactant to be identical to surfactin.     

Production of Glycolipid Biosurfactant from Sponge-Associated Marine Actinobacterium <Emphasis Type="Italic">Brachybacterium paraconglomeratum</Emphasis> MSA21

Potential biosurfactant producers and economic production processes are major considerations for commercialization of biosurfactants. The present study was aimed at exploring marine Actinobacteria for the production of biosurfactants using industrial and agro-industrial wastes under solid state culture (SSC). A biosurfactant producer, Brachybacterium paraconglomeratum MSA21 was isolated from a marine sponge. The strain MSA21 effectively utilized tannery pre-treated effluent as the substrate for the production of a biosurfactant under SSC. The critical control factors influence the production of biosurfactant includes glucose, yeast extract, copper sulfate and inoculum size. The glucose and yeast extract interactively increase the production maxima over a stable area. The surface active compound was characterized as a glycolipid derivative with a hydrophilic part of methyl-2-oxopropyl furan and a hydrophobic dodecanoic acid, methyl ester. The MSA21 biosurfactant displayed antibiotic activity. The domain ketosynthase in MSA21 showed that the polyketide synthase gene might be involved in the synthesis of antimicrobial compounds. The strain B. paraconglomeratum MSA21 could be used for the production of a biosurfactant as a green alternative to replace chemical surfactants.     

Effect of nutrients on optimal production of biosurfactants by <Emphasis Type="Italic">Pseudomonas putida</Emphasis>—A gujarat oil field isolate

Nutritional requirements for maximal production of biosurfactant by an oil field bacterium Pseudomonas putida were determined. The optimal concentrations of nitrogen, phosphate, sulfur, magnesium, iron, potassium, sodium, calcium, and trace elements for maximal production of biosurfactants were ascertained, and a new “Pruthi and Cameotra” salt medium was formulated. Data show that maximal biomass (2.4 g L⁻¹) and biosurfactant production (6.28 g L⁻¹) takes place after 72 h of growth on 2% hexadecane. The biosurfactant was produced optimally over pH and temperature ranges of 6.4–7.2 and 30–40°C, respectively. That the highest biosurfactant yield was obtained during late log phase of growth indicates that the biosurfactant is a secondary metabolite.     

Screening and identification of Pseudomonas aeruginosa AB4 for improved production,characterization and application of a glycolipid biosurfactant using low‐cost agro‐based raw materials

BACKGROUND: The study is focused on (i) screening and taxonomic identity of a bacterial strain for biosurfactant production, and (ii) evaluation of its potential for production of a biosurfactant using agro‐based feedstock(s) and characterization of it for application in the removal of heavy metals. RESULTS: The production of biosurfactant by an isolate Pseudomonas aeruginosa AB4 (identified on the basis of 16S rRNA analysis) using various cost‐effective substrates were examined at conditions 40 °C, 120 rpm for 7 days. It revealed maximum (40 gL^?1) rhamnolipids production and 46% reduction of initial surface tension. Its optimum production was achieved at (i) C:N ratio 10:0.6, (ii) pH 8.5 and (iii) 40 °C. The cell–free supernatant examined for biosurfactant activity by (i) haemolytic assay, (ii) CTAB‐ methylene blue assay, (iii) drop collapse test, (iv) oil spreading technique and (v) EI 24 assay showed its glycolipid nature and stable emulsification. Analysis of partially purified rhamnolipids by (i) thin layer chromatography (TLC), (ii) high performance thin layer chromatography (HPTLC), (iii) high performance liquid chromatography (HPLC), (iv) Fourier transform infrared (FT‐IR) and (v) gas chromatography–mass spectrometry (GC‐MS) confirmed its structure as methyl ester of 3‐hydroxy decanoic acid (a glycolipid) with two major structural congeners (Rha‐C₁₀‐C₁₀ and Rha‐C₁₀‐C₈) of mono‐rhamnolipids. Finally, it showed sequestration of Cd and Pb, suggesting its application in biosurfactant‐assisted heavy metal bioremediation. CONCLUSION: This work has screened and identified a bacterium with superior biosurfactant production capabilities, characterized the glycolipidic biosurfactants as rhamnolipid and indicated the feasibility of biosurfactant production using novel renewable, relatively inexpensive and easily available resources such as non‐edible vegetable de‐oiled seed cakes and showed its utility in remediation of heavy metals. Copyright © 2010 Society of Chemical Industry     

Production and stability studies of the biosurfactant isolated from marine Nocardiopsis sp. B4

A potential biosurfactant producing strain, marine Nocardiopsis B4 was isolated from the West coast of India. Culture conditions involving variations in carbon and nitrogen sources were examined at constant pH, temperature and revolutions per min (rpm), with the aim of increasing productivity in the process. The biosurfactant production was followed by measuring the surface tension, emulsification assay and emulsifying index E24. Enhanced biosurfactant production was carried out using olive oil as the carbon source and phenyl alanine as the nitrogen source. The maximum production of the biosurfactant by Nocardiopsis occurred at a C/N ratio of 2:1 and the optimized bioprocess condition was pH 7.0, temperature 30° C and salt concentration 3%. The production of the biosurfactant was growth dependent. The surface tension was reduced up to 29 mN/m as well as the emulsification index E24 was 80% in 6 to 9 days. Properties of the biosurfactant that was separated by acid precipitation were investigated. The biosurfactant activity was stable at high temperature, a wide range of pH and salt concentrations thus, indicating its application in bioremediation, food, pharmaceutical and cosmetics industries.     

Enhancement of Styrene Adsolubilization and Solubilization by Rhamnolipid Biosurfactant‐Linker Mixtures onto an Aluminum Oxide Surface

The polarity of rhamnolipid, a relatively hydrophilic biosurfactant, can be enhanced by the addition of linker molecules. In this work, rhamnolipid biosurfactant‐modified surfaces were prepared with and without a combination of linkers (1‐butanol, 1‐octanol, and 1‐dodecanol) to investigate effects of linker molecules on styrene adsolubilization and solubilization. Results showed that styrene adsolubilization increased with increasing carbon chain lengths of the linker molecules whereas the solubilization of styrene exhibited the opposite effect. Decreasing the carbon atoms in the linker molecules resulted in higher styrene solubilization capacity because of the change in polarity of the three‐dimensional surfactant aggregates. The higher adsolubilization capacity indicated the enlargement of surfactant tails that was created a larger adsolubilization region in the admicelle while the lesser solubilization of styrene indicated the decreasing of affective area per molecule of the surfactant‐linker system (butanol > octanol > dodecanol).