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脂肽类生物表面活性剂的发酵优化及其性质和应用研究

Production, Characterization and Application of Lipopeptide Biosurfactant

【作者】 彭丽丽

【导师】 王菊芳; Li Shuang(李爽);

【作者基本信息】 华南理工大学 , 发酵工程, 2011, 硕士

【摘要】 生物表面活性剂主要是由微生物产生的天然表面活性化合物,可被分为六大类:糖脂类、脂肽类、磷脂类、脂多糖类、脂肪酸、中性脂以及多糖蛋白复合物等。和化学合成表面活性剂一样,具有降低表面张力、稳定乳化液等功能。除此之外,由于其具有生物可降解性;低毒或无毒,可用于医药、化妆品和食品领域;在极端温度、pH、盐浓度下的耐受性和选择性;具有更高的起泡性等;结构功能多样性,在特殊领域具有一定的更好的可适用性。由于这些显著特点,生物表面活性剂的研究日益增多,应用也迅速扩展至环保、石油开采、制药、食品、土壤修复等领域。本研究以中国科学院南海海洋研究所提供的一株生物表面活性剂产生菌为研究对象,分别从菌株鉴定、发酵条件和产物特性等方面进行了研究,主要结论如下:1.经菌落、菌体形态和16S rDNA序列分析,鉴定为芽孢杆菌属,命名为Bacillus SCUT09。该菌株以葡萄糖为碳源时培养基表面张力可降至46.2 mN/m,对煤油的乳化能力达到34%。2. Bacillus SCUT09的培养基优化研究结果表明,最利于Bacillus SCUT09生长和生物表面活性剂积累的培养条件为:木薯淀粉20 g/L,牛肉膏7 g/L,NaCl 10 g/L,MgSO4·7H2O 0.7 g/L,Na2HPO4·12H2O 3.8 g/L,酵母提取物0.5 g/L,28oC,初始pH值7.0,接种量2%,转速为200 rpm。优化后以木薯淀粉为碳源时,培养基的表面张力达到28 mN/m,对煤油的乳化能力达到57%。发酵40 h后,菌体生长进入稳定期,菌体干重达1.6 g/L,表面活性剂产量约为0.8 g/L。3.产物TLC和FTIR分析结果表明,该生物表面活性剂为脂肽类物质,其临界胶束浓度为0.2 g/L;对盐浓度的耐受性为21% NaCl。在pH4-11范围内性质变化不大。样品经121 oC高温处理2 h后的表面活性和乳化能力保持稳定。4.经重力沉降法分析,脂肽表面活性剂可以作为纳米材料的分散剂。同时利用该脂肽物质经水热法合成的纳米ZnO,经扫描电镜和电子显微镜分析,和利用CTAB合成的纳米ZnO相比,具有更小的粒径和较好的分散稳定性。

【Abstract】 Biosurfactants synthesized by microorganisms are one of natural surface-active compounds produced by microorganisms, which could be classified into six groups. Basically there are six major classes of biosurfactants:, glycolipids, lipopeptide, phospholipids, lipopolysaccharides, substituted fatty acids and neutral lipids. Like synthetic surfactants, biosurfactants can reduce the surface tensions of aqueous media. However, biosurfacants have several outstanding advantages, such as higher biodegradability, lower toxicity, higher selectivity and specific activity at extreme temperatures, pH, and salinity, and they also have higher foaming, better environmental compatibility and the ability to be synthesized from renewable feed stocks and so on.A biosurfactant-producing bacterium provided by South China Sea Institute of Oceanology was studied. The detailed conclusions were as follows:(1). The strain was classified into Bacillus sp. according to its 16S rDNA sequences and designated as Bacillus SCUT09. The surface tension is lowered to 46.2 mN/m and the emulsification in kerosene-water is 34% when glucose was used as carbon source.(2). The culture conditions of the strain were primarily optimized. The best carbon and nitrogen source are the cassava starch and beef extract, respectively. Under the conditions of 1% NaCl, pH 6.5 and 28 oC, the biomass and emulsifying ability reach the maximum values.(3). Fourier transform infrared spectroscopy and thin layer chromatography analysis revealed that SCUT09 produced lipopeptide biosurfactant in the process of metabolism. Its critical micelle concentration (CMC) was 0.2 g/L and its salt tolerance was 21% of NaCl. Also, the biosurfactant has higher stability in the pH range from 4 to 11. Also, the surface tension and the emulsifying ability remained steady after 2 h incubation at 121oC.(4). Gravity sedimentation analysis results showed that lipopeptide prepared in this study was a good kind of nano-materials dispersant. Compared to CTAB, the size of nano-ZnO particles using lipopeptide prepared in this study as dispersant is smaller by scanning electron microscopy and electron microscopy analysis.

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