假单胞菌YZ32产生物乳化剂及原油降解作用

Bioemulsifier Production and Crude Oil Degradation by Pseudomonas YZ32

苟 燕 佘跃惠 舒福昌 董 浩

GOU Yan SHE Yuehui SHU Fuchang DONG Hao

为了分离出一株产生物乳化剂且能降解原油的菌株，以原油为唯一碳源，筛选出能乳化原油的细菌，再通过调整培养基成分来优化假单胞菌YZ32 产生物乳化剂，以乳化指数和乳化剂产量为指标选出最佳的碳源和氮源；用乙醇提取生物乳化剂，再用透析袋纯化，测定乳化剂的糖含量和蛋白质含量，检测乳化剂的乳化活性物质。将乳化剂与7种不同的底物混合后测定乳化指数，考察乳化剂对不同底物的乳化效果。在不同条件下考察生物乳化剂的理化稳定性；通过原油降解实验，用 GC-MS 对原油组分进行分析，考察菌株对原油的去除率和降解效果。结果表明，分离出的能乳化原油的假单胞菌YZ32，以蔗糖（20 g/L）和谷氨酸钠（7 g/L）为碳源和氮源时，可获得最大的生物乳化剂产量。代谢产物中蛋白含量为 64.29%，糖含量为 13.89%，乳化活性物质为蛋白类物质。该乳化剂对液体石蜡和甲苯具有较好的乳化效果，在温度为 15～80 ℃、pH值为4～11和矿化度为 50～350 g/L 的条件下，YZ32产生的生物乳化剂都能保持良好的稳定性。原油降解实验表明，YZ32在24 d内对原油的去除率达到60.3%，对原油饱和烃中的C11—C20组分和芳香烃中的菲类具有较好的降解效果。假单胞菌YZ32 能产生蛋白类的且稳定性良好的生物乳化剂，对原油具有良好的降解效果，在含油污泥土壤修复中具有巨大的应用潜力。

In order to obtain a strain that could produce bioemulsifier and degrade crude oil，the bacteria that could emulsify crude oil was selected using crude oil as the sole carbon source，and the production of bioemulsifiers by Pseudomonas YZ32 was optimized by adjusting the composition of the culture medium. The optimal carbon and nitrogen sources were selected based on the emulsification index and emulsifier yield. bioemulsifiers was extracted with ethanol and purified with dialysis bags，the sugar and protein content of the emulsifiers was measured，the emulsifying active substances of the emulsifiers was tested. By Mixxing bioemulsifiers with 7 kinds of different substrates and measuring the emulsification index，the bioemulsification effect of emulsifiers on different substrates was investigated. Under different conditions，the physicochemical stability of bioemulsifiers was investigated. The crude oil degradation experiments were carried out，the components of crude oil were analyzed using GC-MS to investigate the removal rate and degradation effect of strains on crude oil. The results showed that a strain of Pseudomonas YZ32 capable of emulsifying crude oil was isolated. The maximum bioemulsifier yield could be obtained using 20 g/L of sucrose and 7 g/L of sodium glutamate as carbon and nitrogen sources for YZ32. The protein content of its metabolites was 64.29% and the sugar content was 13.89%，and the emulsifying active substances were protein-like substances. The bioemulsifier had a good emulsifying effect on liquid paraffin and toluene. The bioemulsifier produced by YZ32 could maintain good stability under the temperatures of 15—80 ℃，pH value of 4—11 and salinity of 50—350 g/L. The crude oil degradation experiment showed that YZ32 achieved 60.3% removal of crude oil within 24 d with better degradation effect for C11—C20 components in saturated hydrocarbons and phenanthrene in aromatic hydrocarbons of crude oil. Pseudomonas YZ32 could produce stable protein based bioemulsifiers. YZ32 had a good degradation effect on crude oil and had great potential for application in the remediation of oily sludge soil.

10.19346/j.cnki.1000-4092.2024.01.020