Efficient Degradation Of The Oil Actinomycetes Isolation And Degradation Characteristics

With the development of petroleum exploration and industry, petroleum has become one of the most important energy sources for human survival and development. Because of accidents, abnormal operation and overhaul, there will be petroleum hydrocarbon spilling over and emission in the process of petroleum producing, storage, transport and using. Petroleum pollution brings great harm to humans and environment, and it has threatened the survival and development of humans and other organisms. Therefore scientists around the world are greatly concerned about the problem of petroleum pollution. They are looking for a new, safe, economical and reliable technique to solve this problem. With the development of modern biotechnology, microorganism is used more widely in environmental protection. Compared with physical and chemical methods, bioremediation of petroleum pollution has great advantages and it has become a research hot spot.In this study, an effective oil-degrading bacteria was isolated from long-term petroleum contaminated soil and named Z1a-B. Strain Z1a-B was treated with mutagens in order to obtain efficient oil-degrading bacteria. The degradation conditions were studied with the bacteria. The results of this study are as follows:Through enrichment and domestication experiments, an effective oil-degrading bacteria was purified. The petroleum degradation rate reached 66.4% by shaking flask culture.Strain Z1a-B could produce bio surfactants by petro-degrading activity assay and blood plate hemolysis assay. And Strain Z1a-B was tested to be quite strong in hemolyzing and petro-degrading activity, which implied that it has high ability of producing biosurfactant. With further study of the degradability of crude oil, the components of crude oil before and after degradation were analyzed by GC-MS analysis. The results showed that the strain Z1a-B could almost completely degrade C12～C38 n-alkanes. That indicated that the strain has a wide spectrum of degradation and the degradation of oil pollutants can be very good.By analyzing the colony morphology, cultural characteristics and spore morphology, and using the physiological and biochemical tests, strain Z1a-B was preliminarily identified as Albosporus in Streptomyces.Compared with the UV mutagenesis method, UV plus LiCl mutation was a more appropriate mutagenesis one. Using UV plus LiCl, Z1a-B was mutagenized to ZBⅡ-12, whose oil degradation ability was significantly improved. The production capacity of biosurfactants and petro-degrading activity of ZBⅡ-12 increased. ZBⅡ-12 could degrade a wider variety of petroleum hydrocarbons completely than Z1a-B.The optimum nutrient source experiment and growth condition optimizing experiment indicated that the optimum nutrient was KNO3 and K2HPO4, and that the optimum conditions for oil degradation was pH 7.5, temperature 28℃, inoculation 8%, oil concentration of 7 g·L-1. Under pH 6.0～9.0 and temperature 20～42℃, an oil degradation rate of 60% was gained by using ZBⅡ-12.In the indoor orthogonal bioremediation experiment, the primary and second order of factors affecting the oil degradation rate was inoculation> KNO3> K2HPO4> sawdust. The optimal combination was KNO3 3 g·L-1,K2HPO4 0.75 g·L-1,sawdust 30 g·L-1,inoculation 8%. Adding microbial agent, sawdust stalk and a proper amount of nutrients nitrogen and phosphorus in petroleum contaminated soil processing could improve the degradation efficiency of microbe. Plant growth measurement experiment, the results of which were consistent with the detection results of the degradation rate of oil contamination, directly reflected the effects of bioremediation.To culture the strain, solid medium composed of 2 g of bran,40 g of chicken manure, 52.5 g of peat and 1.5 g of quick-lime, or of 2 g of bran,20 g of rice,28 g of soymeal and 1.5 g of quick-lime was optimal. In these two mediums, streptomyces grows quickly and vigorously and yields plenty of spores. A large number of spores, which could be used for bioremediation of petroleum contaminated soil directly, were prepared from cheap raw materials. With the advantage of higher efficiency and convenient storage and transport, bioremediation of petroleum contaminated soil could be promoted and applied on a large scale using this method.