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为了阐明白腐菌株Phlebia brevispora TMIC34596对有机氯杀虫剂林丹的酶促降解机理及规律,在实验室条件下,通过菌株的纯培养、超声波破碎和高速离心等过程,提取到胞内粗酶液和胞外粗酶液,并研究了胞内及胞外酶对林丹的降解特性、最佳降解条件及动力学参数等。结果表明,胞内酶起主要的降解催化作用,相同处理时间内对林丹的降解率是胞外酶的4~5倍。胞内酶降解林丹的酶促反应最适温度为35℃,最适p H值为5.0,最适条件下反应2 h后的林丹降解率为64.0%。胞内酶在25~40℃、p H值在4.0~6.5时能保持较高的降解活性,对林丹的降解率在50%以上。胞内酶降解林丹的米氏常数Km为1.30μmol/L,最大反应速率Vmax为1.18μmol/min,表明胞内酶对林丹有较强的亲和力,降解林丹速度较快。通过气相色谱-质谱分析,五氯环己醇和四氯环己二醇被鉴定为林丹的胞内酶代谢产物,表明胞内酶可通过连续的脱氯及羟基化作用将林丹转化为多羟基化产物,该途径不同于目前所报道的白腐菌对林丹的降解途径。
In order to elucidate the mechanism and rule of enzymatic degradation of organochlorine insecticide lindane by white-rot fungus Phlebia brevispora TMIC34596, the crude enzyme solution was extracted by pure culture, sonication and high-speed centrifugation under laboratory conditions. And extracellular crude enzyme solution, and studied the degradation characteristics of lindane by intracellular and extracellular enzymes, the optimum degradation conditions and kinetic parameters. The results showed that the intracellular enzymes played a major catalytic role of degradation, degradation rate of lindane 4 to 5 times the extracellular enzyme in the same treatment time. The optimum temperature for enzymatic degradation of lindane by intracellular enzyme degradation was 35 ℃, and the optimum p H value was 5.0. Under the optimum conditions, the degradation rate of lindane was 64.0% after 2 h reaction. Intracellular enzyme at 25 ~ 40 ℃, p H value of 4.0 to 6.5 to maintain a high degradation activity, degradation of lindane in more than 50%. The Michaelis constant Km of intracellular enzyme degradation of lindane was 1.30μmol / L and the maximum reaction rate Vmax was 1.18μmol / min, indicating that the intracellular enzyme had a strong affinity for lindane and the degradation rate of lindane was faster. Pentachlorocyclohexanol and tetrachlorocyclohexanediol were identified as intracellular enzyme metabolites of lindane by gas chromatography-mass spectrometry analysis, indicating that intracellular enzymes can convert lindane to polyhydroxylation by successive dechlorination and hydroxylation Product, this approach is different from the currently reported white rot fungi lindane degradation pathway.