Screening For Producing Cholesterol Oxidase(COD) Strains And Research Of COD Properties

The function of cholesterol oxidase (COD) in food, medicine and biologic resisting insects is now studied in many aspects and show huge potentials. In this paper, four topics were investigated:(1) screening strains producing COD and identification by physiological and biochemical tests;(2) mutagism of high-yield COD strains;(3) purifying crude enzyme by (NH4)2SO4precipitat;(4) study of COD properties.Samples, which came from fermenting mixtures of eggs, squids and soil for2months, were placed by accumulation culture and plate cultivation in only cholesterol carbon source. It obtained45strains. After colorimetry, it received four high COD activity strains, among which No.510is the highest one, I.e135.45U/L. By physiological and biochemical tests, it belonged to Rhodotorula.It used many mutagenesis methods (UV, ultrasound, NaN3and combined mutation) to do induction experiments. It obtained M-24high COD activity strains, I.e445U/L, enhanced by231.87%. Especially, combined mutation has better effect on producing COD strains.No.M-24mutant strains were fermened and sagged to purify crude enzyme by distinct density (NH4)2SO4contained1.854mg/mL protein. After extracting, enzyme liquid was condenced twice, but activity from0.393mg/mL to0.372mg/mL, and recovery rate66.34%. Recovery rate of COD is not high.It studied properties of crude COD which has widely pH stability and strong thermal stability. The optimum pH is6and optimum temperature is50℃. It also studied several metal ions and compound effects on COD activity during low50℃and pH6. Most metal ions had little effects on COD activity, but Cu2+and boric acid had positive effect on COD activity while Cr6+, SDS and EDTA had passive effect on it.