الفهرس 
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Abstract
There is an increasing demand on cholesterol oxidase for its various industrial and clinical applications. Streptomyces aegyptia strain NEAE-102 exhibited high level of extracellular cholesterol oxidase production using a minimum medium containing cholesterol as the sole source of carbon. Fifteen variables were screened using Plackett– Burman design for the enhanced cholesterol oxidase production. The most significant variables affecting enzyme production were further optimized by using the face centered central composite design. The statistical optimization resulted in an overall 4.97-fold increase (15.631 UmL−1) in cholesterol oxidase production in the optimized medium as compared with the unoptimized medium before applying Plackett Burman design (3.1 UmL−1).The crude enzyme extract was purified by two purification steps, protein precipitation using ammonium sulfate followed by ion exchange chromatography using DEAE Sepharose CL-6B. The best conditions for maximum cholesterol oxidase activity were found to be 105 min of incubation time, an initial pH of 7 and temperature of 37 °C. The optimum substrate concentration was found to be 0.4 mM. The higher thermal stability behavior of cholesterol oxidase was at 50 °C. The apparent molecular weight of the purified enzyme as sized by sodium dodecyl sulphate-polyacryalamide gel electrophoresis was approximately 46 KDa. The amino acid contents of the purified Streptomyces aegyptia NEAE-102 was determined using amino acid analyzer. Glutamic acid was the most represented amino acid in the quantified cholesterol oxidase (11.49 µg/mL). The purified cholesterol oxidase was evaluated for its in vitro anticancer activities against five human cancer cell lines. The selectivity index values on rhabdomyosarcoma and breast cancer cell lines were 3.26 and 2.56; respectively. The in vivo anticancer activity of cholesterol oxidase was evaluated against Ehrlich solid tumor model. Compared with control mice, tumors growth was significantly inhibited in the mice injected with cholesterol oxidase alone, doxorubicin alone and cholesterol oxidase/doxorubicin combination by 60.97%, 72.99% and 97.04%; respectively. These results demonstrated that cholesterol oxidase can be used as a promising natural anticancer drug.