RESPONSE SURFACE METHODOLOGY BASED MEDIUM OPTIMIZATION FOR IMPROVEMENT OF CHOLESTEROL OXIDASE ENZYME PRODUCTION BY NEWLY ISOLATED STREPTOMYCES SP.

Cholesterol oxidase is an important enzyme with great commercial value in many clinical and industrial processes, such as determination of cholesterol levels in food, serum and other clinical samples as well as its potential application, as a biocatalyst  for bioconversion of sterol compounds and as an insecticide. A Preliminary comparative study of cholesterol oxidase enzyme production from Streptomyces sp. strain NHIA_CH5 using six different media containing cholesterol in submerged fermentation was investigated. An increase in the production was detected after two days in all of the studied media with a maximum production of 23.7 U/mL, noticed in medium containing  soluble starch, glucose, peptone, beef extract, MgSO4.7H2O, and NaCl. A classical (one-variable-at-a-time) method revealed that the elimination of MgSO4.7H2O   and NaCl favored cholesterol oxidase enzyme production. Statistical optimization of the other components was studied using the screening method Plackett-Burman design followed by the modeling method of the central composite design (CCD). The results showed that beef extract and glucose are the variables that significantly influenced and considered as the most important factors for cholesterol oxidase production. Experiments of CCD based on response surface methodology determine the optimum concentrations of these significant medium components and revealed maximum cholesterol oxidase enzyme production (50.5 U/mL) at beef extract 55 g/L and glucose 40 g/L which represented 2- fold higher titers than acquired from the non-optimized medium.