Microbial transformation of cholesterol produces various steroid drugs and intermediates, such as cholest-4-en-3-one (C4EO), which is an important intermediate of hormone steroidal drugs. Cholest-4-ene-3,6-dione (CEDO) is another product of cholesterol biotransformation, with better medicinal value than cholest-4-en-3-one. It is indicated to improve fat metabolism, resistance to tumors and essential in the treatment of nerve injuries. This study proceeded using Burkholderia cepacia ZWS15, which is known to produce enzymes to transform cholesterol. Organic solvents were chosen to increase the solubility of the substrate cholesterol and the conversion of the product, and the organic solvent affects the mechanism of product conversion and cell membrane permeability. The use of organic solvents in this study prompted shifts in the composition of fatty acids in the cell membrane, inducing the production of more unsaturated fatty acids and reduction of saturated fatty acids. The alkenone products generated after transformation of cholesterol were isolated.