Biotransformation is increasingly exploited as a useful and often unique reaction step in the semi synthesis of pharmaceuticals or for structural modification of complex natural compounds. Estradiol was subjected to biotransformation with B. subtilus and A. niger resulted in the formation of four metabolite which were reported for the first time via B. substillus, whilst the biotransformation of estradiol was carried out with C. elegancs and F. lini also. Inhibitory activity (IC50 (µM) ) of metabolites was carried out against lipoxygenase enzyme. The structures of the compounds were elucidated through extensive spectroscopic analysis including EIMS, HREIMS, 1H NMR, 13C NMR and including advance techniques like NMR 1H-1H COSY, HMBC, and NOESY.