An efficient and improved synthetic route of fluocinolone acetonide with combination of bio-fermentation was developed from 21-acetyloxy-17 alpha-hydroxy-4,9(11)-diene-3,20-dione (1a). Process of the 6 alpha and 9 alpha fluorination steps was studied, and it was observed that the stereoselectivity of 6 alpha fluorination is highly substrate dependent. After an extensive screening on the fluorinating reagents and activation reagents, 6 alpha-F was introduced in 85% yield with 98.9% stereoselectivity. Instead of HF gas, aqueous HF solution was applied in 9 alpha fluorination step to provide the desired product in 89% yield. Starting from 1a, fluocinolone acetonide was prepared in 9 steps with an overall 38.5% yield.