Coupling of acetyl coenzyme A (acetyl-CoA) formation and microbial esterification was achieved with various microorganisms and alcohols. Pichia quercuum IFO 0949 and P. heedii IFO 10019 were selected as microorganisms which had strong double coupling activity of acetyl-CoA formation and microbial esterification with alcohol acetyltransferase (AATFase) as a key enzyme in an interface bioreactor. Both the microorganisms also exhibited triple coupling activity of the acetyl-CoA formation, microbial reduction of citronellal to citronellol, and microbial esterification of produced citronellol with acetyl-CoA. A system based on the double coupling activity was applied to the synthesis of various acetate esters. AATFase in some yeasts could catalyze the esterification of primary aliphatic, aromatic, and terpene alcohols, although it could not catalyze esterification of secondary alcohols. It was proved that the double coupling system can be used for the production of acetate esters of primary alcohols without addition of any acetyl donors.