This paper concentrates on studies of the acylation of alcohols with 3,3-difluoro-4-phenylazetidin-2-one rac-1, trans-3-fluoro-4-phenylazetidin-2-one rac-2 and 4-phenylazetidin-2-one rac-3 in the presence of immobilized lipase PS from Burkholderia cepacia in dry tert-butyl methyl ether (TBME). Fluorine activation in the compounds studied was essential in order to split the β-lactam ring with lipase PS. The highly enantioselective ring opening of rac-1 and rac-2 with methanol (1-butanol was also studied) allowed the preparation of the (R/(3R,4R))-β-lactams as the unreacted enantiomers and (S/(2S,3S))-β-amino esters as the product enantiomers with an ee >99%. Under the same conditions, rac-3 was totally unreactive. The possibility for a competing hydrolysis caused by water in the enzyme preparations is also discussed.