A new lipase which enantioselectively hydrolyzes (+/-)-trans-3-(4-methoxyphenyl)glycidic acid methyl ester [(+/-)-MPGM], a key intermediate in the synthesis of diltiazem hydrochloride, was purified from the culture supernatant of Serratia marcescens Sr41 8000. The apparent kinetic constants (K(m), V(max)) for hydrolysis of (2S,3R)-MPGM [(+)-MPGM] were 350 mM and 1.7 x 10(-3) mol/min/mg protein in a toluene-water (1 : 1) emulsion system. The lipase did not attack (2R,3S)-MPGM [(-)-MPGM], and (-)-MPGM acted as a competitive inhibitor. The molecular mass was estimated to be 62,000+/-2,000 from SDS-PAGE. The lipase preferentially hydrolyzed (2S,3R)-3-phenylglycidic acid esters, but did not hydrolyze cinnamic acid esters. The lipase released glycerol and fatty acid from olive oil, and the optimum pH and temperature for hydrolysis of olive oil were pH 8 and 45-degrees-C, respectively. The lipase was inhibited by Co2+, Ni2+, Fe2+, Fe3+ and EDTA, and activated by Ca2+, Li+ and SDS. It was presumed that the lipase was a metalloenzyme containing approximately one gram atom of calcium per molecular mass of 62,000. The lipase selectively hydrolyzed the 1,3 ester of triglycerides. Sequencing of the N-terminal amino acids revealed that this lipase was distinct from other known lipases.