The activity of dehydropeptidase I in rat tissues decreases in the order of lung > kidney > liver-spleen > other tissues, while aminopeptidase activity is high in the kidney, and lower in the lung than in other tissues.Dehydropeptidase I was solubilized from the membrane fraction of rat lung by treatment with papain and purified by DEAE-cellulose column chromatography, affinity chromatography on concanavalin-A-Sepharose and highperformance liquid chromatography gel filtration. The purified preparation was found to be homogeneous on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The relative molecular mass was estimated to be 150000 by gel filtration, comprising a homodimer of two 80000-Mr subunits. The enzyme activity was inhibited by cilastatin, o-phenanthroline and ATP. This enzyme catalyzed the hydrolysis of S(substituent)-L-cysteinyl-glycine adducts such as L-cystinyLbis(g1ycine) and N-ethylmaleimide-S-L-cysteinyl-glycine, as well as the conversion of leukotriene D4 to F4. Furthermore it catalyzed a hydrolytic splitting of L-Leu-L-Leu, but not S-benzyl-L-cysteine p-nitroanilide, which is a good substrate for aminopeptidase. Our enzyme preparation was immunologically identical to the rat renal dehydropeptidase I. The physiological significance of the pulmonary dehydropeptidase I on the metabolism of glutathione and its adducts is discussed.The degradation of reduced and oxidized glutathione and of glutathione adducts is initiated by y-glutamyl transferase [l , 21, which removes the y-glutamyl moiety to form L-cysteinylglycine(L-Cys-Gly), L-cystinyl-bis(g1ycine) (~-m y s -G l y J and S (substituent)-L-Cys-Gly adducts, respectively. These dipeptides or dipeptide adducts are further hydrolyzed by dipeptidases or by aminopeptidase M [3,4].Campbell et al. purified the dipeptidase hydrolyzing glycyl-dehydrophenylalanine from porcine [S -71 and human [8] kidneys. Recently, it has been reported that this enzyme specifically hydrolyzes the dehydropeptide bond in carbapenem antibiotics, and it has been referred to as dehydrope tiddse I (DHP-I) [9, 101. We have previously shown that L-h ys ys-Gly, and S(substituent)-L-Cys-Gly adducts are mainly hydrolyzed by the renal DHP-I rather than by aminopeptidase M, and we suggested that DHP-I is an important enzyme in the metabolism of glutathione and glutathione adducts [Ill.In the present study, the tissue distribution of activities of DHP-I and aminopeptidase in rats was investigated, and DHP-I was purified from the rat lung, where its specific activity was found to be higher than in the kidney. The immunological identity between pulmonary and renal DHP-I was also compared using anti-(renal DHP-I) antibody.