In vitro metabolism of human and salmon calcitonins in rat liver and kidney evaluated by liquid chromatography-tandem mass spectrometry

1. Using LC-MS and LC-MS/MS, an in vitro study was conducted on the metabol ism of human calcitonin (hCT) and salmon calcitonin (sCT) in rat liver and kidney to determine the rates of metabolism and the positions of hydrolytic cleavage in both peptides. 2. In lysosomal fractions of rat liver and kidney, hCT was degraded 9-12 ti mes faster than sCT. Many metabolites of hCT were produced in the lysosomal fractions, whereas the metabolites of sCT were scarcely found. 3. In the case of the cytosolic fractions, three positions of initial endop roteolytic cleavage were found in hCT, leading to the production of many pe ptide fragments via subsequent exoproteolytic metabolism. The initial cleav age position of sCT could not be identified precisely, but it was postulate d that the rate-determining step in the metabolism of sCT is the endoproteo lytic hydrolysis. 4. The studies using pure proteases and protease inhibitors indicated that the metabolism of calcitonins proceeds by initial endoproteolytic cleavage and subsequent exoproteolytic digestion, catalysed by an aspartate-protease in lysosomes and by a metalloprotease and cysteine-protease in combination in the cytosol. 5. The result suggested that the higher in vivo pharmacological activity of sCT compared with that of hCT may be due to a slower metabolism of the for mer.