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J Drug Target. 1994;2(1):79-89.

The Caco-2 cell monolayers as an intestinal metabolism model: metabolism of dipeptide Phe-Pro.

Author information

1
Department of Pharmaceutical Sciences, College of Pharmacy, Washington State University, Pullman 99164-6510.

Abstract

The metabolism of Phe-Pro was investigated in Caco-2 cell monolayers, a model of small intestinal epithelium. The results indicate that the majority of Phe-Pro was hydrolyzed during passage from the apical (AP) to basolateral (BL) side. The enzyme responsible for the hydrolysis is prolidase, a cytosolic enzyme. Through kinetic studies of a supernatant enzyme preparation, a Km of 30.4 microM and Vmax of 38.9 nmol/min per mg of protein were obtained. The enzyme catalyzed hydrolysis was inhibited by proline (66%), Zn+ (86%), Cu++ (100%), Fe (100%), PCMB (89%), and captopril (66%), but not by leucine. We also studied the transcellular transport of Phe-Pro by measuring the amount of Phe in the receiver media. In the presence of a proton gradient (AP pH6, BL pH7.4), the appearance rate of Phe in the BL media after Phe-Pro was loaded apically was at least 100 times faster than that in the AP media after Phe-Pro was loaded basolaterally. The former is also higher than the appearance rate of Phe without a transepithelial proton gradient (pH 6-pH 6) or against a proton gradient (pH7.4-pH6). The rate of appearance of Phe in the BL media (pH7.4) after Phe-Pro was loaded on the AP side (pH 6) was decreased by the presence in the AP media of proline (42%), leucine (40%), and captopril (17%), but not by Zn++. In conclusion, the transmembrane uptake of Phe-Pro is dependent on a proton gradient, and the intracellular metabolism of Phe-Pro is complete via hydrolysis by prolidase.

PMID:
8069586
DOI:
10.3109/10611869409015895
[Indexed for MEDLINE]

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