Evaluation of inhibitory activity of casein on proteases in rat intestine.
OBJECTIVE
To investigate the possible use of casein as an adjuvant for oral delivery of peptide drugs, the inhibitory activity of casein on proteases in rat small intestine was examined.


METHODS
Male Sprague-Dawley rats weighing 200-300 g were used as the animal model. The luminal contents of the small-intestinal tract and mucosal subcellular fractions of the small intestine were prepared: the enzymatic activities of trypsin, chymotrypsin, aminopeptidase-B, leucine aminopeptidase, dipeptidylaminopeptidase-IV, cathepsin B, and dipeptidylaminopeptidase-II were determined using a specific substrate for each protease; and the effect of casein on the protease activity was examined.


RESULTS
Casein strongly inhibited trypsin and chymotrypsin in the concentration-dependent manner. As to the proteases in the intestinal epithelial cells, casein inhibited an endopeptidase, cathepsin B, but not other exopeptidases. The inhibitory activity was independent of the type of casein. The kinetic analysis characterized the type of inhibition on trypsin and chymotrypsin to be competitive.


CONCLUSIONS
Casein was shown to have strong inhibitory activity on trypsin and chymotrypsin in the intestinal lumen. Taken into consideration that trypsin and chymotrypsin are endopeptidases, it is suggested that casein has strong inhibitory activity only on endopeptidases.
Department of Pharmaceutics, Faculty of Pharmaceutical Sciences, Okayama University, 1-1-1 Tsushima-naka, Okayama 700-8530, Japan.
Seiji
S
Ohtani
Kyoko
K
Shirasu
Ken-ichi
K
Ogawara
Kazutaka
K
Higaki
Toshikiro
T
Kimura
D000818:Animals; D002364:Caseins / Q000493:pharmacokinetics*; D002401:Cathepsin B / Q000037:antagonists & inhibitors / Q000378:metabolism; D002417:Cattle; D002918:Chymotrypsin / Q000037:antagonists & inhibitors / Q000378:metabolism; D007583:Jejunum / Q000187:drug effects* / Q000201:enzymology*; D008297:Male; D011480:Protease Inhibitors / Q000493:pharmacokinetics*; D051381:Rats; D017207:Rats, Sprague-Dawley
D002364:Caseins; D011480:Protease Inhibitors; D002918:Chymotrypsin; D002401:Cathepsin B