A Bernkop-Schnürch1, M K Marschütz. 1. Center of Pharmacy, University of Vienna, Austria/Europe. andreas.bernkop-schnuerch@univie.ac.at
Abstract
PURPOSE: Develop and evaluate systems to prevent aminopeptidase N caused enzymatic degradation of perorally administrated peptide drugs. METHODS: Bacitracin was covalently bound to the unabsorbable carrier matrix poly(acrylic acid) (paa) in order to avoid any dilution effects of the inhibitor in the intestine as well as systemic toxic side effects. The inhibitory effect of this conjugate, of neutralized paa and N-acetylcysteine was evaluated using a brush border membrane model. RESULTS: Whereas within 6 h of incubation 65.3 +/- 3.7 mumol/l of the substrate (L-leucine p-nitroanilide) was hydrolyzed under our assay conditions, this metabolism was reduced to 44.5 +/- 6.3 mumol/l and 49.0 +/- 8.8 mumol/l (n = 3-5; +/- S.D.) using 1.5% bacitracin-polymer conjugate and 0.5% N-acetylcysteine, respectively. The same amount of bacitracin as immobilized to the polymer exhibited a comparably weaker inhibitory effect. Neutralized paa did not inhibit membrane bound aminopeptidase N. Covering the membrane with a thin mucus layer led to a significantly lowered inhibitory effect of all tested agents. CONCLUSIONS: The immobilization of enzyme inhibitors to a carrier matrix and the use of N-acetylcysteine as a novel inhibitor are promising strategies in order to overcome the enzymatic barrier caused by membrane bound peptidases. However the use of effective mucolytic agents seems to be a prerequisite.
PURPOSE: Develop and evaluate systems to prevent aminopeptidase N caused enzymatic degradation of perorally administrated peptide drugs. METHODS:Bacitracin was covalently bound to the unabsorbable carrier matrix poly(acrylic acid) (paa) in order to avoid any dilution effects of the inhibitor in the intestine as well as systemic toxic side effects. The inhibitory effect of this conjugate, of neutralized paa and N-acetylcysteine was evaluated using a brush border membrane model. RESULTS: Whereas within 6 h of incubation 65.3 +/- 3.7 mumol/l of the substrate (L-leucine p-nitroanilide) was hydrolyzed under our assay conditions, this metabolism was reduced to 44.5 +/- 6.3 mumol/l and 49.0 +/- 8.8 mumol/l (n = 3-5; +/- S.D.) using 1.5% bacitracin-polymer conjugate and 0.5% N-acetylcysteine, respectively. The same amount of bacitracin as immobilized to the polymer exhibited a comparably weaker inhibitory effect. Neutralized paa did not inhibit membrane bound aminopeptidase N. Covering the membrane with a thin mucus layer led to a significantly lowered inhibitory effect of all tested agents. CONCLUSIONS: The immobilization of enzyme inhibitors to a carrier matrix and the use of N-acetylcysteine as a novel inhibitor are promising strategies in order to overcome the enzymatic barrier caused by membrane bound peptidases. However the use of effective mucolytic agents seems to be a prerequisite.