Acta pharmaceutica Nordica最新文献

The N-terminal pyroglutamyl group in several peptides is specifically cleaved by pyroglutamyl aminopeptidase (PAPase I). With the aim of protecting this group against enzymatic cleavage by the prodrug approach, various derivatives of L-pyroglutamyl benzylamide, used as a PAPase I sensitive model pyroglutamyl peptide, were prepared and their stability characteristics determined. The derivatives studied included phenoxycarbonyl, phthalidyl, hydroxymethyl and actoxymethyl derivatives, all formed at the pyroglutamyl NH-moiety. Whereas L-pyroglutamyl benzylamide was rapidly hydrolyzed by PAPase I, all the derivatives were resistant to cleavage by the enzyme. On the other hand, these derivatives, with the exception of the N-phenoxycarbonyl derivative, were readily converted to the parent pyroglutamyl benzylamide by spontaneous or plasma catalyzed hydrolysis, the half-lives of conversion in 80% human plasma being in the range 2.3-8.4 h. The major degradation reaction of the N-phenoxycarbonyl derivative in both buffer and plasma solutions was hydrolytic opening of the pyrrolidone ring. The pH-rate profiles for the degradation of the compounds in aqueous solution were obtained and both specific acid and base catalytic reactions as well as a spontaneous reaction were observed. The results suggest that N-phthalidylation, N-hydroxymethylation and N-acyloxymethylation of pyroglutamyl peptides may be useful prodrug approaches to protect such peptides against cleavage by pyroglutamyl aminopeptidase and hence to improve their delivery characteristics.

The formation of Metabolic Intermediate (MI) complexes from a series of beta-alkylsubstituted 2-phenylethanamines and corresponding N-hydroxylamines is investigated during NADPH-dependent metabolism in liver microsomes from phenobarbital pretreated rats. The beta-alkyl substituents are methyl, dimethyl, ethyl, di-ethyl, n-propyl, di-n-propyl and i-propyl groups. The amines are synthesized by LiAlH4-reduction of the corresponding nitriles, which are prepared through alkylation of the enolate anion of phenylacetonitrile. The hydroxylamines are prepared either by oxidation of the corresponding benzylimines with m-chloroperbenzoic acid and subsequent hydrolysis of the initially formed 3-phenyloxaziridines, or by H2O2-mediated oxidation of the corresponding amines in the presence of catalytic amounts of sodium tungstate, followed by reduction with cyanoborohydride. The amines are found to be completely devoid of complexing activity, while the hydroxylamines form the MI complex at high rates. Complex formation from these substrates thus parallels the known behaviour of 2-phenylethanamine and its corresponding N-hydroxylamine. Since N-oxygenation is known to be a prerequisite for MI complex formation from amines our results suggest that the beta-alkylated 2-phenylethanamines are metabolized exclusively through other pathways. In accordance with this hypothesis, capillary GC-analysis of the incubation mixture of 2-phenylpropanamine shows no formation of N-hydroxylated metabolites; only 2-phenylpropanol, a metabolite formed through the deamination pathway, is found.

The influence of auxiliary substances and pH on the chemical transformations of insulin in pharmaceutical formulation, including various hydrolytic and intermolecular cross-linking reactions, was studied. Bacteriostatic agents had a profound stabilizing effect--phenol > m-cresol > methylparaben--on deamidation as well as on insulin intermolecular cross-linking reactions. Of the isotonicity substances, NaCl generally had a stabilizing effect whereas glycerol and glucose led to increased chemical deterioration. Phenol and sodium chloride exerted their stabilizing effect through independent mechanisms. Zinc ions, in concentrations that promote association of insulin into hexamers, increase the stability, whereas higher zinc content had no further influence. Protamine gave rise to additional formation of covalent protamine-insulin products which increased with increasing protamine concentration. The impact of excipients on the chemical processes seems to be dictated mainly via an influence on the three-dimensional insulin structure. The effect of the physical state of the insulin on the chemical stability was also complex, suggesting an intricate dependence of intermolecular proximity of involved functional groups. At pH values below five and above eight, insulin degrades relatively fast. At acid pH, deamidation at residue A21 and covalent insulin dimerization dominates, whereas disulfide reactions leading to covalent polymerization and formation of A- and B-chains prevailed in alkaline medium. Structure-reactivity relationship is proposed to be a main determinant for the chemical transformation of insulin.