The effect of absorption enhancers on the initial degradation kinetics of insulin by alpha-chymotrypsin.
The goal of this investigation was to establish a fast method to screen various insulin absorption enhancers by following their effect on the initial kinetics of insulin incubated with alpha-chymotrypsin at 37 degrees C. A simple, sensitive and reproducible reversed phase high performance liquid chromatography (HPLC) method has been developed to carry out this goal. Linear responses (r > 0.999) were observed over the range of 0.4-4 U/ml for insulin. There was no significant difference (P < 0.05) between inter- and intra-day studies for insulin. The mean relative standard deviations (RSD%) of the results of within-day precision and accuracy of insulin were 12%. The assay was sensitive to detect the existence of any metabolite due to the addition of any absorption enhancers, even if it was not seen with insulin alone. Three metabolites (A-C) were detected only when insulin was incubated with alpha-chymotrypsin at 37 degrees C. Metabolite D was observed when either glycocholic acid (0.5, 1%) or taurochenodeoxycholate (0.5, 1%) was incubated with insulin in the absence of alpha-chymotrypsin at 37 degrees C. The compounds that significantly increased insulin T50% were glycyrrhizic acid (0.5%) > deoxycholic acid (1%) > deoxycholic acid (0.5%) > glycyrrhizic acid (1%) > cholic acid (0.5, 1%). Capric acid (0.5%), hydroxypropyl-alpha-cyclodextrin (0.5, 1%) and dimethyl-alpha-cyclodextrin (0.5, 1, 5%) did not significantly affect insulin T50%. The bile salts increased insulin T50% in this order: deoxycholate > cholate > glycocholate > taurocholate > taurodeoxycholate > taurochenodeoxycholate > glycodeoxycholate. The results obtained would support the feasibility of utilizing such method for screening any compound incorporated in insulin formulation. These compounds should be used in the minimum possible concentration to avoid or minimize insulin degradation.
Radwan
M A
MA
Department of Clinical Pharmacy, College of Pharmacy, Science and Medical Studies, Department for Women Students, King Saud University, P.O. Box 22452, 11495, Riyadh, Saudi Arabia. mradwan@ksu.edu.sa
Aboul-Enein
H Y
HY
D000042:Absorption; D000287:Administration, Topical; D000818:Animals; D000893:Anti-Inflammatory Agents / Q000737:chemistry / Q000378:metabolism; D001647:Bile Acids and Salts / Q000737:chemistry* / Q000378:metabolism; D002417:Cattle; D002626:Chemistry, Pharmaceutical; D019826:Cholic Acid / Q000737:chemistry / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D002918:Chymotrypsin / Q000737:chemistry* / Q000378:metabolism; D003840:Deoxycholic Acid / Q000737:chemistry / Q000378:metabolism; D003902:Detergents / Q000378:metabolism; D006034:Glycyrrhetinic Acid / Q000737:chemistry / Q000378:metabolism; D007004:Hypoglycemic Agents / Q000737:chemistry* / Q000378:metabolism; D007328:Insulin / Q000737:chemistry* / Q000378:metabolism
D000893:Anti-Inflammatory Agents; D001647:Bile Acids and Salts; D003902:Detergents; D007004:Hypoglycemic Agents; D007328:Insulin; D003840:Deoxycholic Acid; D002918:Chymotrypsin; C117608:alpha-chymotrypsin; D019826:Cholic Acid; D006034:Glycyrrhetinic Acid