AIM To establish an HPLC
assay to determine directly, the
propranolol glucuronides in microsomal incubate without hydrolysis to their parent enantiomers. METHODS The standard for this direct assay was prepared by incubation of liver
microsomes with propranolol. The
Glucuronide obtained was purified and concentrated with solid
phase extraction and the concentration was measured by an indirect
METHOD, i.e. HPLC assay of the propranolol after enzymatic hydrolysis with β glucuronidase. The direct assay involved
separation by HPLC using a C 18 reversed phase column, with UV detection at 290 nm. RESULTS The recovery of the assay was 99 4%±1 0% ( n =15), the reproducibility of the assay was less than 3% (RSD) for inter day and 5% (RSD) for intra day. The standard curves showed excellent linearity over the range 0 24-73 17 μmol·L -1 . Followed Michaelis Menten kinetics, the K m and V max for the two enantiomers were very different. The R enantiomer was glucuronidated at a more efficient rate than its enantiomorph, and was a better substrate. CONCLUSION The sensitive, reproducible and accurate HPLC method was developed to measure directly the propranolol glucuronides and has been applied to determine the stereoselectivity of glucuronidation metabolism of racemic propranolol in rat liver microsomes.
More abstracts about the HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY SEPARATION AND QUANTITATION OF PROPRANOLOL GLUCURONIDE IN