Study on the phase II metabolites of phenoprolamine hydrochloride in rat bile by LC/DAD/MSD

AIM To study the phase II metabolites of phenoprolamine hydrochloride (DDPH) in rat bile. METHODS DDPH was administered by ip to bile duct-cannulated rats. Bile samples were collected before drug administration and up to 12 h after drug administration. After being purified and enriched with C-18 SPE columns the rat bile samples were analyzed by LC/DAD/MSD to identify the peaks of phase II metabolites. The fractions of phase II metabolites were prepared by HPLC and treated with β-glucuronidase, and then were purified and enriched with C-18 SPE columns and analyzed by LC/DAD/MSD. The corresponding reference standards of DDPH phase I metabolites were analyzed by LC/DAD/MSD under identical conditions. RESULTS The peaks M₇, M₈ and M₉ in the chromatograms of rat bile samples were the phase II metabolites DDPH and the enzymatic hydrolysates of M₇, M₈ and M₉ were 1-(2, 6-dimethyl-4-hydroxyphenoxy)-2-(3, 4-methoxyphenylethylamino)-propane (M₃), 1-(2, 6-dimethyl-3-hydroxyphenoxy)-2-(3, 4-methoxyphenyl-ethylamino)-propane (M₂) and 1-(2, 6-dimethylphenoxy)-2-(3-methoxy-4-hydroxyphenylethyl-amino)-propane (M₁) respectively. CONCLUSION β-1-O-{3, 5-dimethyl-4-[-2-methyl-2-(3, 4-dimethoxy-phenylethylamino)-ethoxy]-phenyl}-glucuronic acid (M₇, glucuronide of M₃), β-1-O-{2, 4-dimethyl-3-[2-methyl-2-(3, 4-dimethoxy-phenylethylamino)-ethoxy]-phenyl}-glucuronic acid (M₈, glucuronide of M₂) and β-1-O-{2-methoxy-4-[1-methyl-2-(2, 6-dimethylphenoxy)-ethylamino-ethyl]-phenyl}-glucuronic acid (M₉, glucuronide of M₁) were the phase II metabolites of DDPH in rat bile.