This article is available online at http://dmd.aspetjournals.org ABSTRACT:This study uses stable isotope methodology to evaluate the validity of 6␤-hydroxylation clearance of endogenous cortisol as a new index for in vivo CYP3A phenotyping in humans. Important factors contradictory to the use of a conventional index of urinary ratio of 6␤-hydroxycortisol to cortisol (6␤-OHF/F) to evaluate in vivo CYP3A activity are also discussed. Stable isotopically labeled cortisol (3-5 mg) was orally administered to three healthy adult subjects to accurately determine the fractional metabolic clearance specific for the 6␤-hydroxylation of cortisol. Plasma concentrations of labeled cortisol and urinary excreted amounts of labeled cortisol and 6␤-OHF were analyzed by gas chromatography-mass spectrometry simultaneously with their endogenous counterparts. There was a good correlation between endogenous and exogenous 6␤-hydroxylation clearances in the three subjects tested (r ‫؍‬

The conjugation of xenobiotic carboxylic acids with b-Dglucuronic acid to yield 1-O-acyl-b-D-glucopyranuronates (1b-O-acyl glucuronides) is a major metabolic pathway for many compounds, including 2-arylpropionic acids (profens). The 1b-O-acyl glucuronides are labile and reactive in neutral and mildly alkaline aqueous solution, and undergo both hydrolysis and intramolecular acyl migration (Fig. 1). In acyl migration, the aglycone is transferred from the 1b position to the neighboring 2-hydroxyl group on the glucuronic acid moiety via an ortho-acid ester intermediate. 1,2) This process continues on to the OH-groups at C-3 and C-4, thereby forming several regioisomers. The glucuronide regioisomers readily ring-open and mutarotate to give a-and b-anomers. The acyl migrations are generally reversible, but acyl migration from C-2 to C-1 to reform the 1b-O-acyl glucuronide is not commonly observed. These acyl glucuronides are capable of reacting with protein nucleophiles to form covalent adducts.3-12) The protein adducts have been hypothesized as mediators of hypersensitive responses to carboxylate drugs. 1,13) Profens possess a chiral center and are marketed as racemates. Profens are metabolized predominantly to the diastereomeric acyl glucuronides with different physicochemical properties. Consequently, the diastereomeric acyl glucuronides might have distinct toxicological actions and be differentially eliminated. Although the stability of the diastereomeric acyl glucuronides has been investigated in several profens, 7,[14][15][16][17][18][19] little information is currently available on the factors that contribute to stereoselective differences in the degradation process.Enantiomeric 2-phenylpropionic acids (PAs) possess the fundamental structure of profens (Chart 1) and have been used as model substrates of profens to study the stereoselective metabolism and disposition of profens. PAs show similar pharmacokinetic properties to other profens, as observed in stereoselective isomerization and glucuronidation in rodents. The stereoselective acyl migration of diastereomeric 1b b-O-acyl glucuronides of (R)-and (S)-2-phenylpropionic acid [(R)-1PG and (S)-1PG, respectively] in phosphate buffer (pH 7.4) at 310 K was investigated using HPLC. The disappearance of (R)-1PG was faster than that of (S)-1PG according to pseudo first-order kinetics. A kinetic model describing the degradation reactions was constructed. The rate constant for acyl migration from the 1b b-O-isomer to the 2-O-acyl isomer (k 12 ) was about one order magnitude larger than that for hydrolysis from 1b b-O-acyl isomer to aglycone (k 10 ). The k 12 of (R)-1PG (0.377؎0.005 h ؊1 ) was about two times larger than that of (S)-1PG (0.184؎0.003 h ؊1 ). The results indicated that the stereoselectivity in the degradation of 1PG was apparently governed by the acyl migration from 1-isomer to 2-isomer. The kinetic parameters for acyl migration from 1-isomer to 2-isomer were estimated from temperature-dependent experiments using the transition state theory. The value o...

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The aim of this study was to evaluate the absorption of a citrus flavonoid, naringin, as its glycosylated form. Six healthy volunteers (three males and three females) were studied. After a single oral administeration of 500 mg of naringin, intact naringin was isolated from 2-4 h urine. Isolated naringin was identified by the LC/electrospray ionization mass spectrometry (ESI-MS), MS/MS, and MS/MS/MS techniques. The cumulative urinary excretion of naringin and its metabolites (naringenin and naringenin glucuronides) was determined by HPLC for 0-24 h. Approximately 0.02% of the administered dose was recovered in urine as unchanged naringin, whereas urinary recoveries of naringenin and naringenin glucuronides were approximately 0.4 and 3.6% of the administered dose, respectively. It was concluded that trace amounts of orally administered naringin can be absorbed as the glycoside. However, it is not clear whether the glycoside is cleaved before or after absorption to generate naringenin.