Identification of rat cytochrome P450 forms involved in the metabolism of clausenamide enantiomers

Abstract

To identify which cytochrome P450 (CYP) isoform(s) are responsible for the metabolism of clausenamide (CLA) enantiomers in rats, effects of various CYP isoform inducers and inhibitors on the formation of CLA metabolites were investigated in liver microsomes. In incubations with rat liver microsomes, CLA enantiomers were mainly converted to 4-hydroxy, 5-hydroxy, and 7-hydroxy-metabolites. 4-OH-CLA was the major metabolite of (+)-3R, 4S, 5S, 6R-CLA [(+)-CLA], while 7-OH-CLA was the major one of (-)-3S, 4R, 5R, 6S-CLA [(-)-CLA]. In induction studies, enzymatic parameters were used to assess the role of different CYP forms in CLA hydroxylation reactions. A marked increase in the rate of metabolism of CLA enantiomers was observed in microsomes of dexamethasone treated rats, V(max)/K(m) values for 4-OH-(+)-CLA, 7-OH-, 5-OH-, and 4-OH-(-)-CLA were 5.3, 6.5, 3.0, and 5.9 times higher than those in control microsomes, respectively. Rifampicin treatment caused corresponding 1.7-, 2.6-, 3.1-, and 2.8-fold increases. Dex and Rif also increased in the amount of (+)-5- and (+)-7-OH-CLA that were not detectable in the control group. These results suggested that inducible CYP3A1 was involved in the hydroxylation of CLA enantiomers. In inhibition studies, ketoconazone (6.25 microM) completely inhibited the production of main metabolites of (-)-CLA (100%) and (+)-CLA (97%). Triacetyloleandomycin (12.5 microM) strongly inhibited the corresponding metabolites by 34-85%. These findings also indicated that institutive CYP3A2 shared a major role in the hydroxylation of CLA enantiomers with CYP3A1 in untreated rats. Together, the data suggested that CYP3A was the predominant isoform responsible for the metabolism of CLA enantiomers.

Application of the PIFA–mediated alkyne amidation reaction to the formal synthesis of (±)-clausenamide

“…8 This alkaloid has shown efficiency as a liver-protecting agent against chemical toxins, 9 as an inductor effect over P450 cytochrome, 10 and also as a protector against cerebral hypoxia. 11 In fact, the possible application of this compound for the prevention and/or treatment of degenerative processes, such as Alzheimer's disease, is under study.…”

Stereoselective plasma protein binding and target tissue distribution of clausenamide enantiomers in rats

“…Concentrations of CLA enantiomers were determined by a reversed phase HPLC method described previously. 1,7,14 In short, HPLC system was comprised a SP-9A chromatograph, a SPD-6A UV detector (Shimadzu, Tokyo, Japan), a 7125 sampler (Rheodyne, USA), a 3394A integrator (Hewlett Packard, Palo Alto, CA) and a Kromasil C 18 column (4.6 3 250 mm internal diameter, 5 lm particle size; Eka Chemical AB, Bohus, Sweden). The mobile phase consisted of acetonitrile, methanol, and water at the ratio of 23:17:60 (v/v/v) and was delivered at a flow rate of 1.2 ml/min.…”

“…Each homogenate (2 ml) was taken into a 10-ml centrifuge tube containing 50 ll of internal standard solution(33 lg/ml in chloroform), extracted with 4 ml of chloroform, and analyzed by HPLC method. 1,14 Kinetics of Target Tissue Distribution of CLA Enantiomers…”

“…The homogenates were extracted and concentrations of CLA enantiomers were determined by the HPLC method. 1,14 Curves of CLA concentration in target tissue versus time were fitted by a two-compartment open model 15 plus the last measured plasma concentration divided by the terminal phase elimination rate constant (b). b was determined by log-linear regression analysis of the terminal data points.…”

Enhanced bioavailability of a new thiazolidine derivative FPFS-410, an antidiabetic and lipid-lowering drug, after oral administration of its hydroxypropyl-β-cyclodextrin complex to bile duct-cannulated rats

“…8). As CYP1A, CYP2A, CYP3A, CYP2B, CYP2C, and CYP2D are known to account for a major part of the CYP activity in rats, [21][22][23][24][25][26][27][28][29] specific inhibitors were used as follows: a-naphthoflavone (CYP1A), coumarin (CYP2A), ketoconazole (CYP3A), metyrapone (CYP2B), sulfaphenazole (CYP2C), and quinidine (CYP2D). As a result, ketconazole strongly suppressed the bioconversion of FPFS-410 to M1, but other specific inhibitors did not inhibit this bioconversion.…”