Ex vivo conversion of prodrug prothionamide to its metabolite prothionamide sulfoxide with different extraction techniques and their estimation in human plasma by LC-MS/MS.

BACKGROUND: The present work reports an ex vivo conversion study of prothionamide to its active metabolite prothionamide sulfoxide in human plasma during sample preparation by three conventional extraction techniques. RESULTS: The chromatography was done on a Hypersil™ Gold C18 (50 × 2.1 mm, 3.0 µm) column using 0.1% acetic acid and acetonitrile (20:80, v/v) as the mobile phase. Quantitation of the analytes was done by MS/MS in the positive ionization mode. The method was validated over a wide concentration range of 30 to 6000 ng/ml for prothionamide and 50 to 10,000 ng/ml for prothionamide sulfoxide. The recovery for both the analytes was greater than 89%. Stability was extensively validated under different storage conditions. CONCLUSION: The extraction protocol was optimized using acetonitrile as protein precipitant for their simultaneous determination in human plasma by LC-MS/MS. The method was applied to a bioequivalence study of 250 mg prothionamide tablet formulation in 14 healthy Indian subjects.

Disposition of prothionamide in rats and armadillos.

A new method for the sensitive and selective measurement of prothionamide (PTH) and its S-oxide metabolite (PTHSO) in biological fluids was described. The limit of sensitivity was approximately 0.01 microgram of drug/ml of plasma. Endogenous materials, 2-propylisonicotinamide, ethionamide, dapsone, or monoacetyl dapsone did not interfere or contribute. Rats receiving PTH, intravenously or orally, showed a sexual dimorphism in the ability to oxidize PTH to PTHSO, with males exhibiting greater capacities for this conversion. Both sexes cleared the administered PTH more rapidly from the plasma than the metabolite, PTHSO. Following oral or intravenous administration of equimolar doses of PTHSO, both sexes exhibited an ability to reduce the administered PTHSO to PTH, with the female showing greater capacities for this conversion. Clearances after oral PTHSO administration were again more rapid for PTH than for PTHSO in both sexes. However, the total of PTH and PTHSO in the plasma during 8 hr following PTHSO administration was consistently less than following PTH dosing. Therefore, although PTHSO is retained longer than PTH after either PTH or PTHSO administration, giving PTHSO yielded less total active drug in the circulation. Comparison of plasma patterns of PTH and PTHSO in unfasted rats receiving one oral or eight daily oral doses of PTH did not indicate that PTH induces its own metabolism. Limited studies in armadillos receiving PTH and PTHSO intravenously led to the same general conclusions as those we derived from the rat studies regarding the disposition of PTH and PTHSO.

Mutagenic activity of antileprosy drugs and their derivatives.

We tested the mutagenic activity of antileprosy drugs (clofazimine, ethionamide, prothionamide, prothionamide-S-oxide, rifampin, and dapsone and many of its derivatives) using the Ames Salmonella/microsome assay system. None of these, including N-acetylated and N-hydroxylated derivatives of dapsone, were found to be positive with or without metabolic activation of this test. However, the sulfoxide and sulfide analogs of dapsone were found to be mutagenic with metabolic activation. These two analogs could not be detected in pharmaceutical preparations of dapsone (less than 0.01%), nor could they be found (in either unconjugated or conjugated form) in urine from volunteers taking a single oral dose of 50 mg of dapsone or from patients receiving daily oral doses of 100 mg of dapsone. Also, urine concentrates from volunteers taking 50 mg of dapsone did not exhibit mutagenic activity in the Ames screen. These results indicate that patients receiving antileprosy therapy with clofazimine, dapsone, ethionamide, prothionamide, and/or rifampin are not being exposed to mutagenic (and thereby possible carcinogenic) drugs.