Determination of a novel selective inhibitor of type 1 5alpha-reductase in human plasma by liquid chromatography with atmospheric pressure chemical ionization tandem mass spectrometry.
A sensitive and specific assay of human plasma for the determination of (5alpha,7beta,16beta)-16[(4-chlorophenyl)oxy]-4,7- dimethyl-4-aza-androstan-3-one (I), a selective inhibitor of human type 1 5alpha-reductase, has been developed. The method is based on high-performance liquid chromatography (HPLC) with tandem mass spectrometric (MS-MS) detection. The analyte (I) and internal standard, Proscar (II), were isolated from the basified biological matrix using a liquid-liquid extraction with methyl-tert.-butyl ether (MTBE). The organic extract was evaporated to dryness, the residue was reconstituted in mobile phase and injected into the HPLC system. The MS-MS detection was performed on a PE Sciex API III Plus tandem mass spectrometer using a heated nebulizer interface. Multiple reaction monitoring using the precursor-->product ion combinations of m/z 430-->114 and 373-->305 was used to quantify I and internal standard (II), respectively. The assay was validated in the concentration range of 0.5 to 500 ng/ml in human plasma. The precision of the assay, expressed as coefficient of variation (C.V.), was less than 7% over the entire concentration range, with adequate assay specificity and accuracy. The HPLC-MS-MS method provided sufficient sensitivity to completely map the 24 h pharmacokinetic time-course following a single 0.5 mg dose of I.
Constanzer
M L
ML
Merck Research Laboratories, West Point, PA 19486, USA.
Chavez-Eng
C M
CM
Matuszewski
B K
BK
D001378:Azasteroids / Q000097:blood* / Q000494:pharmacology; D042944:Cholestenone 5 alpha-Reductase; D002851:Chromatography, High Pressure Liquid; D013196:Dihydrotestosterone / Q000031:analogs & derivatives* / Q000097:blood / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000097:blood* / Q000494:pharmacology; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors; D013058:Mass Spectrometry; D010088:Oxidoreductases / Q000037:antagonists & inhibitors*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D001378:Azasteroids; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D013196:Dihydrotestosterone; C115387:L 751788; D010088:Oxidoreductases; D042944:Cholestenone 5 alpha-Reductase