科研之友
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摘要
For quantitative assaying tylerdipine hydrochloride, and its two primary metabolites (M2 and M4) in human urine, two sensitive and accurate LC-MS/MS methods were firstly developed and validated, where multiple reaction monitoring (MRM) was applied under positive electrospray ionization mode for tylerdipine and negative electrospray ionization mode for M2/M4, respectively. Urinary proteins were precipitated using acetonitrile, and deuterated isotopes of tylerdipine and M4 ([D5]-tylerdipine and [D6]-M4) were used as internal standards. Triton X-100, a good surfactant, was used to prevent the adsorption. An Agilent Poroshell 120 column was employed for chromatographic separation of the analytes with the mobile phases of 2 mM ammonium formate solution (containing 0.1% formic acid) and acetonitrile (45:55 for tylerdipine and 75:25 for the M2/M4, v/v). Flow rate was 0.3 mL/min. Calibration curves for tylerdipine, M2 and M4 in urine were linear over the ranges of 0.02-10 ng/mL, 2-1500 ng/mL and 0.5-200 ng/mL, respectively. The precision, accuracy, specificity and stability of two methods all evaluated and achieved the acceptable criteria. The LC-MS/MS methods were successfully applied to assay urinary excretion of tylerdipine and the metabolites in healthy Chinese subjects who orally received a single dose of 20 mg tylerdipine tablet. Generally, the urinary excretion of the two primary metabolites accounted for 11.7% of the total dose of tylerdipine in healthy Chinese subjects, while little tylerdipine was recovered in urine.
