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目的:建立大鼠尿液中阿仑膦酸钠柱前衍生HPLC-UV测定方法,并应用于大鼠口服阿仑膦酸钠后尿液排泄动力学研究。方法:大鼠单次灌胃给予阿仑膦酸钠溶液(4.5 mg·kg~(-1)),在给药后不同时间段收集尿液,尿液固相萃取预处理后,采用异硫氰酸苯酯(PITC)对尿液中的阿仑膦酸钠衍生化。液相色谱分析采用Venusil AA分析柱(4.6 mm×250 mm,5μm),以乙酸钠溶液-乙腈为流动相,梯度洗脱,正亮氨酸为内标,流速0.8 mL·min~(-1),检测波长270 nm。结果:阿仑膦酸钠质量浓度在2.5~250.0μg·mL~(-1)范围内线性良好(r=0.999);方法的日内和日间精密度小于3.0%,提取回收率在87.5%~89.2%之间;检测限为0.2μg·mL~(-1)。尿液排泄动力学曲线显示阿仑膦酸钠溶液口服后达峰时间约为2.16 h,清除半衰期为(4.5±0.12)h,36 h尿液累积量为(42.6±1.2)μg,占给药剂量的4.7%。结论:本方法经方法学验证,可用于阿仑膦酸钠尿液排泄动力学研究。
OBJECTIVE: To establish a HPLC-UV method for the determination of alendronate in urine of rats and to study the urine excretion kinetics after oral administration of alendronate in rats. Methods: Rats were given alendronate sodium solution (4.5 mg · kg -1) orally by gavage. Urine was collected at different time points after administration. After pretreatment with urine solid phase extraction, Phenyl cyanate (PITC) derivatization of alendronate in urine. The liquid chromatographic analysis was performed on a Venusil AA column (4.6 mm × 250 mm, 5 μm) using gradient elution with sodium acetate-acetonitrile as mobile phase and n-leucine as an internal standard at a flow rate of 0.8 mL · min -1 ), Detection wavelength of 270 nm. Results: The calibration curve of alendronate was linear in the range of 2.5-250.0 μg · mL -1 (r = 0.999). The intra- and inter-day precision was less than 3.0% and the recovery was 87.5% 89.2%; the detection limit was 0.2μg · mL -1. Urine excretion kinetics curves showed that alendronate sodium peaked at about 2.16 h after oral administration, the half-life of elimination was (4.5 ± 0.12) h, and the accumulated urine volume at 36 h was (42.6 ± 1.2) μg, accounting for 4.7% of dose. Conclusion: This method is validated by methodology and can be used to study urinary excretion kinetics of alendronate.