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目的:建立大鼠组织中阿德福韦(PMEA)含量测定的柱前衍生化HPLC-荧光检测法,研究新型阿德福韦前药M1、M2在Wister大鼠组织中的分布情况。方法:Benetnach C18(150 mm×4.6 mm,5μm)色谱柱,柱前衍生化;流动相A:含2%乙腈的25 mmol·L-1磷酸二氢钾缓冲溶液(5 mmol·L-1氢氧化四丁基铵,pH 6.0),流动相B:含65%乙腈的25 mmol·L-1磷酸二氢钾缓冲溶液(5 mmol·L-1氢氧化四丁基铵,pH 6.0),梯度洗脱;流速1.0 mL·min-1;柱温35℃;荧光检测波长为λex 236nm和λem 420 nm。以泰诺福韦(PMPA)为内标,测定Wister大鼠肝、肾中的药物浓度,比较新型前药M1、M2与阿德福韦酯(ADV)的组织分布特点。结果:组织匀浆中PMEA线性关系、最低检测限、准确度、精密度均符合分析要求。大鼠肝组织中药物浓度:M2>M1>ADV,肾组织中药物浓度:ADV>M1>M2。结论:该方法准确、稳定,灵敏度高,可用于PMEA的检测及药动学研究。在大鼠组织中M2比M1和ADV具有更强的肝靶向性和更弱的肾靶向性,具有继续研发的价值。
OBJECTIVE: To establish a pre-column derivatization HPLC-fluorescence detection method for the determination of adefovir (PMEA) in rat tissues and to study the distribution of novel adefovir prodrugs M1 and M2 in Wister rats. METHODS: Benetnach C18 (150 mm × 4.6 mm, 5 μm) column was used to derivatize the column. Mobile phase A was a 25 mmol·L-1 potassium dihydrogen phosphate buffer solution containing 5% Tetrabutylammonium hydroxide, pH 6.0), mobile phase B: 25 mmol·L-1 potassium dihydrogen phosphate buffer (5 mmol·L -1 tetrabutylammonium hydroxide, pH 6.0) containing 65% acetonitrile, Eluting at a flow rate of 1.0 mL · min-1. The column temperature was 35 ℃. The detection wavelength was λex 236nm and λem 420nm. Tenofovir (PMPA) was used as an internal standard to determine the drug concentration in the liver and kidney of Wister rats. The tissue distribution characteristics of novel prodrugs M1, M2 and adefovir dipivoxil (ADV) were compared. Results: The linear relationship of PMEA in tissue homogenate, the lowest detection limit, the accuracy and the precision all met the analytical requirements. The concentration of drug in rat liver tissue: M2> M1> ADV, the concentration of drug in kidney tissue: ADV> M1> M2. Conclusion: The method is accurate, stable and sensitive. It can be used for the detection and pharmacokinetics of PMEA. M2 has greater liver targeting and weaker kidney targeting than M1 and ADV in rat tissue, with the potential to continue its development.