液相色谱-质谱联用同时测定大鼠血浆中替硝唑、达克罗宁和氯己定的浓度及其药动学研究

目的：建立大鼠血浆中替硝唑、达克罗宁和氯己定的HPLC-MS/MS定量分析方法。方法：采用固相萃取预处理血浆样品。使用Phenomenex Gemini C₁₈色谱柱（50 mm×2.0 mm，5 μm），以甲醇-10 mmol·L^-1甲酸铵-甲酸（56：44：0.2）为流动相，流速0.2 mL·min^-1，柱温25℃，酚妥拉明为内标;通过电喷雾离子化四极杆串联质谱，以正离子多反应监测方式（MRM）检测，用于定量分析的离子反应分别为m/z 247.9→m/z 82.0（替硝唑）、m/z 290.0→m/z 98.0（达克罗宁）、m/z 505.0→m/z 335.5（氯己定）、m/z 282.1→m/z 212.0（酚妥拉明）。结果：测定替硝唑、达克罗宁和氯己定的线性范围均为2～1000 ng·mL^-1，最低定量限为2 ng·mL^-1，提取回收率均 > 81.3%，日内RSD均 < 11.0%，日间RSD均 < 14.1%，准确度在-7.2%～2.9%范围内;替硝唑、达克罗宁、氯己定T_max分别为（1.50±0.93）、（1.50±0.43）、（1.50±1.05）h，C_max分别为（111.23±39.14）、（42.17±23.15）、（8.95±6.45） ng·mL^-1，t_1/2分别为（5.09±2.17）、（5.93±1.93）、（7.34±6.28）h，AUC_0-t分别为（804.52±87.66）、（145.57±21.86）、（99.31±24.01） ng·h·mL^-1。结论：该方法快速、灵敏、准确，可用于替硝唑、达克罗宁和氯己定在大鼠体内的药动学研究。

Objective: To develop and validate a rapid and sensitive HPLC-MS/MS method for simultaneous determination of tinidazole,dyclonine and chlorhexidine in rat plasma. Methods: The plasma samples were pretreated by the solid phase extraction (SPE) method.Separation was achieved on a Phenomenex Gemini C₁₈ column (50 mm×2.0 mm,5 μm) using an isocratic mobile phase system composed of methanol-ammonium formate (10 mmol·L^-1)-formic acid (56: 44: 0.2) at a flow rate of 0.2 mL·min^-1.The column temperature was controlled at 25℃.The internal standard was phentolamine mesylate.Analytes were determined by tandem mass spectrometry with electrospray positive ionization and multiple-reaction monitoring (MRM) mode.The monitoring ions were m/z 247.4→m/z 82.0 for tinidazole,m/z 290.0→m/z 98.0 for dyclonine,m/z 505.0→m/z 335.5 for chlorhexidine and m/z 282.1→m/z 212.0 for phentolamine. Results: The calibration curves were linear in the range of 2-1000 ng·mL^-1 and LLOQ were all 2 ng·mL^-1for the three compounds.The mean extraction recoveries were all above 81.3%.The intra-day and inter-day RSD values were less than 11.0% and 14.1%,respectively.The RE values were all within the range of-7.2%-2.9%.The main pharmacokinetic parameters oftinidazole, dyclonine, chlorhexidine were as follows: T_max (1.50±0.93), (1.50±0.43) and (1.50±1.05)h; C_max (111.23±39.14), (42.17±23.15) and (8.95±6.45) ng·mL^-1; t_1/2 (5.09±2.17), (5.93±1.93) and (7.34±6.28) h;AUC_0-t (804.52±87.66), (145.57±21.86) and (99.31±24.01) ng·h·mL^-1; respectively. Conclusion: The method is rapid,sensitive,accurate and can be applied to the pharmacokinetic research of tinidazole,dyclonine and chlorhexidine in rat plasma.