RRLC-MS/MS法测定大鼠血浆中咖啡酸四聚体的含量_及药代动力学研究

目的: 建立大鼠血浆中咖啡酸四聚体(CAT)的测定方法,并研究其药代动力学。方法: 大鼠尾静脉注射CAT后不同时间经眼静脉采血,血浆样品经液-液萃取后,采用RRLC-MS/MS法,色谱柱为Agilent SB-C₁₈(4.6 mm×100 mm,1.8 μm),流动相为0.1%甲酸溶液(A)-乙腈(B),梯度洗脱(0~3 min,68%A;3~6.5 min,68%A→2%A;6.5~10 min,2%A),流速0.4 mL·min^-1,柱温30 ℃,进样量10 μL;质谱采用电喷雾离子源(ESI源),正离子模式检测,扫描方式为多反应监测(MRM)。结果: 咖啡酸四聚体检测限和最低定量限分别为5.7和11.4 ng·mL^-1;在11.4~1420 ng·mL^-1浓度范围内线性关系良好(r=0.9991);提取回收率为85.2%~97.1%;药动学参数:AUC_0-t为(1.25±0.25)mg·L^-1·h,t_1/2为(4.66±1,07)h^-1,K为(0.15±0.04)h^-1,CLz为(3.19±0.52)L·h^-1·kg^-1,Vz为(1.87±0.59)L·kg^-1。结论: 本方法专属性强、灵敏度高、准确性好,可用于咖啡酸四聚体的血药浓度测定。

Objective: To establish a method to determine the plasma concentration of caffeic acid tetramer(CAT)in rats by using RRLC-MS/MS,and study the pharmacokinetics of CAT. Methods: CAT in rat tail vein and plasma samples were collected at different time points through vena ophthalmica.After a liquid-liquid extraction process with methl tert-mutyl ether,the HPLC separation for plasma samples was performed on a Agilent SB-C_l8 column(4.6 mm×100 mm,1.8 μm)in gradient elute mode(0-3 min,68%A;3-6.5 min,68%A→2%A;6.5-10 min,2%A)with water containing 0.1% formic acid(A)and acetonitrile(B)as the mobile phase at a flow rate of 0.4 mL·min^-1.MS detection was performed with multiple reaction monitoring(MRM)mode using positive electrospray ionization. Results: The linearity was achieved in the range of 11.4-1420 ng·mL^-1(r=0.9991)for CAT.The extraction recovery of CAT was from 85.2%-97.1%.The lowest limit of detection(LOD)and quantification(LOQ)of caffeic acid tetramer were 5.7 ng·mL^-1and 11.4 ng·mL^-1.The pharmacokinetic parameters of AUC_0-t,t_1/2,K,CLz and Vz were as follows:(1.25±0.25)mg·L^-1·h,(4.66±1,07)h^-1,(0.15±0.04)h^-1,(3.19±0.52)L·h^-1·kg^-1,and(1.87±0.59)L·kg^-1. Conclusion: A sensitive,accuracy and suitable RRLC-MS/MS method for determining CAT was developed.