UPLC-MS/MS法测定小鼠血浆中紫杉醇脂肪酸酯前药及其药代动力学研究

Determination and pharmacokinetics investigation of prodrugs of paclitaxel fatty acid esters in mouse plasma by UPLC-MS/MS

目的建立小鼠血浆中紫杉醇肉豆蔻酸酯(PTX-MA)、紫杉醇棕榈酸酯(PTX-PA)和紫杉醇硬脂酸酯(PTX-SA)等3种紫杉醇脂肪酸酯的超高效液相色谱-串联质谱(UPLC-MS/MS)测定方法,并初步考察其脂质体的小鼠体内药代动力学特性。方法采用EclipsePlus C_(8)色谱柱(2.1 mm×50 mm,1.8μm),0.2%甲酸水溶液(A)和甲醇(B)的不同比例混合液为流动相,梯度洗脱,三重四极杆串联质谱多重反应监检测(MRM),流速:0.3 ml/min,柱温:30℃,进样量:10μl。结果PTX-MA、PTX-PA和PTX-SA在5.0~500.0 ng/ml范围内均呈良好的线性关系(r>0.9950),日内和日间精密度、稳定性、提取回收率和基质效应试验结果的RSD均小于10%;3种紫杉醇脂肪酸酯脂质体PTX-MA-L、PTX-PA-L和PTX-SA-L在小鼠体内的半衰期(t_(1/2))分别为14.78、44.49和69.32 h,清除率(CL)分别为29.06、24.94和13.74 L·kg/h。结论该方法专属性高、灵敏、操作简便、稳定性好,可用于小鼠血浆中紫杉醇脂肪酸酯的含量测定。小鼠体内药代动力学研究结果表明,随脂肪酸碳链增加,紫杉醇脂肪酸酯在小鼠体内t_(1/2)呈大幅延长,清除率则显著降低,提示紫杉醇经不同链长饱和脂肪酸酯化修饰可改变其体内药代动力学特性,从而为紫杉醇脂肪酸酯前药的纳米制剂研发提供科学依据。

Objective To establish an UPLC-MS/MS method for determinating content of three paclitaxel fatty acid esters such as paclitaxel myristate(PTX-MA),paclitaxel palmitate(PTX-PA)and paclitaxel myristate(PTX-SA)in mouse plasma,and preliminarily investigate the pharmacokinetic characteristics of their liposomes in mice.Methods Eclipse Plus C_(8)chromatography column(2.1 mm×50 mm,1.8μm)was used with different proportions of 0.2%formic acid aqueous solution(A)and methanol(B)mixture as mobile phase for gradient elution at a flow rate of 0.3 ml/min.The collum temperature was 30℃.The sample injection volume was 10μl.The triple quadrupole mass series spectrometer was used as multi-reaction monitoring(MRM).Results PTX-MA,PTX-PA and PTX-SA all exhibited a good linear relationship in the range of 5.0~500.0 ng/ml(r>0.9950).Their RSD of precision,stability,extraction recovery rate and matrix effect test results was all less than 10%.The half-lives(t_(1/2))for liposomes of three paclitaxel fatty acid esters PTX-MA-L、PTX-PA-L and PTX-SA-L in mice were 14.78 h,44.49 h and 69.32 h individually,and their clearance rates(CL)were 29.06 L·kg/h,24.94 L·kg/h and 13.74 L·kg/h,respectively.Conclusion This method had high specificity,sensitivity,easy operation and good stability,which could be used for the determination of paclitaxel fatty acid esters in mouse plasma.The results of pharmacokinetic studies in mice showed that t_(1/2)for paclitaxel fatty acid esters were significantly prolonged,and the clearance rate were significantly reduced with the length of fatty acid carbon chains increasement,which indicated that esterification of paclitaxel with different chain length saturated fatty acids could obviously alter its in vivo pharmacokinetic properties,which provided scientific basis for the research and development of nano formulations of paclitaxel fatty acid ester prodrug.