超高效液相色谱-串联质谱法研究紫杉醇对多柔比星在小鼠体内药动学的影响

The Effect of Paclitaxel on Pharmacokinetics of Doxorubicin in Mice by UPLC-MS/MS

目的建立测定小鼠血浆和心、肝、脾、肺、肾以及肿瘤各组织中多柔比星(doxorubicin,DOX)及其活性代谢产物多柔比星醇(doxorubicinol,DOXol)含量的超高效液相色谱-串联质谱(UPLC-MS/MS)方法,并探究紫杉醇(paclitaxel,PTX)对DOX和DOXol的药动学影响。方法柔红霉素(daunorubicin,DAU)为内标,乙腈(A)-0.1%甲酸水(B)为流动相进行梯度洗脱:0 min 10% A,0.5 min 30% A,1.0 min 60% A,1.5 min 60% A,2.0 min 10% A,2.2 min 10% A。用于定量分析的离子分别为DOX m/z 544.25→396.96、DOXol m/z 546.25→399.00和DAU m/z 528.38→321.00。UPLC-MS/MS测定小鼠血浆和各组织中DOX和DOXol浓度,比较各组DOX和DOXol的体内药动学差异。结果小鼠血浆和组织中DOX和DOXoL在测定范围内,均呈现良好线性关系(r>0.99),日内和日间精密度RSD均<6.9%,准确度RE为±6.2%,提取回收率为93.1%~107.4%,基质效应均在85%~115%,该检测方法稳定可靠;各组织中的DOX和DOXol浓度为肾>肝>脾>肺>心>肿瘤,其中由于肿瘤中DOXol的浓度低于定量下限而未检测到。结论该方法专属性强、灵敏度高,可以用于DOX和DOXol药动学研究。PTX对DOX的小鼠体内分布有影响,对DOX和DOXol的心脏分布浓度无明显影响。

OBJECTIVE To establish an ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the determination of doxorubicin (DOX) and its active metabolite doxorubicinol (DOXol) in mouse plasma and heart, liver, spleen, lung, kidney and tumor tissues, and to explore the pharmacokinetic effects of paclitaxel (PTX) on DOX and DOXol. METHODS The daunorubicin (DAU) was used as the internal standard. The mobile phase consisted of acetonitrile (A)-0.1% formic acid water (B) for gradient elution:0 min 10% A, 0.5 min 30% A, 1.0 min 60% A, 1.5 min 60% A, 2.0 min 10% A, 2.2 min 10% A, and the separation was carried out on an ACQUTTY UPLC BEH C18 column(2.1 mm×50 mm, 1.7 μm). The transitions of m/z 544.25→396.96, m/z 546.25→399.00, m/z 528.38→321.00 were used to quantify DOX, DOXol, and DAU. UPLC-MS/MS method was used to determine the concentration of DOX and DOXol in plasma and tissues of Balb/C mice at different times in the three groups. The pharmacokinetic differences of DOX and DOXol were compared between the single administration group and the combined administration group. RESULTS DOX and DOXoL in the plasma and tissues of mice showed a good linear relationship (r>0.99), the intra-and inter-day precision (RSD) was <6.9%, the accuracy (RE) was ±6.2%, the extraction recovery rate was 93.1%-107.4% and the matrix effect was 85%-115%. The detection method was stable and reliable. The concentration of DOX and DOXol in each tissue was kidney>liver>spleen>lung>heart>tumor, and DOXol was not detected because its concentration in the tumor was lower than the lower limit of quantitation. Compared with the DOX alone group, the concentrations of DOX and DOXoL in the plasma, liver and kidney were increased within 4 h after administration in the PTX→DOX group. CONCLUSION The method is highly specific and sensitive, and can be used for pharmacokinetic studies of DOX and DOXol. PTX affects the distribution of DOX in mice, but has no significant effect on the cardiac distribution of DOX and DOXol.