HPLC-MS/MS测定硫酸长春新碱热敏脂质体及其比格犬药动学初步研究

Quantification of Vincristine Sulfate in Dog Plasma by HPLC-MS / MS and Pharmacokinetics of Vincristine Sulfate Thermosensitive Liposomes in Dogs

目的建立一种简便、快速、灵敏的测定比格犬血浆中硫酸长春新碱（vincristine sulfate，VCR）浓度的液相串联质谱（LC—MS／MS）法，并研究硫酸长春新碱热敏脂质体（VSTL）比格犬体内药动学特征。方法采用叔丁基甲醚对血浆进行提取，液相分离采用Agela Venusil XBP C18（2．1mm×30mm，3．0μm）分析柱，柱温30℃，以甲醇-水为流动相，采用梯度洗脱，流速为0．4mL·min^-1，进样量10μL；选用气动辅助电喷雾离子化（ESI），在正离子电离模式下，在多反应监测（MRM）的模式下检测离子对m／z825．4→807．2（VCR）和m／z811．3→223．9（硫酸长春碱）。两组比格犬分别静脉推注0．07mg·kg^-1。VsTL与硫酸长春新碱注射液，采用高效液相．串联质谱测定比格犬血浆中VCR浓度，计算主要药动学参数并比较研究比格犬注射VSTL与硫酸长春新碱注射液（VSU的药动学特征。结果血浆中无干扰测定的内源性物质，VCR在0．25—500ng·mL^-1内线性良好（r=0．9943），定量限为0．25ng·mL^-1，日内、日间精密度（RSD）均小于15％。VSTL和VSI的ρmax分别为（121．00±42．31）、（61±23．36）ng·mL^-1；t1/2λz分别为（23．95±9．03）、（37．91±8．02）h；CLz分别为（0．37±0．07）、（0．35±0．09）L·h·kg^-1；Vz分别为（12．15±2．14）mL·kg^-1、（18．95±3．27）L·kg^-1；AUC0-t分别为（144．87±1．10）、（127．7±2．45）ng·h·mL^-1。AUC0-∞分别为（152．97±12．56）、（131．61±13．22）ng·h·mL^-1。结论LC—MS／MS方法快速、准确、灵敏，适合于临床前药动学研究。静脉输注VSTL后的ρmax和AUC显著高于VSI，其他药动学参数没有显著差异。

OBJECTIVE To establish a rapid and sensitive liquid chromatography-tandem mass spectrometry（LC-MS/MS） meth- od for quantification of vincristine sulfate（VCR） in dog plasma and to study the pharmaeokinetics of vincristine sulfate thermosensitive liposomes（VSTL） in dogs. METHODS The plasma was extracted with tert-butyl methyl ether（TBME）. VCR and IS （ vinblastine sulfate） were separated on an Agela Venusil XBP Cl8 （2. 1 mm ×30mm,3.0μm） with a mobile phase gradient at a flow rate of 0. 4 mL · min^-1. The injection volume was 10 μL. An Agilent 6460A QQQ triple-quadrople mass spectrometer equipped with an electrospray ionization（ESI） source was used as detector and was operated in positive ion mode. Multiple-reaction monitoring（MRM） was performed and the m/z of ions selected for quantitation were m/z 825.4→807.2 （VCR） and m/z 811.3→223.9 （IS, vinblastine sulfate）. Dogs were injected VSTL and vincristine sulfate injection（VSI） via vein at the dose of 0. 07 mg · kg^-1 , respectively. VCR and internal standard were quantified in dog plasma using a high-performance liquid chromatography-tandem mass spectrometry, Pharmacokinetic parameters were calculated, and a comparative study of VSTL and VSI with six clogs was conducted. RESULTS The chromatograms showed no endogenous interfering peaks in blank dog plasma. The linear range of VCR in plasma was 0. 25 - 500 ng · mL^-1 （r = 0. 994 3 ）. The lower limit of quantification was 0. 25 ng · mL^-1. The intra-run and inter-run relative standard deviations （RSD） were less than 15%. The pharmacokinetic parameters of VSTL and VSI were as following：ρmax（ 121.00 ±42. 31 ） and （61 ±23.36） ng · mL^-1; t1/2λz（23.95 ±9.03） and （37.91 ±8.02） h; CL·（0. 37 ±0.07） and （0.35 ±0.09） L · h · kg^-1 ;Vz（12. 15 ±2. 14） and （18.95 ±3.27） L·kg^-1; AUC0-t（144.87±1.10） and （127.7 ±2.45） ng· h · mL^-1; AUC0-∞（152.97±12.56） ng· h· mL^- 1 and （ 131.61 ±13.22） ng · h · mL - 1. CONCLUSION The method is shown to be sensitive, accurate, and convenient for as saying the concentration of vincristine sulfate in preclinical pharmacokinetic studies. The ρmax and AUC of VSTL are significantly higher than VSI after intravenous infusion, the other pharmacokinetic parameters are no significant difference.