黄体酮缓释制剂在Beagle犬中的血药浓度测定方法的建立及其药代动力学研究  

Development of a method for determination of progesterone sustained-release formulation in Beagle dog plasma and its pharmacokinetic study

作　　者：王玥 程艺 张慧 郑爱萍 WANG Yue;CHENG Yi;ZHANG Hui;ZHENG Ai-ping(School of Pharmacy,Xuzhou Medical University,Xuzhou 221000,China;Institute of Pharmacology and Toxicology,Academy of Military Medical Sciences,Academy of Military Sciences,Beijing 100850,China)

机构地区：[1]徐州医科大学药学院,江苏徐州221000 [2]军事科学院军事医学研究院毒物药物研究所,北京100850

出　　处：《药学学报》2022年第7期2191-2196,共6页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(81573357)。

摘　　要：本研究建立了高效液相色谱串联质谱法(LC-MS/MS),测定Beagle犬体内黄体酮的血药浓度,并进行黄体酮缓释微球在Beagle犬体内药代动力学的研究。以醋酸甲地孕酮为内标,蛋白沉淀法处理血样。采用电喷雾离子化源(ESI)以多反应监测(MRM)方式进行正离子检测,监测离子对分别为黄体酮m/z 315.2→97.0,内标醋酸甲地孕酮m/z 385.2→267.1。雄性Beagle犬肌注黄体酮缓释微球,于给药后不同时间点采集静脉血,结合WinNonlin 8.1软件计算相关药代动力学参数。在0.1~500.0 ng·mL^(-1)内,黄体酮的线性关系良好(r^(2)=0.9989);验证样品的批内、批间精密度(RSD)小于13.25%,准确度(RE)在8.92%以内;样品在室温放置12 h、-20℃反复冻融3次、-20℃放置60 d条件下均稳定;提取回收率为71.43%~77.97%。Beagle犬肌肉注射黄体酮缓释微球后,t_(max)为19.00±25.36 h,C_(max)为137.72±11.59 ng·mL^(-1),t_(1/2)为83.83±26.43 h。药物在体内持续释放,且呈多次连续吸收过程,具有良好的缓释效果。本研究所建立的LC-MS/MS测定方法灵敏、快速、稳定,适用于Beagle犬中黄体酮血药浓度的测定,可应用于黄体酮相关制剂的临床前药代动力学的研究。动物实验方案已获得军事医学研究院动物伦理委员会的批准。A liquid chromatography-tandem mass spectrometry(LC-MS/MS)method was developed to determine the plasma concentration of progesterone in Beagle dogs,and apply it to the study of the pharmacokinetics of progesterone sustained-release formulation in Beagle dogs.The plasma samples were processed by protein precipitation method and megestrol acetate was used as an internal standard(IS).The quantitation analysis was performed using multiple-reaction monitoring(MRM)mode at the specific ion transitions of m/z 315.2→97.0 for progesterone and m/z 385.2→267.1 for megestrol acetate(IS)under the positive ion condition.Male Beagle dogs were injected intramuscularly with progesterone sustained-release microspheres and the plasma samples were collected at different time points after administration.The relevant pharmacokinetic parameters were calculated by WinNonlin 8.1 software.A good linearity over the range of 0.1−500.0 ng·mL^(-1) was yielded by this method.The intra-and inter-day precision(RSD)were all less than 13.25%and the accuracy(RE)was within 8.92%.Stability test showed that progesterone in dog plasma was stable at room temperature for 12 h,up to 60 days at−20℃and after three cycles of freeze-thaw.The recovery of it ranged from 71.43%−77.97%.After intramuscular injection of progesterone sustained-release microspheres in Beagle dogs,t_(max)was 19.00±25.36 h,C_(max) was 137.72±11.59 ng·mL^(-1),t_(1/2) was 83.83±26.43 h.The drug was released continuously in vivo and in a continuous absorption process for many times with good sustained-release effect.The method developed in this study is sensitive,rapid and stable.It is suitable for the determination of progesterone plasma concentration in Beagle dogs,and can be applied to the preclinical pharmacokinetic study of progesterone-related formulations.The animal experiment scheme of this study was approved by the Animal Ethics Committee of the Academy of Military Medical Sciences.

关 键 词：黄体酮 高效液相色谱串联质谱法 BEAGLE犬 缓释微球 药代动力学 

分 类 号：R917[医药卫生—药物分析学]