HPLC测定大鼠全血中虫草素药代动力学方法的建立  

HPLC Optimization for the Determination of the Pharmacokinetics of Cordycepin in Whole Blood of Rats

作　　者：吴灵玉 王雪艳[2] 陈志宝[1] 徐红月 李燕[3] 唐旭东[2] 于录 Wu Lingyu;Wang Xueyan;Chen Zhibao;Xu Hongyue;Li Yan;Tang Xudong;Yu Lu(College of Life Science and Technology,Heilongjiang Bayi Agricultural University,Daqing 163319;Key Laboratory of New Drug Research and Development,Research Institute of Tsinghua University;Key Laboratory of Zoonosis,Ministry of Education of Jilin University)

机构地区：[1]黑龙江八一农垦大学生命科学技术学院,大庆163319 [2]深圳清华大学研究院新药研发重点实验室 [3]吉林大学人兽共患病研究教育部重点实验室

出　　处：《黑龙江八一农垦大学学报》2018年第6期69-74,共6页journal of heilongjiang bayi agricultural university

基　　金：深圳市科学技术创新委员会项目(JSGG20160301100442775);吉林省科技发展计划项目(20150101108JC);吉林省教育厅项目(No.2016444)

摘　　要：建立一种灵敏、稳定的检测血液中虫草素及其代谢物的高效液相色谱方法,并报告采用该方法进行药代动力学的初步研究。大鼠全血0.5 mL,加入甲醇1 mL,涡旋振荡2 min,超声10 min,4℃条件下10 000 r·min^(-1)离心15 min,取上清室温条件下氮气吹干,用体积分数为0.06的乙腈溶液0.5 mL复溶,4℃条件下15 000 r·min^(-1)离心10 min,取上清过0.22μm微孔滤膜进行HPLC检测;色谱条件为Agilent TC-C18色谱柱(4.6 mm×250 mm,5μm),流动相为乙腈/水,6:94(V/V),流速0.8 mL·min^(-1),柱温30℃,紫外检测波长259 nm。虫草素在0.1～5.0μg·mL-1浓度范围内线性关系良好,线性回归方程为Y=10.97X-11.73,相关系数r=0.998 0血液样品中虫草素在高、中、低浓度的回收率分别为99.63%、95.37%和93.26%。全血中虫草素的药物代谢动力学初步研究的结果为给药后10～120 min内均未检测到虫草素原型,相同色谱条件下,保留时间为8.8 min左右检测到一虫草素的未知代谢物,且该物质在30 min时含量最高,随后逐渐降低。对该产物富集后通过质谱检测得未知代谢物分子量为252 M。该方法灵敏度高,重现性好,可用于大鼠全血中虫草素的代谢研究;研究结果同时表明,虫草素在大鼠体内代谢速度快,按40 mg·kg^(-1)剂量灌胃,即使灌胃给药后10 min也未检测到虫草素原型。It was aimed to establish a sensitive and stable HPLC method for the determination of Cordycepin and its metabolites in blood and to report the preliminary study of pharmacokinetics.The rat whole blood0.5mL was added with1mL of methanol,vortexed for2min,sonicated for10min and centrifuged at10000r·min-1for15min at4℃,the supernatant was dried under nitrogen at room temperature with volume fraction of0.06in acetonitrile of0.5mL reconstituted,centrifuged at15000r·min-1at4℃for10min,the supernatant was filtered through a0.22μm microporous membrane for HPLC detection;the chromatographic conditions were Agilent TC-C18column(4.6mm×250mm,5μm).The mobile phase consisted of acetonitrile/water,6:94(V/V),the flow rate was0.8mL·min-1,the column temperature was30℃and the UV detection wavelength was259nm.Cordycepin had a good linearity in the range of0.1~5.0μg·mL-1with a linear regression equation of=10.97-11.73and a correlation coefficient of=0.9980.The recoveries of Cordycepin in the blood samples were99.63%,95.37%and93.26%at high,medium and low concentrations,respectively.Using this condition,the pharmacokinetics of Cordycepin in whole blood were studied preliminarily:no prototype of Cordycepin was detected within10~120min after administration,and the retention time was8.8min under the same chromatographic conditions Cordycepin unknown metabolites,and the substance at30min when the highest content,then gradually decreased.The mass of the unknown metabolite detected by mass spectrometry after enrichment of this product was252M.The method had high sensitivity and good reproducibility and could be used to study the metabolism of Cordycepin in rat whole blood.The results also showed that Cordycepin was metabolized rapidly in rats at a dose of40mg·kg-1,even if Cordycepin was not detected at10min after intragastric administration.

关 键 词：虫草素 全血 HPLC 

分 类 号：R917[医药卫生—药物分析学]