世界中医药

目的：评价大鼠体内葛根素浓度及药代动力学，为临床葛根素用药方案制定选择提供参考。方法：建立葛根素在大鼠体内浓度测定方法，采用该方法进行葛根素药代动力学实验室研究，分析文献报道的高效液相色谱（HPLC）分析法检测葛根素含量方法，制定色谱条件：色谱柱：AcclaimC18色谱柱（5 μm，4.6 mm×150 mm），流动相：甲醇：水，检测器：Modle 500 UV检测器；检测波长：250 nm，色谱工作站：N2000双通道工作站。配制葛根素标准液、外标物溶液、待测血浆样品液、空白对照液等。分别进样行梯度洗脱，验证方法的专属性、线性关系、回收率、精密度、稳定性。选择12只洁净大鼠，雌雄各半，随机分为2组，将采用药典葛根素推荐剂量下限给予腹腔灌胃设为低剂量组，给予药典推荐剂量上线腹腔灌胃设为高剂量组。2组大鼠分别于灌胃后不同时间采集眼眶静脉血，采用已验证的HPLC分离检测方法检测血浆葛根素药物浓度，采用DAS2.0统计软件处理葛根素浓度-时间数据，绘制药时曲线图，计算药代动力学指标并行组间比较。结果：1）拟定色谱条件下，大鼠血浆空白溶液、葛根素对照溶液、葛根素血浆样品溶液中主要成分保留时间差异明显，未发生重叠，说明该方法专属性较好。2）加入不同浓度葛根素对照品于大鼠血浆中，所得浓度-峰面积呈良好的线性关系，回归方程为：Y=3.957×106X-697.329，r=0.998 3，检测限：0.055~0.660 μg/mL。不同浓度葛根素对照品回收率为（103.110±0.521）%，高浓度和低浓度2组回收率比较,差异无统计学意义（P>0.05）。日内精密度为：3.14%，日间精密度5.26%；12 h内，放置不同时间的样品间葛根素含量检测相对偏差（RSD）为3.95%；不同灌胃剂量大鼠药时曲线均符合二室模型;大鼠灌胃葛根素后，不同剂量大鼠的AUC0-t、Cmax、MRT0-t、CL差异有统计学意义（P<0.05）。结论：采用HPLC检测大鼠体内葛根素方法可行，采用低剂量用药方案即可获得较好的药物吸收代谢效果，高剂量则影响葛根素吸收。

English Summary:

To establish a method for concentration determination of puerarin in rats，and to study pharmacokinetics of puerarin in laboratory by this method，and to provide reference for formulation and selection of clinical puerarin medication scheme.Methods:Methods for determination of puerarin content by high performance liquid chromatography(HPLC)reported in literature were analyzed.Chromatographic conditions were established.Chromatographic column：Acclaim C18 chromatographic column(5 μm，4.6 mm×150 mm); mobile phase：methanol:water; detector：Modle 500 UV detector; detection wavelength：250 nm; chromatographic workstation：N2000 dual channel workstation.Puerarin standard solution，external standard solution，plasma sample solution for detection and blank control solution and other solutions were prepared.Gradient elution was used respectively.Specificity，linear relationship，recovery rate，precision and stability of the methods were validated.Twelve clean rats，half male and half female，were randomly divided into 2 groups.The group treated by intraperitoneal administration and gavage with lower limit of recommended dose of puerarin in the pharmacopoeia was taken as low dose group，and the group treated by intraperitoneal administration and gavage with upper limit of recommended dose of puerarin in the pharmacopoeia was taken as high dose group.Orbital venous blood was collected immediately，0.25 h，0.5 h，0.75 h，1 h，1.5 h，2 h，4 h，6 h and 8 h after the gavage.Plasma puerarin concentration was detected by the validated method of HPLC separation and detection.Concentration-time data of puerarin were processed by DAS 2.0 software.Drug-time curves were drawn and pharmacokinetic index was calculated and compared between the groups.Results:1)Under the proposed chromatographic conditions，retention time of main components in rat plasma blank solution，puerarin control solution and puerarin plasma sample solution was significantly different，and no overlap occurred，indicating that the method was specific.2)Different concentrations of puerarin control products were added into rat plasma，and there was a good linear relationship between the concentration and peak area.The regression equation was Y=3.957×106X-697.329，r=0.998 3; detection limit was 0.055～0.660 μg/mL.3)Recovery rate：the recovery rate of puerarin control products in different concentrations was(103.110±0.521)%，and there was no significant difference in the recovery rate between the high dose group and the low dose group(P>0.05).4)Precision：intraday precision was 3.14%，and day to day precision was 5.26%.5)Stability：within 12 h，relative deviation(RSD)of puerarin content between samples placed at different times was 3.95%.6)The drug-time curves of rats with different doses of gavage conformed to two-compartment model.7)After the gavage of puerarin，the AUC0-t，Cmax，MRT0-t and CL of rats with different doses were significantly different(P<0.05).Conclusion:It is feasible to detect puerarin in rats by high performance liquid chromatography.Good efficacy of drug absorption and metabolism can be achieved by using low dose medication scheme，while high dose can affect puerarin absorption.

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