柴贝止痫汤在大鼠体内主要入血成分及其药代动力学研究

Study on the main components absorbed into blood of Chaibei Zhixian decoction in rats and their pharmacokinetics

目的借助于高效液相色谱(Ultra Performance Liquid Chromatography,UPLC)-串联质谱(Tandem mass spectrometry,MS/MS)技术以及气相色谱(Gas Chromatography,GC)-MS/MS技术确定大鼠灌服柴贝止痫汤后主要吸收入血成分,并计算其在大鼠体内的药代动力学特征。方法采用Thermo Syncronis C18(2.1×100 mm,1.9μm)为色谱柱,乙腈-0.01%甲酸水为流动相,分析时间0~8分钟,进样量5μL,流速0.25 mL/min;电喷雾电离源ESI(+),监测模式为二级数据依赖性扫描;GC载气为He气,恒流模式,流速为1 mL/min,进样量1μL,气化室温度250℃,柱温为程序升温,起始温度为80℃,维持1分钟。雄性Wistar大鼠6只,给予中药柴贝止痫汤(生药8.125 g/kg)灌胃,分别于灌胃前及灌胃后各时间点眶静脉取血,样品处理后采用液质联用及气质联用技术检测17种单体成分在不同时间点大鼠血液中的浓度,采用DAS 2.0软件计算药代动力学参数:AUC_(0-t)、AUC_(0-∞)、MRT_(0-t)、MRT_(0-∞)、t 1/2、T max、C max。结果大鼠血浆中检测出15种单体成分,其含量由高到低排列依次为:天麻素、对羟基苯甲醛、胡芦巴碱、天麻苷元、原儿茶醛、柴胡皂苷A、香草醛、细辛醛、贝母素乙、α-细辛醚、茴香脑、β-细辛醚、柴胡皂苷C、贝母素甲、贝母辛,柠檬醛及柴胡皂苷D未检测出。柴贝止痫汤整体入血成分血药浓度在0.25~0.5小时达到高峰,半衰期平均值为4.28小时。结论该方法日内精密度和日间精密度良好,回收率在标准范围内,符合定量检测要求;柴贝止痫汤灌胃后贝母素甲、贝母素乙等15种化合物可吸收入血,0.25~0.5小时入血成分浓度达到高峰,可作为复方含药血清最佳采血时间。

Objective To determine the main absorbed into the blood components of Chaibei Zhixian decoction by using ultra performance liquid chromatography-tandem mass spectrometry(UPLC/MS)technology,and to calculate its pharmacokinetics characteristics in rats.Methods Thermo Syncronis C18(2.1×100 mm,1.9 m)was used as the chromatographic column,acetonitrile-0.01%formic acid water was used as mobile phase,the analysis time was 0~8 minutes,the injection volume was 5μL,and the flow rate was 0.25 mL/min,the gasification chamber temperature is 250℃,column temperature is program heating,the starting temperature is 80℃for 1 minute.6 male Wistar rats were given Chaibei Zhixian decoction(dried medicine herbs 8.125 g/kg)by gavage.Blood was collected from the orbital vein before and after gavage.After sample treatment,the concentrations of 17 monomer components in rat blood at different time points were detected by lC-MS and GC-MS.DAS 2.0 software was used to calculate pharmacokinetic parameters,including AUC_(0-t),AUC_(0-∞),MRT_(0-t),MRT_(0-∞),t 1/2,T max and C max.Results 15 monomer components were detected in rat plasma,and the order of their contents from high to low was gastrodin,p-hydroxybenzaldehyde,trigonelline,p-hydroxybenzoic alcohol,protocatechualdehyde,saikosaponin A,vanillin,asarylaldehyd,verticinone,α-asarone,anethole,β-asarone,saikosaponin C,verticine,peimisine.Citral and saikosaponin D were not detected at any time points.The blood concentration of the whole component absorbed into blood in Chaibei Zhixian decoction peaked at 0.25~0.5 hours,and the average half-life was 4.28 hours.Conclusion The method has good within-day precision and day to day precision,and the recovery rate is within the standard range,meeting the quantitative detection requirements.15 compounds,such as fritillary a and fritillary b,could be absorbed into the blood after Chaibei Zhixian decoction was given by gavage,and the concentration of components absorbed into blood reached the peak at 0.25~0.5 hours,which can be used as the best time for blood collection of compound drug-containing serum.