UPLC-MS-MS同时测定比格犬血浆中阿魏酸、香草酸、芍药苷和芍药内酯苷的含量及其药动学研究

Simultaneous Determination of Ferulic Acid,Vanillic Acid,Peoniflorin and Albiflorin in Beagle Plasma and Pharmacokinetics by UPLC-MS-MS

目的:建立利用超高效液相质谱联用色谱(UPLC-MS-MS)同时测定比格犬血浆中阿魏酸、香草酸、芍药苷和芍药内酯苷的分析方法,并研究比格犬灌胃当归-白芍药对提取物后以上4种成分的药动学特征。方法:采集比格犬灌胃当归-白芍药对提取物后不同时间点的血浆样本,甲醇沉淀蛋白,色谱采用Waters BEH C18反相柱,流动相为水(含0.1%的甲酸)-乙腈梯度洗脱,采用电喷雾离子化三重四极杆串联质谱,以多反应监测(MRM)方式进行检测。正离子条件下用于定量分析的离子对分别为m/z 195.3～144.5(阿魏酸)、m/z 169.0～92.9(香草酸)、m/z 481.3～197.1(芍药内酯苷),负离子条件下用于定量分析的离子对为m/z 525.4～449.2(芍药苷)。DAS 2.0软件计算主要药动学参数。结果:阿魏酸、香草酸、芍药苷和芍药内酯苷分别在1.13～1 089,1.23～1 080,0.82～787.5,0.36～337.5μg.L-1线性关系良好,样品提取回收率、精密度、准确度及稳定性等均符合要求。阿魏酸和香草酸2种酚酸类成分在比格犬体内吸收均较迅速,阿魏酸消除较快,香草酸消除相对较慢;芍药苷和芍药内酯苷2种单萜苷类成分药动学特征相似,在比格犬体内吸收和消除均较慢。结论:UPLC-MS-MS法选择性好、灵敏度高、专属性强,可用于当归-白芍药对中主要化学成分的体内药动学研究,为当归-白芍配伍研究提供一定科学依据。

Objective： To develop a UPLC-MS-MS method to simultaneously determine the concentration of ferulic acid,vanillic acid,peoniflorin and albiflorin in beagle plasma and study the pharmacokinetics of these four components after ig administration of Angelicae Sinensis Radix-Paeoniae Radix Alba drug pair extract to beagle dogs.Method： Following a protein precipitation with methanol for plasma samples at different time after ig Angelicae Sinensis Radix-Paeoniae Radix Alba drug pair extract to Beagle dogs.The analysis was achieved by BEH C18 column and the mobile phase was consisted of water（0.1% formic acid included） and acetonitrile followed step gradient elution.A TQ tandem mass spectrometry equipped with electrospray ionization source was used as detector and operated by multiple reaction monitoring（MRM）.The ion pairs for quantitation were m / z 195.3-144.5 for ferulic acid,m / z 169.0-92.9 for vanillic acid,and m / z 481.3-197.1 for albiflorin in positive ion mode and m / z 525.4-449.2 for peoniflorin in negative ion mode.The pharmacokinetic parameters were calculated by DAS 2.0 program.Result： Ferulic acid,vanillic acid,peoniflorin and albiflorin showed a good liner relationship of 1.131 089,1.23-1 080,0.82-787.5,0.36-337.5 μg.L-1,respectively.The method was validated according to the requirements.The pharmacokinetic characteristics of ferulic acid and vanillic acid were similar,both of their absorption was very fast.Elimination of ferulic acid was fast while the elimination of vanillic acid was slower.The pharmacokinetic characteristics of peoniflorin and albiflorin were very similar,both of their absorption and elimination were slow in beagle dogs.Conclusion： The method is proved to be sensitive,selective,rapid and successfully applied to a pharmacokinetic study of major components of Angelicae Sinensis Radix-Paeoniae Radix Alba drug pair in beagle dogs.This research laid a good foundation for the study of compatibility of Angelicae Sinensis Radix-Paeoniae Radix Alba drug pair.