Pang Qian, Tian Yuanyuan, Mi Jianping, Wang Jin, Xu Yuanjin
State Key Laboratory of Conservation and Utilization of Subtropical Agro-bioresources, Guangxi University, Nanning 530004, China; School of Chemistry and Chemical Engineering, Guangxi University, Nanning 530004, China.
State Key Laboratory of Conservation and Utilization of Subtropical Agro-bioresources, Guangxi University, Nanning 530004, China; Beihai Center for Disease Control and Prevention, Beihai 536000, China.
J Pharm Biomed Anal. 2016 Jan 25;118:228-234. doi: 10.1016/j.jpba.2015.10.027. Epub 2015 Oct 26.
A rapid and sensitive assay based on ultra high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) was established and validated for the simultaneous determination of gallic acid, protocatechuic acid, vanillic acid, caffeic acid, epicatechin, isoquercitrin, vincetoxicoside B and quercetin in rat plasma using catechin and daidzein as the internal standards (IS). Plasma samples added internal standards were acidified with formic acid then pretreated by direct protein precipitation with acetonitrile. The separation of eight constituents was achieved on a C18 column with gradient elution using methanol and 0.2% acetic acid aqueous solution as the mobile phase and detected by multiple reaction monitoring using electrospray ionization source in the positive-negative ionization mode. The method was validated for sufficient specificity, precision, accuracy, and sensitivity over the concentration range of 10-6000 ng mL(-1) for gallic acid, 1.5-3000 ng mL(-1) for protocatechuic acid, 10-15000 ng mL(-1) for vanillic acid, 2-3600 ng mL(-1) for caffeic acid, 1.5-3600 ng mL(-1) for epicatechin, 4-6000 ng mL(-1) for isoquercitrin, 2-9000 ng mL(-1) for vincetoxicoside B, and 20-18000 ng mL(-1) for quercetin. The overall intra‑run precision and the inter‑run precision were showed in the range of 1.0-14.2% and 2.8-12.9%, respectively, and the accuracy was no more than 12.8%. This analytical method was successfully applied to investigate the pharmacokinetics of eight ingredients in rats after oral administration of Hypericum japonicum Thunb extract.
建立了一种基于超高效液相色谱串联质谱（UHPLC-MS/MS）的快速灵敏测定方法，以儿茶素和大豆苷元为内标（IS），同时测定大鼠血浆中的没食子酸、原儿茶酸、香草酸、咖啡酸、表儿茶素、异槲皮苷、罗布麻苷B和槲皮素。加入内标的血浆样品用甲酸酸化，然后用乙腈直接进行蛋白沉淀预处理。在C18柱上以甲醇和0.2%乙酸水溶液为流动相进行梯度洗脱，分离8种成分，并采用电喷雾电离源在正负离子模式下进行多反应监测检测。该方法在没食子酸浓度范围为10 - 6000 ng mL(-1)、原儿茶酸为1.5 - 3000 ng mL(-1)、香草酸为10 - 15000 ng mL(-1)、咖啡酸为2 - 3600 ng mL(-1)、表儿茶素为1.5 - 3600 ng mL(-1)、异槲皮苷为4 - 6000 ng mL(-1)、罗布麻苷B为2 - 9000 ng mL(-1)、槲皮素为20 - 18000 ng mL(-1)内具有足够的特异性、精密度、准确度和灵敏度。批内精密度和批间精密度分别在1.0 - 14.2%和2.8 - 12.9%范围内，准确度不超过12.8%。该分析方法成功应用于研究大鼠口服地耳草提取物后8种成分的药代动力学。
