HPLC―MSMS同时测定广金钱草中8种成分在大鼠胆汁中浓度及其排泄动力学研究.docVIP

HPLC―MSMS同时测定广金钱草中8种成分在大鼠胆汁中浓度及其排泄动力学研究 　　[摘要] 建立同时测定灌胃给予大鼠广金钱草提取物后胆汁中7种黄酮和1种酚酸类化合物浓度的HPLC-MS/MS，并用于胆汁排泄研究。大鼠灌胃给予广金钱草提取物后，分别在0～1，1～2，2～4，4～6，6～8，8～12，12～24 h时间段收集胆汁。色谱柱采用Diamonsil C18 column （4.6 mm×150 mm，5 μm），柱温 40 ℃。以甲醇-0.01%乙酸水溶液为流动相，梯度洗脱，流速0.8 mL?min-1。质谱采用电喷雾离子源（ESI），负离子模式下检测，多反应离子监测（MRM）模式进行定量。结果表明8种成分在测定浓度范围内线性关系良好（r≥0.991 5），日内、日间精密度RSD63.2%，内标归一化的基质因子RSD均未超过15%，符合要求。大鼠胆汁排泄研究结果显示，8种成分在胆汁中的原型排泄率均比较小，并且不同个体之间差别较大。该研究方法灵敏度高，选择性好，样品处理简单，可用于灌胃给予大鼠广金钱草提取物后胆汁中8种成分的排泄动力学研究。 　　[关键词] 广金钱草； 黄酮； 酚酸； HPLC-MS/MS； 含量测定； 胆汁排泄 　　[Abstract] To establish a HPLC-MS/MS quantitative method for analysis of 8 constituents in rat bile. The method was applied in the biliary excretion study after oral administration of Desmodii Styracifolii Herba extract. The rat bile samples were collected during 0-1， 1-2， 2-4， 4-6， 6-8， 8-12， 12-24 h. Diamonsil C18 column （4.6 mm×150 mm， 5 μm） was adopted and eluted with methanol and 0.01% acetic acid in a gradient mode. The flow rate was 0.8 mL?min-1， and the column temperature was set at 40 ℃. The detection was carried out by a triple quadrupole linearion trap mass spectrometer in the negative ion mode with an electrospray source. Multiple reactions monitoring （MRM） mode was employed. The calibration curves showed a good linearity， with correlation coefficients （r≥0.991 5） for all of the analytes within the concentration range. The intra-day and inter-day precisions （RSD） were both less than 15%， and the accuracies （RE） ranged from -15% to 15%. The method had a good stability， and was suitable for the content determination of 8 constituents in rat bile. The extraction recoveries of the 8 analytes were more than 63.2%， and the RSD of IS-normalized matrix factors were no more than 15%， which met the requirements for analysis. According to the biliary excretion study， the bile excretion rates of the 8 analytes were relatively low， with great difference among the individuals. The results showed that the established method was simple， selective a