Zhao Yi-Ning, Shi Rong, Zan Bin, Li Yuan-Yuan, Wang Tian-Ming, Liu Shao-Yong, Yang Li, Ma Yue-Ming
School of Pharmacy, Shanghai University of Traditional Chinese Medicine Shanghai 201203, China.
Shanghai Kai Bao Pharmaceutical Co., Ltd. Shanghai 200120, China.
Zhongguo Zhong Yao Za Zhi. 2021 Oct;46(20):5372-5381. doi: 10.19540/j.cnki.cjcmm.20210706.205.
Due to the limited resource of bear bile powder, the major raw material of Tanreqing Capsules(TRQ), cultured bear bile powder is used as a replacement to develop the Tanreqing Capsules Substitute(TRQS). An LC-MS/MS method was established in this study for simultaneous quantitation of 8 compounds from TRQS in rat plasma: tauroursodeoxycholic acid(TUDCA), taurocheno-deoxycholic acid(TCDCA), ursodeoxycholic acid(UDCA), chenodeoxycholic acid(CDCA), ferulic acid, wogonoside, baicalin, and forsythoside A. Thereby, the pharmacokinetic behaviors of TRQ and TRQS were evaluated. Concentration of endogenous compounds TUDCA, TCDCA, UDCA, and CDCA was determined with the stable isotope surrogate analytes: D4-TUDCA, D4-TCDCA, D4-UDCA, and D4-CDCA. Plasma samples were extracted by acetonitrile-induced protein precipitation. The LC conditions are as follows: Waters BEH C_(18) column(2.1 mm×100 mm, 1.7 μm), mobile phase of 10 mmol·L(-1) ammonium formate aqueous solution(containing 0.01% formic acid) and acetonitrile-methanol mixture(1∶5). MS conditions are as below: multiple reaction monitoring(MRM), ESI(+/-). Concentration of UDCA, CDCA, TUDCA, and TCDCA was corrected with a response factor, which is the ratio between the responses recorded for the surrogate and the authentic analyte at the equal concentration. Each of the plasma components showed good linearity(r > 0.995 1). Accuracy and precision met the criteria(inter-day RSD<7.0%, RE 89.98%-112.0%; intra-day RSD<12%, RE 90.41%-111.2%). The recovery was 64.83%-119.9% and matrix effect was 87.15%-113.8%. The validated method was applied for pharmacokinetic study of TRQS and TRQ(po, 0.94 g·kg~(-1)). There was no significant difference in C_(max) and AUC_(0-24 h) of baicalin, UDCA, TUDCA, and TCDCA between the two groups, indicating similar pharmacokinetic behaviors between TRQS and TRQ in rats.
由于痰热清胶囊（TRQ）的主要原料熊胆粉资源有限，故采用人工养殖熊胆粉作为替代原料研制痰热清胶囊替代物（TRQS）。本研究建立了一种液相色谱 - 串联质谱（LC - MS/MS）法，用于同时定量大鼠血浆中TRQS的8种成分：牛磺熊去氧胆酸（TUDCA）、牛磺鹅去氧胆酸（TCDCA）、熊去氧胆酸（UDCA）、鹅去氧胆酸（CDCA）、阿魏酸、汉黄芩苷、黄芩苷和连翘酯苷A。据此，对TRQ和TRQS的药代动力学行为进行了评价。内源性化合物TUDCA、TCDCA、UDCA和CDCA的浓度用稳定同位素替代分析物D4 - TUDCA、D4 - TCDCA、D4 - UDCA和D4 - CDCA进行测定。血浆样品通过乙腈诱导的蛋白沉淀法进行提取。液相色谱条件如下：Waters BEH C₁₈柱（2.1 mm×100 mm，1.7 μm），流动相为10 mmol·L⁻¹甲酸铵水溶液（含0.01%甲酸）和乙腈 - 甲醇混合液（1∶5）。质谱条件如下：多反应监测（MRM），电喷雾电离（ESI⁺/⁻）。UDCA、CDCA、TUDCA和TCDCA的浓度用响应因子进行校正，响应因子为替代分析物与等浓度真实分析物记录响应的比值。各血浆成分均显示出良好的线性关系（r＞0.995 1）。准确度和精密度符合标准（日间相对标准偏差＜7.0%，相对误差89.98% - 112.0%；日内相对标准偏差＜12%，相对误差90.41% - 111.2%）。回收率为64.83% - 119.9%，基质效应为87.15% - 113.8%。该验证方法应用于TRQS和TRQ（口服，0.94 g·kg⁻¹）的药代动力学研究。两组间黄芩苷、UDCA、TUDCA和TCDCA的Cmax和AUC₀ - 24 h无显著差异，表明TRQS和TRQ在大鼠体内的药代动力学行为相似。
