Li Guo-Qing, Shu Ting, Chai Yuan-Yuan, Huang Xin, Jiang Zhen-Zhou, Zhang Lu-Yong
New Drug Research and Development Center, Guangdong Pharmaceutical University Guangzhou 510006, China.
New Drug Screening Center, Jiangsu Center for Pharmacodynamics Research and Evaluation,China Pharmaceutical University Nanjing 210009, China.
Zhongguo Zhong Yao Za Zhi. 2022 Aug;47(15):4183-4189. doi: 10.19540/j.cnki.cjcmm.20220507.501.
This study aims to establish an ultra-high performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS) method for determining the concentrations of triptolide(TP) in plasma and liver, and to explore the toxicokinetics of TP and the relationship between TP exposure and liver injury in C57 BL/6 mice, so as to provide reference for dissecting the toxicity mechanism of TP. The liquid chromatography was conducted with ZORBAX SB-C_(18) column(3.0 mm×100 mm, 3.5 μm) and the mobile phase of methanol-0.05 mmol·L(-1) ammonium acetate. Electrospray ionization(ESI) and multiple reaction monitoring(MRM) mode were employed for mass spectrometry. After oral administration of TP(toxic dose 600 μg·kg(-1)), the blood and liver tissues of the C57 BL/6 mice were collected at different time points to measure the TP concentrations in plasma and liver tissues. Furthermore, the blood biochemical indexes, including alkaline phosphatase(ALP), alanine aminotransferase(ALT), aspartate aminotransferase(AST), and total bile acid(TBA), were determined. After being processed by DAS 2.0, the experiment data showed that the TP in mice had the toxicokinetic parameters of T_(max)=5 min, C_(max)=14.38 ng·mL(-1), t_(1/2)=0.76 h, AUC_(0-t)=5.63 h·ng·mL(-1), MRT_(0-t)=0.56 h, and CL_(Z/F)=103.19 L·h(-1)·kg(-1). The trend of TP concentration in mouse liver tissue was consistent with that in plasma. The concentration of TP peaked at the time point of 5 min and then decreased until TP was completely metabolized. The plasma biochemical indexes(ALT, AST, ALP, and TBA) showed no significant changes within 3 h after TP administration. TP had high clearance rate and short residence time and did not significantly increase the blood biochemical indexes in mice. The results suggested that the exposure amount of free TP in vivo cannot directly cause liver injury, which might be caused by the binding of TP to some substances or the stimulation of inflammation and immune response.
本研究旨在建立一种超高效液相色谱 - 串联质谱法（UHPLC - MS/MS）用于测定血浆和肝脏中雷公藤甲素（TP）的浓度，探讨C57 BL/6小鼠中TP的毒代动力学以及TP暴露与肝损伤之间的关系，为剖析TP的毒性机制提供参考。液相色谱采用ZORBAX SB - C₁₈柱（3.0 mm×100 mm，3.5 μm），流动相为甲醇 - 0.05 mmol·L⁻¹乙酸铵。质谱采用电喷雾电离（ESI）和多反应监测（MRM）模式。口服给予TP（毒性剂量600 μg·kg⁻¹）后，在不同时间点采集C57 BL/6小鼠的血液和肝脏组织，测定血浆和肝脏组织中TP的浓度。此外，还测定了血液生化指标，包括碱性磷酸酶（ALP）、丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）和总胆汁酸（TBA）。经DAS 2.0处理后，实验数据表明小鼠体内TP的毒代动力学参数为：Tₘₐₓ = 5 min，Cₘₐₓ = 14.38 ng·mL⁻¹，t₁/₂ = 0.76 h，AUC₀₋ₜ = 5.63 h·ng·mL⁻¹，MRT₀₋ₜ = 0.56 h，CL₍Z/F₎ = 103.19 L·h⁻¹·kg⁻¹。小鼠肝脏组织中TP浓度的变化趋势与血浆中一致。TP浓度在5 min时达到峰值，随后下降直至TP完全代谢。TP给药后3 h内血浆生化指标（ALT、AST、ALP和TBA）无明显变化。TP具有高清除率和短停留时间，且未显著升高小鼠血液生化指标。结果提示，体内游离TP的暴露量不能直接导致肝损伤，可能是由于TP与某些物质结合或炎症及免疫反应的刺激所致。
