液质联用技术评估汞接触污染蟾蜍泌酥中蟾蜍甾烯水平

目的:运用液质联用技术研究重金属汞皮肤接触中华大蟾蜍后生理状态及泌酥中蟾蜍甾烯水平变化,为中药蟾酥品质评价提供参考。方法:配制不同质量浓度的氯化汞溶液(0.5,0.75,1.0 g·L~(-1))对蟾蜍进行半静水式处理,观察其生理状态,且在30 d后收集蟾蜍耳后腺分泌液,进一步采用液相-质谱联用技术测定蟾蜍泌酥中游离型蟾蜍甾烯的水平,结合Hem I热图软件进行数据分析。结果:氯化汞接触明显影响蟾蜍泌酥中蟾蜍甾烯水平,和正常组比较,低质量浓度(0.5 g·L~(-1))汞污染组有20种蟾蜍甾烯水平明显上调,其中有12个物质发生1.5倍变化。而中质量浓度(0.75 g·L~(-1)),高质量浓度(1.0 g·L~(-1))污染组分别有15,13种物质发生下调,且大部分为中低脂溶性蟾蜍甾烯(异沙蟾毒精、去乙酰华蟾毒精等)。结论:重金属汞污染对蟾蜍甾烯水平有潜在影响,且随着浓度的升高呈现明显降低趋势。

Assessing Mercury Pollution Impacts on Levels of Bufadienolides Based on LC-MS/MS

Objective: To investigate the effects of HgCl₂ pollution on the physiological status and bufadienolides level of toad by using liquid chromatography-mass spectrometry (LC-MS/MS), and provide references for quality evaluation of Chinese medicine venenum bufonis. Method: The different concentrations of HgCl₂ solution (0.5, 0.75 and 1.0 g·L^-1) were prepared to treat the toad with semi-static water method, and then their physiological status was observed. Venenum bufonis was collected after 30 days. Furthermore, ultra performance liquid chromatography-mass spectrometry was used to determine the level of bufadienolides in toad venom and HemI heat map software was used for data analysis. Result: The results showed that HgCl₂ solution could significantly influence the bufadienolides level. As compared with normal group, 20 kinds of bufadienolides were up-regulated in low concentration (0.5 g·L^-1) Hg²⁺ exposure group, and there were 12 kinds of bufadienolides with 1.5 fold change. However, 15 kinds of bufadienolides in 0.75 g·L^-1 Hg²⁺ exposure group and 13 kinds in 1.0 g·L^-1 Hg²⁺ exposure group were down-regulated, and most of them were medium and low fat-soluble toad steroids (bufarenogin, dehydrated cinobufagin, etc.). Conclusion: These illustrated that Hg²⁺ exposure could affect the levels of bufadienolides in toad venom, showing an obvious decrease tendency with the increase of HgCl₂ solution concentration.