藏药仁青常觉中砷在大鼠尿液中的HPLC-ICP-MS研究

目的：采用HPLC-ICP-MS法对藏药仁青常觉中砷（As）在大鼠体内代谢产物—尿液中的形态进行研究。方法：采用Dionex Ion Pac AS 19阴离子色谱柱，以2 mmol·L^-1磷酸二氢钠-0.2 mmol·L^-1乙二胺四乙酸二钠-10 mmol·L^-1无水乙酸钠-3 mmol·L^-1硝酸钾（pH=10.5~11.3）-无水乙醇为流动相；流速1.0 mL·min^-1；进样量10μ L，柱温为室温。结果：实验表明，该方法不受⁴⁰Ar³⁵Cl⁺干扰，常见的5种As形态化合物的线性范围为0.5~100μ g·L^-1，甜菜碱（AsB）、二甲基胂（DMA）、三价砷［As（Ⅲ）］、一甲基胂（MMA）及五价砷［As（Ⅴ）］的定量下限分别为0.07、0.34、0.135、0.13、0.2μ g·L^-1，相关系数（R²）均大于0.999。通过对尿液中As总量及As形态进行分析，结果As总量在给药第15天达到最大值，主要As形态化合物为AsB、DMA、As（Ⅲ）和As（Ⅴ）；在第26天即停药后第11天As总量降到最低，主要As形态化合物为AsB、DMA和As（Ⅴ）。仁青常觉中As在大鼠体内的代谢研究表明：As（Ⅲ）通过大鼠给药吸收进入体内，后在大鼠体内被部分氧化为As（Ⅴ），经过生物甲基化反应，转化为DMA，最终通过尿液排出体外。4种As形态在大鼠尿液中的排泄速度以DMA最快。结论：藏药仁青常觉中As在大鼠体内代谢产物—尿液中的形态分析的研究，为藏药中As的毒理学研究提供了依据。

Objective: To establish an HPLC-ICP-MS method for the study of the metabolic fate of arsenic speciation in urine metabolites of Renqing Changjue in rats. Methods: Analysis was performed on a Dionex Ion Pac AS19 anion chromatographic column, with 2 mmol·L^-1 dihydrogen phosphate dihydrogen -0.2 mmol·L^-1 disodium tetraacetic acid disodium -10 mmol·L^-1 sodium acetate -3 mmol·L^-1 potassium nitrate (pH=10.5-11.3) -ethyl alcohol as the mobile phase. The flow rate was 1.0 mL·min^-1, the injection volume was 10μL, and the column temperature was room temperature. Results: The experiments showed that the method was not affected by ⁴⁰Ar³⁵Cl⁺ interference. The linears of AsB, DMA, As(Ⅲ), MMA and As(Ⅴ) were in the range of 0.5-100 μg·L^-1 and the limits of quantitation were 0.07,0.34, 0.135, 0.13 and 0.2μg·L^-1, respectively. The correlation coefficients(R²) were above 0.999. The results of repeatability, accuracy and precision test accorded with the requirements of the measurement. Through the analysis of the content of total arsenic and arsenic speciation in urine samples,the results showed that the total amount of arsenic reached the maximum value on the 15th day. And the mainly arsenic compounds were presented as AsB, DMA, As(Ⅲ) and As(Ⅴ). On the 26th day(the 11th day after discontinuation),urine arsenic reduce to a minimum content. And the major compounds were observed as AsB、DMA and As (Ⅴ). The study of arsenic speciation in vivo rat metabolites indicated that As (Ⅲ) in Renqing Changjue was absorbed into the body, then was partial oxidized to As(Ⅴ), finally was transformed into DMA through biological methylation reaction. Comparing with the other forms of arsenic in rat urine,the DMA had the fastest excretory speed. Conclusion: This study of arsenic speciation in the urine metabolites of rats can provide basis for the research of arsenic toxicology in Tibetan medicine.

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