成分分析

SPE-HPLC测定川乌炮制过程中6个乌头类生物碱动态变化及其毒性分析*

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  • 1.湖北科技学院, 咸宁 437000;
    2.湖北通山县第一人民医院, 咸宁 437000
第一作者 毕健丽 Tel: 15512151802; E-mail: 3192999525@qq.com
陈婷 Tel: 15872827994; E-mail: 274435228@qq.com
**范宝磊 Tel: 18986638283; E-mail: fanbl_1980@163.com
王琦 Tel: 13872190370; E-mail: 594230783@qq.com

收稿日期: 2020-10-23

  网络出版日期: 2024-07-12

基金资助

*湖北省食品药品监督管理局重点项目(20180103); 湖北省卫生健康委员会中医药面上项目(ZY2019M029); 咸宁市自然科学基金(2020ZRKX05); 湖北科技学院五官专项重点项目(2020WG04)

SPE-HPLC determination of the processing process of Aconiti Radix dynamic changes and toxicity analysis of 6 aconitine alkaloids*

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  • 1. Hubei University of Science and Technology, Xianning 437000, China;
    2. The First People's Hospital of Tongshan County, Xianning 437000, China

Received date: 2020-10-23

  Online published: 2024-07-12

摘要

目的:进一步优化固相微萃取和高效液相色谱(SPE-HPLC)结合测定川乌炮制品中6个乌头类生物碱的方法。进行口服制川乌炮制品提取物后肺和肾的病理切片分析, 确定川乌炮制的最适方法和时间。方法:采用强阳离子吸附树脂MCX 固相萃取小柱对川乌炮制品进行纯化, 高效液相色谱法测定, 流动相为0.01 mol·L-1 甲酸铵水溶液(甲酸0.1%)-乙腈(32% 四氢呋喃), 梯度洗脱, 流速为1.0 mL·min-1, 检测波长为235 nm。HE 染色观察大鼠口服川乌炮制品提取物后肺、肾组织的形态学变化。结果:6个乌头类生物碱在优化条件下均可有效分离, 苯甲酰新乌头原碱、苯甲酰乌头原碱、苯甲酰次乌头原碱、新乌头碱、次乌头碱和乌头碱分别在各自线性范围内呈良好的线性关系, 加样回收率在98.1%~104.0%之间, RSD 均<5.0%。川乌炮制时采用蒸制法, 在7 h 时, 单酯型生物碱总量达到最大。HE 染色观察大鼠口服川乌炮制品提取物后肺、肾组织的形态产生变化。肺泡间质水肿导致弥漫性变化, 大量炎性细胞浸润, 肺泡壁增厚。肾小球体积增大, 球内细胞核相对减少, 肾小球与肾小囊间距变小, 管腔之间无空隙, 且管腔变大。结论:本实验采用强阳离子交换树脂MCX 和高效液相色谱法结合, 测定川乌炮制品中乌头类生物碱动态变化, 确定川乌最适炮制方法为蒸制法, 时间为7 h, 方法操作简便, 时间短, 测定结果准确, 可为控制川乌中毒性成分和有效成分的含量提供科学依据。

本文引用格式

毕健丽, 陈婷, 金文芳, 徐胜, 屈曾义, 饶梦凡, 蔡杏, 王琦, 范宝磊 . SPE-HPLC测定川乌炮制过程中6个乌头类生物碱动态变化及其毒性分析*[J]. 药物分析杂志, 2021 , 41(8) : 1389 -1398 . DOI: 10.16155/j.0254-1793.2021.08.13

Abstract

Objective:To further optimize the analysis method of alkaloids in Aconiti Radix preparata, solid phase microextraction combined with high performance liquid chromatography (SPE-HPLC) was used to determine the 6 aconitine alkaloids in Aconiti Radix preparata products. Pathological section of lung and kidney after oral administration of processed Aconiti Radix extract was analyzed, so as to determine the optimum processing method and time. Methods:The processed Aconiti Radix was purified by solid-phase extraction using a strong cationic adsorption resin MCX column. The mobile phase was 0.01 mol·L-1 ammonium formate aqueous solution (formic acid 0.1%) -acetonitrile (32% tetrahydrofuran) with gradient elution, the flow rate was 1.0 mL·min-1 and the detection wavelength was 235 nm. HE staining was used to observe the morphological changes of lung and kidney tissues after oral administration of Aconiti Radix processed product extract in rats. Results:The six aconitine alkaloids could be effectively separated under the optimized conditions. The benzoylmesaconine, benzoylaconine, benzoylhypaconine, mesaconitine, hypaconitine and aconitine had a good linear relationship within their respective linear range. The recovery rate of spiked samples was between 98.1%-104.0%, and RSD<5.0%. Aconiti Radix was processed by steaming and the amount of monoester alkaloids reached the maximum at 7 h. HE staining was used to observe the morphological changes of lung and kidney tissues after oral administration of extracts from Aconiti Radix in rats. The alveolar interstitial edema led to diffuse change, a large number of inflammatory cell infiltration and thickening of the alveolar wall. The glomerulus increased in size, and the nuclei in the glomerulus was relatively reduced. The space between the glomerulus and renal vesicles became smaller, and there was no space between the lumens, which became larger. Conclusion:The dynamic changes of aconitine alkaloids in Aconiti Radix preparata products were determined by MCX with strong cation exchange resin and high performance liquid chromatography in this experiment. The best processing method of Aconiti Radix is steaming for 7 h. The method is simple, short in time and accurate in determination, which could provide scientific basis for controlling the content of toxic components and active components in Aconiti Radix preparata products.

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