中国中药杂志

2020, v.45(17) 4158-4162

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中药黄曲霉毒素测定酶联免疫吸附法的研制
Development of enzyme linked immunosorbent assay of aflatoxin of Chinese herbal medicines

南铁贵;洪小栩;徐昕怡;黄璐琦;袁媛;
NAN Tie-gui;HONG Xiao-xu;XU Xin-yi;HUANG Lu-qi;YUAN Yuan;State Key Laboratory Breeding Base of Dao-Di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences;Chinese Pharmacopoeia Commission;

摘要(Abstract):

《中国药典》2020年版通则<2351>《真菌毒素测定法》中黄曲霉毒素测定法第三法采用了酶联免疫吸附法。该方法利用高通量单克隆抗体筛选技术,筛选得到了中药材专属性可仅特异性识别黄曲霉毒素B_1和可以特异性识别黄曲霉毒素B_1,B_2,G_1,G_2总量的2类特异性单克隆抗体。通过优化包被抗体、酶标抗原浓度,以及酶联免疫吸附法的反应体系,分别建立了可用于中药材、饮片及中药制剂中黄曲霉毒素B_1和黄曲霉毒素B_1,B_2,G_1,G_2总量测定的专属性酶联免疫吸附法。该方法在实际样品测定中回收率为60%~120%,相对标准偏差<15%。另外,针对现行《中国药典》中药材黄曲霉毒素测定前处理方法复杂、昂贵等问题,通过筛选和方法优化,建立了不依赖于免疫亲和柱的中药材黄曲霉毒素液液萃取前处理方法,该提取方法可以对中药材中的黄曲霉毒素进行高效提取。黄曲霉毒素酶联免疫吸附法作为中药黄曲霉毒素控制的有效方法,有效降低检测成本,便于操作,实现检测关口前移,有利于实现全生产过程的质量监管。
The enzyme linked immunosorbent assay of aflatoxin has been adopted in Chinese Pharmacopoeia(2020 edition). Based on high-throughput screening of monoclonal antibodies technology, monoclonal antibodies that can specifically recognize the aflatoxin B_1 and the total amount of aflatoxin B_1, B_2, G_1, and G_2 in Chinese herbal medicines were prepared. By optimizing the concentration of coating antibody, enzyme-labeled antigen, and the reaction system of enzyme-linked immunosorbent assay, the enzyme linked immunosorbent assay(ELISA) were developed for detection of aflatoxins in Chinese herbal medicines, decoction pieces, and preparation of Chinese medicine. In this method, the recovery test of actual samples is 60%-120%, and the relative standard deviation is less than 15%. In addition, in view of the complicated and expensive pretreatment methods for the determination of aflatoxin in Chinese herbal medicine, we developed a highly efficient pretreatment method of liquid-liquid extraction of aflatoxin in Chinese herbal medicine without immunoaffinity column. As an effective method for the detection of aflatoxin, the ELISA can effectively reduce the aflatoxins testing cost of traditional Chinese medicine, and promote the detection ability at earlier stages of production, and strengthen the quality supervision of traditional Chinese medicine.

关键词(KeyWords): 《中国药典》2020年版;黄曲霉毒素;酶联免疫吸附法
Chinese Pharmacopoeia(2020 edition);aflatoxin;enzyme linked immunosorbent assay

Abstract:

Keywords:

基金项目(Foundation): 国家药典委员会药品医疗器械审评审批制度改革专项(ZG2016-4-02);; 中央本级重大增减支项目(2060302);; 中央级公益性科研院所基本科研业务费专项(ZZ10-008)

作者(Author): 南铁贵;洪小栩;徐昕怡;黄璐琦;袁媛;
NAN Tie-gui;HONG Xiao-xu;XU Xin-yi;HUANG Lu-qi;YUAN Yuan;State Key Laboratory Breeding Base of Dao-Di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences;Chinese Pharmacopoeia Commission;

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DOI: 10.19540/j.cnki.cjcmm.20200608.101

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