药学学报, 2021, 56(5): 1497-1508
引用本文:
辛天怡, 闫海霞, 李冉郡, 娄千, 郝利军, 廖保生, 刘颖, 陈晶, 陈有根, 杜小伟, 郭洪祝, 傅欣彤, 宋经元. DNA条形码技术在国家药品抽验中的应用研究[J]. 药学学报, 2021, 56(5): 1497-1508.
XIN Tian-yi, YAN Hai-xia, LI Ran-jun, LOU Qian, HAO Li-jun, LIAO Bao-sheng, LIU Ying, CHEN Jing, CHEN You-gen, DU Xiao-wei, GUO Hong-zhu, FU Xin-tong, SONG Jing-yuan. Application of DNA barcoding technology to national drug sampling inspection[J]. Acta Pharmaceutica Sinica, 2021, 56(5): 1497-1508.

DNA条形码技术在国家药品抽验中的应用研究
辛天怡1, 闫海霞2, 李冉郡1,3, 娄千1, 郝利军1, 廖保生4, 刘颖2, 陈晶2, 陈有根2, 杜小伟2, 郭洪祝2, 傅欣彤2*, 宋经元1,5*
1. 中国医学科学院、北京协和医学院药用植物研究所, 国家中医药管理局中药资源保护重点研究室, 北京 100193;
2. 北京市药品检验研究所, 国家药品监督管理局中成药质量评价重点实验室, 中药成分分析与生物评价北京市重点实验室, 北京 102206;
3. 西南交通大学生命科学与工程学院, 四川 成都 610031;
4. 中国中医科学院中药研究所, 北京 100700;
5. 中药资源教育部工程研究中心, 北京 100193
摘要:
国家药品抽验工作中发现中成药红金消结胶囊、红金消结片和柴黄颗粒处方中部分饮片来源复杂,存在掺伪现象,但常规检验方法难以确定混伪品基原物种。故本研究收集其处方中部分中药,包括柴胡、八角莲、黑蚂蚁和鼠妇虫饮片及相关药材样品184份,采用DNA条形码技术进行基原物种鉴定。结果显示,115份市售柴胡类药材及饮片获得111条ITS2序列,其中71份为柴胡(Bupleurum chinense)、3份为狭叶柴胡(B.scorzonerifolium)、31份为同属混伪品,另检出非同属伪品臭椿(Ailanthus altissima)1份、防风(Saposhnikovia divaricata)2份、一枝黄花(Solidago decurrens)3份;22份市售八角莲药材样本获得21条ITS2序列,其中15份为混伪品南方山荷叶(Diphylleia sinensis)、6份为八角莲(Dysosma versipellis)等鬼臼属植物;22份市售黑蚂蚁饮片基因组DNA降解均未能获得COI序列;38份市售鼠妇虫饮片样本获得24条COI序列,其中9份为平甲虫(寻常卷甲虫Armadillidium vulgare)、11份为光滑鼠妇(Porcellio laevis)、2份为中华蒙潮虫(Mongoloniscus sinensis),另有2份不能判定物种。本研究证实中药材DNA条形码分子鉴定技术适用于中成药处方投料的基原鉴定,有助于建立中药材、中药饮片、中成药生产流通使用全过程追溯体系。
关键词:    国家药品抽验      DNA条形码      ITS2      COI      市场监控     
Application of DNA barcoding technology to national drug sampling inspection
XIN Tian-yi1, YAN Hai-xia2, LI Ran-jun1,3, LOU Qian1, HAO Li-jun1, LIAO Bao-sheng4, LIU Ying2, CHEN Jing2, CHEN You-gen2, DU Xiao-wei2, GUO Hong-zhu2, FU Xin-tong2*, SONG Jing-yuan1,5*
1. Key Lab of Chinese Medicine Resources Conservation, State Administration of Traditional Chinese Medicine of the People's Republic of China, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100193, China;
2. Beijing Institute for Drug Control, NMPA Key Laboratory for Quality Evaluation of Traditional Chinese Medicine(Traditional Chinese Patent Medicine), Beijing Key Laboratory of Analysis and Evaluation on Chinese Medicine, Beijing 102206, China;
3. School of Life and Science, Southwest Jiaotong University, Chengdu 610031, China;
4. Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China;
5. Engineering Research Center of Chinese Medicine Resource, Ministry of Education, Beijing 100193, China
Abstract:
Adulterants and counterfeits were found in some of the commercial traditional Chinese medicine (TCM) decoctions in Hongjin Xiaojie Jiaonang, Hongjin Xiaojie Pian, and Chaihuang Keli during the national drug sampling inspection. However, it was difficult to determine the species of the adulterants and counterfeits by conventional testing methods. Therefore, a total of 184 samples of the TCM decoctions and raw materials belong to the prescriptions of above mentioned traditional Chinese patent medicines, including Bupleuri Radix, Bajiaolian, Heimayi, and Shufuchong, were collected and authenticated by DNA barcoding technology. 111 ITS2 sequences were obtained from 115 commercial TCM decoctions and raw materials of Bupleuri Radix, among which 71 were Bupleurum chinense, three were B. scorzonerifolium, and 31 were closely related species in the same genus. In addition, counterfeits derived from different genera, such as Ailanthus altissima (one sample), Saposhnikovia divaricate (two samples), and Solidago decurrens (three samples), were also detected. 21 ITS2 sequences were obtained from 22 commercial TCM raw materials of Bajiaolian, among which 15 were Diphylleia sinensis and six were Dysosma versipellis and other species in genus Dysosma. For 22 Heimayi samples, PCR amplification of COI sequence was failed due to genomic DNA degradation. Among 38 Shufuchong samples, 24 COI sequences were obtained and only nine of them were the genuine species (Armadillidium vulgare) recorded in the Chinese Pharmacopoeia, 11 were Porcellio laevis, two were Mongoloniscus sinensis, and two samples could not be identified due to the limitation of database. This study demonstrates that DNA barcoding technology is suitable for the species authentication of the decoctions of traditional Chinese patent medicine prescription. It is a conductive way for the establishment of traceability system for the whole TCM industrial chain.
Key words:    national drug sampling inspection    DNA barcoding    ITS2    COI    market supervision   
收稿日期: 2021-01-18
DOI: 10.16438/j.0513-4870.2021-0089
基金项目: 重大新药创制(2019ZX09201005-006-003);中国医学科学院医学与健康科技创新工程(2020-I2M-2-011).
通讯作者: 宋经元,Tel:86-10-57833199,E-mail:jysong@implad.ac.cn;傅欣彤,Tel:86-10-52779633,E-mail:fuxintong68@163.com
Email: jysong@implad.ac.cn;fuxintong68@163.com
相关功能
PDF(1204KB) Free
打印本文
0
作者相关文章
辛天怡  在本刊中的所有文章
闫海霞  在本刊中的所有文章
李冉郡  在本刊中的所有文章
娄千  在本刊中的所有文章
郝利军  在本刊中的所有文章
廖保生  在本刊中的所有文章
刘颖  在本刊中的所有文章
陈晶  在本刊中的所有文章
陈有根  在本刊中的所有文章
杜小伟  在本刊中的所有文章
郭洪祝  在本刊中的所有文章
傅欣彤  在本刊中的所有文章
宋经元  在本刊中的所有文章

参考文献:
[1] Lin R. Standard Directory of Chinese Medicinal Materials (中国药材标准名录)[M]. Beijing:Science Press, 2011:502.
[2] Wang C, Wu J. Taxonomic studies on the genus Polyrhachis Mayr of China (Hymenoptcra:Formicidae)[J]. Forest Res (林业科学研究), 1991, 4:596-601.
[3] State Pharmacopoeia Committee. Pharmacopoeia of the People's Republic of China (中华人民共和国药典)[S]. Part 4. 2020 Ed. Beijing:China Medical Science Press, 2020:558
[4] Lin R. Standard Directory of Chinese Medicinal Materials (中国药材标准名录)[M]. Beijing:Science Press, 2011:527.
[5] Kang S, Yue J, Jiang Z, et al. Varieties textual research and species identification of Chinese medicine Armadillidium[J]. J Chin Med Mater (中药材), 2019, 42:1761-1765.
[6] Chen G. Terrestrial Isopoda fauna of typical zones in China[J]. Acta Zool Sin (动物学报), 2000, 46:255-264.
[7] Chen S, Pang X, Song J, et al. A renaissance in herbal medicine identification:from morphology to DNA[J]. Biotechnol Adv, 2014, 32:1237-1244.
[8] Zhang N, Xin T, Jin Y, et al. Identification of Alisma plantago-aquatica seeds based on traditional Chinese medicine DNA barcoding system[J]. World Sci Technol Mod Tradit Chin Med (世界科学技术-中医药现代化), 2016, 18:18-23.
[9] Lv R, Feng Z, Ma T, et al. Use of ITS2 and psbA-trnH DNA barcode sequences of wild Phytolacca resources in Guanzhong area of Shaanxi Province[J]. Acta Pharm Sin (药学学报), 2020, 55:1951-1956.
[10] Li J, Zhang R, Yu S, et al. Identification of Chinese medicinal materials Stellariae Radix and its adulterants using DNA barcoding[J]. Acta Pharm Sin (药学学报), 2019, 54:937-943.
[11] Xin T, Lou Q, Hao L, et al. DNA barcoding identification of the commercial decoction pieces of traditional Chinese medicine[J]. Acta Pharm Sin (药学学报), 2021, 56:879-889.
[12] Xin T, Li X, Yao H, et al. Survey of commercial Rhodiola products revealed species diversity and potential safety issues[J]. Sci Rep, 2015, 5:8337.
[13] Xin T, Su C, Lin Y, et al. Precise species detection of traditional Chinese patent medicine by shotgun metagenomic sequencing[J]. Phytomedicine, 2018, 47:40-47.
[14] Xin T, Xu Z, Jia J, et al. Biomonitoring for traditional herbal medicinal products using DNA metabarcoding and single molecule, real-time sequencing[J]. Acta Pharm Sin B, 2018, 8:488-497.
[15] Xin T, Li X, Yao H, et al. A two-dimensional DNA barcode system for circulation regulation of traditional Chinese medicine[J]. Sci Sin Vit (中国科学:生命科学), 2015, 45:695-702.
[16] Lou Q, Xin T, Song J. Application of DNA barcoding technology in the whole industrial chain of traditional Chinese medicine[J]. Acta Pharm Sin (药学学报), 2020, 55:1784-1791.
[17] State Pharmacopoeia Committee. Pharmacopoeia of the People's Republic of China (中华人民共和国药典)[S]. Part 4. 2020 Ed. Beijing:China Medical Science Press, 2020:490-492.
[18] Wang Y, Han X, Zhao Y, et al. Identification of commercial Bupleuri Radix and its adulterants based on ITS2 barcode[J]. Chin Tradit Herb Drugs (中草药), 2017, 48:3590-3596.
[19] Yu J, Zhao S, Ren M, et al. Identification of Bupleurum chinense and B. longiradiatum based on ITS2 barcode[J]. China J Chin Mater Med (中国中药杂志), 2014, 39:2160-2163.
[20] Han P, Dong H, Cheng L, et al. Study on the medicinal properties of "Jiangbianyiwanshui"[J]. Chin Tradit Pat Med (中成药), 2012, 34:1151-1154.
[21] Zhang P, Li N, Li M, et al. Report of supervision and quality of Chinese materia medica and decoction pieces in 2019[J]. Mod Chin Med (中国现代中药), 2020, 22:663-671.
[22] Lin Q, Huang Q, Teng A, et al. On quality and regulatory countermeasures of Tetrapanacis Medulla based on national drug sampling inspection[J]. China Pharm Aff (中国药事), 2019, 33:1419-1423.
[23] Liu J, Guo L, Ma S, et al. Analysis of quality problems of Centipeda minima based on national drug evaluation surveillance[J]. China Pharm Aff (中国药事), 2020, 34:325-329.
[24] Gao Y, Li D, Guo L, et al. Quality analysis and study on Paridis Rhizoma pieces based on national drug evaluation sampling inspection[J]. China Pharm Aff (中国药事), 2019, 33:754-759.
相关文献:
1.辛天怡, 娄千, 郝利军, 李冉郡, 林余霖, 许文杰, 马婷玉, 林烨暄, 董一昕, 蒋澄宇, 宋经元.市售中药饮片DNA条形码鉴定研究[J]. 药学学报, 2021,56(3): 879-889
2.吕瑞华, 冯昭, 马添翼, 吕蕊花, 张明英, 彭亮, 高静, 张岗.陕西关中野生商陆资源的ITS2和psbA-trnH条形码序列研究[J]. 药学学报, 2020,55(8): 1951-1956
3.魏妙洁, 刘金欣, 赵晴, 徐晓兰, 任萍, 李明玥, 石林春, 孙伟.普乐安片DNA条形码分子鉴定研究[J]. 药学学报, 2020,55(6): 1327-1333
4.郑辉, 邓楷煜, 陈安琪, 付绍兵, 周德, 王威威, 倪典墨, 任瑶瑶, 周嘉裕, 廖海.基于DNA条形码的川贝母及其近缘种的分子鉴定与亲缘关系研究[J]. 药学学报, 2019,54(12): 2326-2334
5.刘旭朝, 周丽思, 刘金欣, 贾静, 宋经元, 石林春.基于COI序列的水牛角及其易混伪品DNA条形码鉴定研究[J]. 药学学报, 2017,52(3): 494-499
6.向丽, 汤欢, 成金乐, 陈一龙, 邓雯, 郑夏生, 赖智填, 陈士林.超微破壁饮片DNA条形码基原物种追溯[J]. 药学学报, 2015,50(12): 1660-1667