采用DNA条形码技术鉴别面膜粉中常用的药妆同源植物源性成分

doi:10.3969/j.issn.1001-1803.2021.06.009

摘要/Abstract

摘要：

以市面上常用于面膜粉的22种药妆同源植物原料为研究对象,比较了常用的5组基因的物种鉴别效果,筛选出适用于面膜粉中药妆同源植物源性成分鉴别的DNA条形码。结果表明:ITS 2/3和matK基因联合能够准确的鉴定药妆同源植物源性成分。同时采用该方法,结合本实验室建立的混合样品克隆测序流程,对14份市售样品（包括11份单一样品,3份混合产品）进行了适用性分析。实验结果显示面膜粉中的药妆同源植物成分存在35.71%的掺假率。本研究建立的基于ITS 2/3和matK基因的DNA条形码技术能够准确的鉴定面膜粉中的药妆同源植物源性成分,为该类产品的质量和安全提供技术支撑。

关键词: 面膜粉, 药妆同源植物, DNA条形码, 质量, 安全

Abstract:

Took 22 common cosmeceutical plant ingredients in mask powder products as the research object, and screened out the most suitable genes for species identification from five genes. The results show that combined ITS 2/3 and matK can accurately identify cosmeceutical homologous plants. Furthermore, combined with the clone sequencing process for mixture established by our laboratory, 14 commercial mask powder products （including 11 single samples and 3 mixed products） are used to analyze the applicability. The results show that the adulteration rate is 35.71% in the cosmetics and edible homologous plant in mask powder products. The establishment of DNA barcoding based on ITS 2/3 and matK genes can accurately and reliably identify cosmeceutical homologous plants, which provides technical support for the quality and safety control of mask powder products.

Key words: mask powder, cosmeceutical homologous plants, DNA barcoding, quality, safety

中图分类号:

TQ658

王洪越,王迎春,江丽,杨艳歌,刘鸣畅,吴亚君. 采用DNA条形码技术鉴别面膜粉中常用的药妆同源植物源性成分[J]. 日用化学工业, 2021, 51(6): 528-534.

WANG Hong-yue,WANG Ying-chun,JIANG Li,YANG Yan-ge,LIU Ming-chang,WU Ya-jun. Identification of the common cosmeceutical homologous plants in mask powder products based on DNA barcoding[J]. China Surfactant Detergent & Cosmetics, 2021, 51(6): 528-534.

图/表 5

表 1

表 2

5种DNA条形码对药妆同源植物样品物种鉴别结果"

类型	类别	物种分类		拉丁名	rbcL		matK		psbA-trnH		ITS 2/3		ITS 5/4
类型	类别	物种分类		拉丁名	测序比对结果	相似度/%	测序比对结果	相似度/%	测序比对结果	相似度/%	测序比对结果	相似度/%	测序比对结果	相似度/%
药妆同源植物	绿豆	豆科	豇豆属	Vigna radiata	Vigna radiata	100	Vigna radiata	97.01	Vigna radiata	91.19	Vigna radiata	97.31	Vigna radiata	99
	黄芪	豆科	黄耆属	Astragalus membranaceus	Astragalus ernestii	98.12	Astragalus mongholicus	93.45	Astragalus ernestii	83.59	Astragalus membranaceus	99.13	Astragalus membranaceus	99.85
	甘草	豆科	甘草属	Glycyrrhiza uralensis	Thermopsis mongolica	98.15	Glycyrrhiza uralensis	94.99	Glycyrrhiza uralensis	98.04	Glycyrrhiza uralensis	92.41	Thermopsis mongolica	99.55
	苦参	豆科	槐属	Sophora flavescens	Sophora flavescens	97.01	Sophora flavescens	91.36	Sophora flavescens	90.56	Sophora flavescens	98.63	Sophora flavescens	99.10
	百合	百合科	百合属	Lilium brownii	Lilium lancifolium	99.60	Nelumbo nucifera	95.93	Lilium brownii	95.73	Lilium brownii	91.1	—	—
	莲子	睡莲科	莲属	Nelumbo nucifera	Nelumbo nucifera	99.52	Nelumbo nucifera	97.12	Naravelia zeylanica	92.71	Nelumbo nucifera	96.44	—	—
	人参	五加科	人参属	Panax ginseng	Panax ginseng	91.87	Panax ginseng	97.79	Panax ginseng	98.42	Panax ginseng	98.61	Panax ginseng	90.35
	三七	五加科	人参属	Panax notoginseng	Panax notoginseng	97.22	Panax notoginseng	99.52	—	—	Panax notoginseng	99.34	Panax notoginseng	99.71
	白芍	毛茛科	芍药属	Paeonia lactiflora	—	—	Angelica sinensis	94.25	Paeonia lactiflora	98.52	Paeonia lactiflora	94.06	Paeonia lactiflora	91.82
	黄连	毛茛科	黄连属	Coptis chinensis	—	—	Coptis quinquefolia	98.62	Coptis quinquefolia	94.04	Coptis chinensis	99.77	—	—
	白附子	天南星科	犁头草属	Typhonium giganteum	Typhonium giganteum	98.16	Typhonium giganteum	96.68	Typhonium giganteum	91.97	Senna tora	92.87	—	—
	益母草	唇形科	益母草属	Leonurus japonicus	Leonurus japonicus	99.05	Leonurus japonicus	99.26	Leonurus japonicus	86.08	Leonurus japonicus	91.04	Leonurus japonicus	99.70
	丹参	唇形科	鼠尾草属	Salvia miltiorrhiza	Salvia miltiorrhiza	99.13	Salvia miltiorrhiza	97.77	Salvia miltiorrhiza	98.60	Salvia miltiorrhiza	92.19	—	—
	大黄	蓼科	大黄属	Rheum officinale	Rheum palmatum L.	98.62	Rheum rhomboideum	92.26	Rheum officinale	91.46	Rheum officinale	98.78	Rheum officinale	99.55
	当归	伞形科	当归属	Angelica sinensis	Angelica sinensis	98.09	Angelica sinensis	97.07	Angelica sinensis	97.31	Angelica sinensis	92.12	Angelica sinensis	99.55
	川芎	伞形科	藁本属	Ligusticum sinense	Ligusticum sinense	91.19	Ligusticum sinense	95.83	—	—	Ligusticum sinense	97.73	Ligusticum sinense	98.86
	红花	菊科	红花属	Carthamus tinctorius	Carthamus tinctorius	98.50	Carthamus tinctorius	99.86	Carthamus tinctorius	94.75	—	—	Carthamus tinctorius	94.23
	白术	菊科	苍术属	Atractylodes macrocephala	Atractylodes chinensis	98.14	Atractylodes macrocephala	93.46	Atractylodes macrocephala	99.58	—	—	Atractylodes macrocephala	95.48
	白芨	兰科	白芨属	Bletilla striata	Bletilla striata	97.20	Bletilla striata	91.93	Bletilla striata	95.68	Bletilla striata	97.25	Bletilla striata	97.74
	白蔹	葡萄科	蛇葡萄属	Ampelopsis japonica	Ampelopsis japonica	98.15	Ampelopsis japonica	94.80	Ampelopsis japonica	97.96	Ampelopsis japonica	99.55	Ampelopsis japonica	99.27
	白薇	萝藦科	鹅绒藤属	Vincetoxicum atratum	Vincetoxicum atratum	97.24	Vincetoxicum atratum	98.51	Vincetoxicum atratum	96.77	Vincetoxicum atratum	90.67	Vincetoxicum atratum	98.51
	玫瑰	蔷薇科	蔷薇属	Rosa rugosa	Rosa rugosa	97.54	Rosa rugosa	98.57	Rosa rugosa	98.85	Rosa rugosa	98.96	Rosa rugosa	90.40
对照样品	罂粟	罂粟科	罂粟属	Papaver somniferum L.	Papaver somniferum	98.78	Papaver triniifolium	99.16	Papaver somniferum	98.81	Papaver somniferum	99.79	Papaver somniferum	99.23
对照样品	白芷	伞形科	当归属	Angelica dahurica	Angelica dahurica	98.62	Angelica anomala	95.62	—	—	Angelica dahurica	98.87	—	—

表 2

图 1

表 3

面膜粉中药妆同源植物成分检测结果"

编号	样品名称	产地	标签成分	ITS			matK			结果分析
编号	样品名称	产地	标签成分	鉴定结果	物种名称	匹配率/%	鉴定结果	物种名称	匹配率/%	结果分析
1	现磨纯益母草粉	广西	益母草	Leonurus japonicus	益母草	99.07	Leonurus japonicus	益母草	99.87	鉴定物种为益母草,标签标识成分为益母草,与标签相符
2	黄连粉	四川	黄连	Rheum officinale	药用大黄	98.78	—	—	—	鉴定物种为药用大黄,标签标识成分为黄连,与标识不符
3	超细百合粉	湖南	百合	Lilium pumilum	细叶百合	97.14	Nelumbo nucifera	莲子	95.86	ITS鉴定物种为细叶百合,matK鉴定物种为莲子,根据标准样品鉴定结果,ITS鉴定结果准确,因此判定物种成分为细叶百合,根据2020版药典,细叶百合为百合的一种,因此判定与标签相符
4	莲子粉	安徽	莲子	Nelumbo nucifera	莲子	97.31	Nelumbo nucifera	莲子	95.97	鉴定物种为莲子,标签标识成分为莲子,与标签相符
5	白术粉	四川	白术	—	—	—	Atractylodes macrocephala	白术	93.46	鉴定物种为白术,标签标识成分为白术,与标签相符
6	白芨超细粉	四川	白芨	Astragalus membranaceus	黄芪	99.13	Astragalus membranaceus	黄芪	99.14	鉴定物种为黄芪,标签标识成分为白芨,与标识不符
7	当归粉	湖北	当归	Angelica sinensis	当归	99.13	Angelica sinensis	当归	99.57	鉴定物种为当归,标签标识成分为当归,与标签相符
8	黄芪粉	湖北	黄芪	Astragalus membranaceus	黄芪	99.53	Astragalus mongholicus	蒙古黄芪	93.45	鉴定物种为黄芪或蒙古黄芪,均为黄芪的一种,与标签标识成分黄芪相符
9	白芍粉	湖北	白芍	Paeonia lactiflora	白芍	98.74	Angelica sinensis	当归	93.99	ITS鉴定为白芍,matK鉴定为当归,标识成分为白芍,根据标准样品鉴定结果,ITS鉴定结果准确,因此判定物种成分与标签相符
10	白附子粉	河南	白附子	Senna tora	决明	76.87	Typhonium giganteum	白附子	96.68	ITS鉴定物种为决明,matK鉴定物种为白附子,标签标识成分为白附子,根据标准样品鉴定结果,matK鉴定结果准确,因此判定物种成分与标签相符
11	三七粉	云南	三七	Panax notoginseng	三七	99.34	Panax notoginseng	三七	100	鉴定物种为三七,标签标识成分为三七,与标签相符
12	七子白面膜粉	广东	白芍、白蔹、红花、白芨、甘草、绿豆、白薇、人参	Vigna radiata, Leonurus japonicas, Paeonia lactiflora, Glycine max	绿豆、益母草、白芍、大豆	92.93~ 98.98	Vigna radiata, Leonurus japonicus, Paeonia lactiflora, Glycine max, Carthamus tinctorius	绿豆、益母草、大豆、红花	94.18~ 98.87	鉴定出标签标识成分绿豆、白芍、红花,鉴定出标签未标识成分益母草和大豆,未检出标签标识成分白蔹、白芨、甘草、白薇、人参
13	面膜粉	广东	当归、白术、白芨、白蔹、白芍、三七	Angelica sinensis, Paeonia lactiflora, Glycine max	当归、白芍、大豆	89.68~ 100	Angelica sinensis, Paeonia lactiflora, Glycine max	当归、大豆	93.67~ 99.78	鉴定出标签标识成分当归、白芍,鉴定出标签未标识成分大豆,未检出标签标识成分白术、白芨、白蔹、三七
14	草本祛痘面膜粉	广东	丹参、苦参、川芎、三七、绿豆	Vigna radiata, Angelica sinensis, Sophora flavescens	绿豆、当归、苦参	85.33~ 99.13	Vigna radiata, Angelica sinensis, Sophora flavescens	绿豆、当归、苦参	91.36~ 98.60	鉴定出标签标识成分绿豆、苦参,鉴定出标签未标识成分当归,未检出标签标识成分丹参、川芎、三七

表 3

图 2

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基因	引物名称	引物方向（5'-3'）	扩增条件	扩增长度/bp	参考文献
rbcL	rbcL-F	TTGGCAGCATTCCGAGTAAC	95 ℃ 10 min; 95 ℃ 30 s, 50 ℃ 30 s, 72 ℃ 30 s, 35个循环; 72 ℃ 5 min	250	/
rbcL	rbcL-R	AGTAAACATGTTAGTAACAG		250	/
psbA-trnH	psbA-trnH-fwd	GTTATGCATGAACGTAATGCTC	95 ℃ 4 min; 94 ℃ 30 s, 64℃ 30 s, 72 ℃ 30 s, 35个循环; 72 ℃ 5 min	450	[13]
psbA-trnH	psbA-trnH-rev	CGCGCATGGTGGATTCACAATCC		450	[13]
matK	matK-3F-KIM	CGTACAGTACTTTTGTGTTTACGAG	94 ℃ 1 min; 94 ℃ 30 s, 52 ℃ 20 s, 72 ℃ 50 s, 35个循环; 72 ℃ 5 min	890	/
matK	matK-1R-KIM	ACCCAGTCCATCTGGAAATCTTGGTTC		890	/
ITS	2F	ATGCGATACTTGGTGTGAAT	94 ℃ 5 min; 94 ℃ 30 s, 56 ℃ 30 s, 72 ℃ 45 s, 40个循环; 72 ℃ 10 min	500	[14]
ITS	3R	GACGCTTCTCCAGACTACAAT		500	[14]
ITS	5F	GGAAGTAAAAGTCGTAACAAGG	94 ℃ 5 min; 94 ℃ 1min, 50 ℃ 1min, 72 ℃ 1 min33 s, 40个循环; 72 ℃ 7 min	750	[14]
ITS	4R	TCCTCCGCTTATTGATATGC		750	[14]