ATP生物荧光增幅法在化妆品微生物检测中的验证研究

doi:10.3969/j.issn.1001-1803.2020.11.006

日用化学工业 ›› 2020, Vol. 50 ›› Issue (11): 771-775.doi: 10.3969/j.issn.1001-1803.2020.11.006

ATP生物荧光增幅法在化妆品微生物检测中的验证研究

翟磊¹(),葛媛媛¹,崔生辉²,刘吉泉³,洪海军⁴,Annemieke Masolijn⁵,姚粟¹()

1.中国食品发酵工业研究院有限公司,北京 100015
2.中国食品药品检定研究院,北京 100050
3.Procter & Gamble International Operations,新加坡 138547,新加坡
4.联合利华（中国）有限公司,上海 200335
5.Charles River, Microbial Solutions,马斯特里赫特 6199,荷兰

收稿日期:2020-07-24 修回日期:2020-10-20 出版日期:2020-11-22 发布日期:2020-11-20
通讯作者: 姚粟
作者简介:翟磊（1983-）,男,河北人,高级工程师,博士,电话:（010）53218288,E-mail:zhailei@china-cicc.org。

Validation study of amplified ATP bioluminescence assay for microbial detection in cosmetics

ZHAI Lei¹(),GE Yuan-yuan¹,CUI Sheng-hui²,LIU Ji-quan³,HONG Hai-jun⁴,Annemieke Masolijn⁵,YAO Su¹()

1. China National Research Institute of Food & Fermentation Industries Co., Ltd., Beijing 100015, China
2. National Institutes for Food and Drug Control, Beijing 100050, China
3. Procter & Gamble International Operations, Singapore 138547, Singapore
4. Unilever （China） Limited, Shanghai 200335, China
5. Charles River, Microbial Solutions, Maastricht 6199, Netherlands

Received:2020-07-24 Revised:2020-10-20 Online:2020-11-22 Published:2020-11-20
Contact: Su YAO

摘要/Abstract

摘要：

选取清洁类、护理类、美容修饰类6种化妆品和7株代表菌株,开展了ATP生物荧光增幅法与《化妆品安全技术规范》（2015年版）中菌落总数、霉菌和酵母菌检验方法的部分等效性研究。将代表菌株按照目标浓度10,5,1和0.1 CFU/g分别接种于6种化妆品,每个浓度重复检测5次,经Fisher exact test统计分析发现ATP生物荧光增幅法的检测限显著优于标准方法（p=0.000 1）。7株代表菌株在MTAT培养基中生长良好,ATP生物荧光增幅法检测人工污染样品结果均为阳性,表明该方法的专属性良好。选取3家不同的实验室,采用两种方法对相同产品进行比对检测,检测结果无统计学差异（p>0.05）,表明该方法的重现性良好。选取不同品牌TAT肉汤配制的富集培养基MTAT进行增菌,ATP检测结果均为阳性,表明该方法的耐用性良好。因此,ATP生物荧光增幅法与标准方法具有部分等效性。

关键词: ATP生物荧光增幅法, 化妆品, 微生物检测, 方法验证

Abstract:

Six kinds of cosmetics, including cleanser, care cosmetics and makeup categories, and seven representative microorganisms were selected to conduct the partial equivalence study between amplified ATP bioluminescence assay and the detection method of aerobic bacterial count and molds and yeast count in Safety and Technical Standard for Cosmetics （2015th Edition）. The seven representative microorganisms were inoculated into the six kinds of cosmetics at target concentrations of 10, 5, 1 and 0.1 CFU/g, respectively. Tests on each concentration were repeated five times. Fisher exact test statistical analysis show that the detection limit of amplified ATP bioluminescence assay is significantly better than that of the standard method （p=0.000 1）. Seven representative strains grew well in the MTAT medium, and the results of artificially contaminated samples detected by the amplified ATP bioluminescence assay were all positive, indicating that the method has good specificity. Three different laboratories were selected to compare the two methods using the same samples, and no statistical difference was found in the results, indicating the method is reproducible （p>0.05）. The enrichment media MTAT, prepared with selected brands of TAT base medium, were assessed using growth promotion, and their ATP test results were all positive, indicating that the method is has good durability. Therefore, the amplified ATP bioluminescence assay is partially equivalent to the standard method.

Key words: amplified ATP bioluminescence assay, cosmetics, microbial detection, method validation

中图分类号:

TQ658

翟磊,葛媛媛,崔生辉,刘吉泉,洪海军,Annemieke Masolijn,姚粟. ATP生物荧光增幅法在化妆品微生物检测中的验证研究[J]. 日用化学工业, 2020, 50(11): 771-775.

ZHAI Lei,GE Yuan-yuan,CUI Sheng-hui,LIU Ji-quan,HONG Hai-jun,Annemieke Masolijn,YAO Su. Validation study of amplified ATP bioluminescence assay for microbial detection in cosmetics[J]. China Surfactant Detergent & Cosmetics, 2020, 50(11): 771-775.

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参考文献 12

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菌株	检测方法	接种浓度/（CFU·g^-1）
		洗发水				沐浴液				护发素				定型水				面霜				面膜
		10	5	1	0.1	10	5	1	0.1	10	5	1	0.1	10	5	1	0.1	10	5	1	0.1	10	5	1	0.1
A. niger	CSTSC¹	0	0	0	0	0	0	0	0	5	5	3	0	0	0	0	0	5	5	2	1	5	5	3	1
A. niger	ATP²	5	5	4	2	5	5	5	1	5	5	5	1	5	4	1	1	5	5	5	3	5	5	5	2
C. albicans	CSTSC	0	0	0	0	3	0	0	0	5	4	2	0	5	4	2	0	5	5	2	0	5	4	3	0
C. albicans	ATP	5	5	5	0	5	5	4	1	5	5	5	0	5	5	5	1	5	5	4	1	5	5	4	1
B. subtilis	CSTSC	0	0	0	0	1	0	0	0	0	0	0	0	0	0	0	0	2	1	0	0	3	0	0	0
B. subtilis	ATP	5	5	5	0	5	5	5	2	5	5	3	1	5	5	5	0	5	5	2	0	5	5	5	0
P. aeruginosa	CSTSC	5	4	2	0	4	3	0	0	4	3	1	0	5	4	2	0	5	4	2	0	5	4	2	0
P. aeruginosa	ATP	5	5	5	1	5	5	5	0	5	5	4	2	5	5	5	2	5	5	4	2	5	5	5	1
S. aureus	CSTSC	0	0	0	0	0	0	0	0	1	0	0	0	3	1	0	0	5	5	3	0	5	4	2	0
S. aureus	ATP	5	5	5	0	5	5	5	3	5	5	5	0	5	5	5	2	5	5	5	1	5	5	5	3
E. coli	CSTSC	5	4	3	0	4	4	0	0	5	4	3	0	5	4	3	0	5	4	2	0	5	5	3	0
E. coli	ATP	5	5	5	3	5	5	5	2	5	5	5	3	5	5	5	1	5	5	5	1	5	5	5	0
B. cepacia	CSTSC	5	3	1	0	5	3	1	0	5	4	2	0	4	3	2	0	3	2	2	0	4	2	0	0
B. cepacia	ATP	5	5	5	0	5	5	5	0	5	5	3	0	5	5	5	1	5	5	4	0	5	5	5	1

菌株	检测方法	LOD₅₀/（CFU·g^-1）	菌株	检测方法	p值	LOD₅₀/（CFU·g^-1）
A. niger	CSTSC¹	8.33	B. subtilis	CSTSC	0.000 1	12.82
A. niger	ATP²	0.27	B. subtilis	ATP	0.000 1	0.62
C. albicans	CSTSC	5.35	P. aeruginosa	CSTSC	0.000 1	3.63
C. albicans	ATP	0.51	P. aeruginosa	ATP	0.000 1	0.35
S. aureus	CSTSC	9.75	B. cepacia	CSTSC	0.000 1	4.83
S. aureus	ATP	0.14	B. cepacia	ATP	0.000 1	0.59
E. coli	CSTSC	2.68
E. coli	ATP	0.13

菌株	检测方法	检测结果			p值
菌株	检测方法	实验室1	实验室2	实验室3	p值
S. aureus	CSTSC¹	4	4	4	1.000
S. aureus	ATP²	5	5	5	1.000
C. albicans	CSTSC	5	5	5	1.000
C. albicans	ATP	5	5	5	1.000

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ATP生物荧光增幅法在化妆品微生物检测中的验证研究

Validation study of amplified ATP bioluminescence assay for microbial detection in cosmetics

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