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Research and Application of Screening and Detection Technology for Cosmetics Risk Substances
Research and Application of Screening and Detection Technolog y for Cosmetics Risk Substance
Wang Haiyan
China National Institute for Food and Drug Control / State Drug Administration Key Laboratory of Cosmetics Research and Evaluation
WANG Hai-yan
National Institutes for Food and Drug Control/ NMPA Key Laboratory for Researching and Evaluation of Cosmetics
Wang Xinran
China Institute for Food and Drug Control/State Drug Administration Cosmetics Research and Evaluation Key Laboratory
WANG Xin-ran
National Institutes for Food and Drug Control/ NMPA Key Laboratory for Researching and Evaluation of Cosmetics< /span>
Lu Yong*
China National Institute for Food and Drug Control/State Drug Administration Cosmetics Research and Evaluation Key Laboratory
LU Yong*
National Institutes for Food and Drug Control/ NMPA Key Laboratory for Researching and Evaluation of Cosmetics
Summary
Main
< p>Abstract
Cosmetic safety risk substances mainly include prohibited substances in cosmetics, analogs and derivative modifications of prohibited substances, as well as exogenous risk compounds such as environmental pollutants . This paper introduces the research and application status of cosmetics risk substance screening and detection technology, aiming to promote the improvement and improvement of cosmetics safety risk substance detection technology, timely discover potential safety problems and hidden dangers of cosmetics, and protect the legitimate rights and interests of consumers, thereby promoting cosmetics in my country. The safety level has been improved as a whole.
Risk substances in cosmetics mainly include prohibited substances and their analogs, derivative modifications, and exogenous risk compounds such as environmental pollutants. This paper introduces the research on and application of screening and detection technology for risk substances in cosmetics. By promoting improvement of screening and detection technology for cosmetics risk substances, we aim to uncover potential safety issues in time, so as to protect consumers and improve the overall safety level of cosmetics.
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Key
Word
Key words
Cosmetics; Risk Substances; Targeted Risk Substance Screening; Non-targeted Risk Substance Structural Identification
< p>cosmetics; risk substances; targeted risk substance screening; structural identification of non-targeted risk substances
With the growing demand for cosmetics by consumers, the safety of cosmetics has been increasingly valued by the public . Illegal addition of drugs and prohibited components is an important factor in the safety of cosmetics. Prohibited components refer to substances that cannot be used as cosmetic raw materials. A total of 1,393 prohibited components are listed in the Safety Technical Specification for Cosmetics (2015 Edition), including 1,284 prohibited raw materials for cosmetics and 109 prohibited plant (animal) raw materials for cosmetics. Traditional detection methods have been unable to meet the detection requirements of safety risk substances. In order to improve the efficiency of cosmetic safety supervision and expand the scope of inspection of risk substances, high-throughput rapid screening and detection technology came into being.[1-6], It breaks through the limitations of existing detection methods and increases the screening dimension of illegally adding banned components. With the continuous concealment of the hidden safety hazards of cosmetics, the phenomenon of illegally adding dangerous substances other than prohibited components, such as adding derivatives of prohibited components, and even adding modified and synthesized risk compounds to avoid supervision, therefore, cosmetics The screening and identification technology of unknown risk substances also needs to be established and perfected to deal with the ever-changing security risks in a timely manner.
1 Cosmetics High-Throughput Targeted Risk Substance Screening and Detection Technology
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In view of the prohibited and restricted substances in cosmetics, illegal additions driven by economic interests, pollution, derivative effect substances, etc., the China Institute for Food and Drug Control (hereinafter referred to as: China Institute for Inspection and Quarantine) has established a cosmetic safety risk High-throughput screening and detection technology for substances, actively carry out research and application of integrated methods for high-throughput screening of banned and restricted cosmetics and other risk substances, and integrate Chinese and international standards and regulations, as well as newly discovered pollutants and derivative effect substances etc., optimize and integrate the pretreatment technology of various substrates such as cosmetic creams, liquids, gels, powders, blocks, muds, wax bases, sprays, aerosols, etc., and establish hormone substances, anti-infective drugs, Antihistamines, antikeratinizing drugs, antipyretic and analgesic drugs, epinephrine drugs, narcotic drugs, psychotropic drugs, preservatives, colorants, fluorescent whitening agents, packaging migratory substances, N-nitrosamine compounds, organic solvent residues, pesticide residues, prostaglandins and their analogs, allergenic fragrances, perfluorinated compounds and other comprehensive cosmetic risk substances screening library. After the sample is injected, the parent ion mass-to-charge ratio, product ion mass-to-charge ratio, retention time, and isotopic information of each compound are obtained. High-throughput screening of safety risk substances is performed by comparing with the information in the established risk compound database. Below is information on some of the risk compounds that the laboratory has established.
1.1 Experimental Section
1.1.1 Instruments< /p>
ACQUITY UPLC I-Class Ultra Performance Liquid Chromatograph (Waters Corporation); Xevo G2 XSQ-TOF Quadrupole Time-of-Flight Mass Spectrometer (Waters Corporation).
1.1.2 Reagents
Acetonitrile, ammonium formate, and formic acid, all of which were mass spectrometric pure .
1.1.3 Standard solution preparation
Accurately weigh 10 mg of each compound standard ( Accurate to 0.0001g) into a 10ml brown volumetric flask, make up to the mark with acetonitrile, and prepare a stock solution with a concentration of 1.0mg/ml.
Accurately pipette 10μl of the stock solution of each compound into a 10ml volumetric flask, make up to the mark with acetonitrile, and prepare a mixed standard intermediate solution with a concentration of 1μg/ml.
Accurately pipette 1ml of the mixed standard intermediate solution into a 10ml volumetric flask, make up to the mark with water, and prepare a mixed standard solution with a concentration of 100ng/ml.
1.1.4 Sample preparation
1.1.4.1 Creams, lotions, liquid water Base, gel, and mask samples
Weigh 0.2g of the sample (accurate to 0.001g), put it in a 10ml colorimetric tube with a stopper, add a small amount of acetonitrile, Vortex and mix on a vortex mixer, then add acetonitrile to near the mark, ultrasonically extract for 20 minutes, let stand to room temperature, make up to the mark with acetonitrile, shake well, centrifuge at 4000 r/min for 10 minutes, filter, and use an equal volume of acetonitrile solution Diluted, the diluted solution was filtered through a 0.22 μm filter membrane, and the filtrate was used as the solution to be tested.
1.1.4.2 Liquid oil-based samples
Weigh 0.2g of the sample (accurate to 0.001g) and place it in the In the centrifuge tube, add 2ml of n-hexane, disperse on a vortex mixer, then add 3ml of 70% acetonitrile, vortex to mix, centrifuge at 4000r/min for 10min, suck the lower layer solution into a 10ml colorimetric tube, and use the upper n-hexane layer with 3ml of 70% acetonitrile, mix by vortex, centrifuge at 4000r/min for 10min, draw the lower layer solution into a 10ml colorimetric tube, combine the two solutions, add 50% acetonitrile to the volume, filter, dilute with an equal volume of acetonitrile solution, The diluted solution was filtered through a 0.22 μm membrane, and the filtrate was used as the solution to be tested.
1.1.4.3 Block samples
Weigh 0.2g of the sample (accurate to 0.001g) and place it in 10ml In a colorimetric tube with a stopper, add acetonitrile to near the mark, vortex and mix, ultrasonically extract for 20 minutes, make up to the mark with acetonitrile, shake well, centrifuge at 4000 r/min for 10 minutes, filter, and dilute with an equal volume of acetonitrile solution. Filter through a μm filter, and the filtrate is used as the solution to be tested.
1.1.4.4 Wax-based samples
Weigh 0.2g of the sample (accurate to 0.001g) and place it in 10ml In a colorimetric tube with stopper, add 1 ml of tetrahydrofuran, vortex to disperse the sample, add acetonitrile mixed solution (containing 0.2% formic acid) to near the mark, vortex to mix, ultrasonically extract for 20 min, let stand to room temperature, use acetonitrile mixed solution (containing 0.2% formic acid) 0.2% formic acid) to the mark, centrifuge at 4000r/min for 10min, take 0.5ml of supernatant, add 0.5ml of water, vortex to mix, centrifuge at 4000r/min for 10min, take the supernatant, filter through 0.22μm membrane, The filtrate was used as the solution to be tested.
1.1.4.5 Powder and mud samples
Weigh 0.2g of the sample (accurate to 0.001g), put it in a 10ml colorimetric tube with a stopper, add acetonitrile mixed solution (containing 0.2% formic acid) to near the mark, vortex to mix evenly, ultrasonically Extraction for 20min, let stand to room temperature, use acetonitrile mixed solution (containing 0.2% formic acid) to make up to the mark, centrifuge at 4000r/min for 10min, take 0.5ml of supernatant, add 0.5ml of water, vortex to mix, centrifuge at 4000r/min for 10min , take the supernatant, filter through a 0.22 μm filter membrane, and use the filtrate as the solution to be tested.
1.1.5 Experimental Conditions
1.1.5.1 UPLC Separation Conditions
Injection volume: 3.0μl; Column: ACQUITYUPLC®BEH C18 (2.1mm×100mm, 1.7μm); Column temperature: 40℃; Flow rate: 0.35 ml/min; positive ion mode mobile phase: A=2mmol/L ammonium formate aqueous solution (containing 0.05% formic acid), B=acetonitrile; negative ion mode mobile phase: A=2mmol/L ammonium formate aqueous solution, B=acetonitrile. The UPLC gradient elution procedure is shown in Table 1.
1.1.5.2 Q-TOF-MS assay conditions
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Electrospray ion source conditions: ion polarity: ESI+/-; capillary voltage: 1kV; ion source temperature: 120°C; sampling cone voltage: 20V; desolvation temperature: 550°C; cone gas flow: 50L /h; Desolvation gas flow: 1000L/h; Low collision energy: 4eV; High collision energy: 15~50eV; Mass scanning range: 50~1200m/z; Real-time calibration solution: Leucine enkephalin solution (200pg/μl ).
1.2 Results and discussion
The process of screening and analysis of cosmetic risk substances is shown in Figure 1. Each compound entry in the established risk substance screening database contains basic information such as Chinese name, English name, chemical structure, molecular formula, average molecular weight, monoisotopic precise molecular weight, and CAS number. In addition, each compound has key qualitative information: retention time and high-resolution secondary fragments. The retention time in the screening library corresponds to the recommended chromatographic conditions, and the secondary fragment information corresponds to the recommended collision energy. The data-independent acquisition mode consists of alternate scans of “low collision energy” and “high collision energy”, which can effectively match high and low energy spectra and reduce false positives. One-shot injection simultaneously obtains primary and secondary high-resolution mass spectrometry information, which preserves the integrity of the data. When additional screening targets are required later, there is no need to reprocess the sample and injection, and re-analyze the existing full-scan data. analyze.
1.2.1 Establishment of database of anti-infective drugs and hormones strong>
Anti-infective drugs and hormonal compounds have always been the work of screening for illegal additives in laboratory cosmetics by virtue of their rapid anti-inflammatory and antipruritic properties and the illusion of making the skin smooth and delicate Emphasis [7]. Existing standard test methods contain a limited number of compounds and cannot cover the types of risk substances. The method established in the laboratory is gradually expanded by the database of anti-infective drugs and hormone compounds by chromatography-high resolution mass spectrometry, and the database is verified by matrix-spiked samples. After sample injection, the parent ion mass-to-charge ratio, product ion mass-to-charge ratio, retention time, and isotopic information of each compound are obtained, and the parent ion mass-to-charge ratio, product ion mass-to-charge ratio, retention time, and isotopic information of each compound in the sample are detected. Compare with the information in the database, and conduct qualitative screening of the safety risk substances contained in the platform through calculation and scoring; construct a standard curve for the samples confirmed as positive by screening, and calculate the content of safety risk substances. The data information of some anti-infective drugs and hormone compounds is shown in Table 2.
In hormone compounds, there are many groups of compounds that are isomers of each other, especially It is the cis-trans isomer and epimer which are difficult to separate. The isomers can be well separated under the experimental conditions. Figure 2 shows the separation effect of some hormone isomers.
1.2.2 Establishment of database of allergenic flavors and fragrances
Fragrants and fragrances can impart pleasant odors to cosmetics, mask the odor of product bases, and make cosmetics emit unique fragrances. However, with the continuous application and development of flavors and fragrances, the safety issues presented are increasingly concerned. Among the cosmetic safety incidents caused by flavors and fragrances, the most representative one is cosmetic allergy. Allergy cases caused by allergenic flavors and fragrances account for more than 1/3 of cosmetic allergy cases. The European Union stipulates the use limits of 26 common allergenic flavors and fragrances. Most of the current common detection methods only contain 26 compounds specified by the European Union, which is far from enough to cover the risk-sensitizing flavors and fragrances of cosmetics. The laboratory expanded the database of allergenic flavor and fragrance compounds, and the data information of some allergenic flavor and fragrance compounds is shown in Table 3.
4-methoxycoumarin, 6-hydroxy-4-methylcoumarin and 4-hydroxy- 7-Methylcoumarins are isomers of each other and can be well separated under the experimental conditions, as shown in Figure 4.
Among them, hydromorphone and norcodeine are isomers of each other , a good separation can be obtained under the experimental conditions, as shown in Figure 5.
1.2.4 Creation of perfluorinated compound database >
Perfluorinated compounds are a class of substances with a hydrophobic structure[8], which have unique physicochemical properties and are widely used in cosmetics. Studies have shown that long-term exposure to perfluorinated compounds increases the risk of disease. In 2019, 11 ministries and commissions including the Ministry of Ecology and Environment of my country jointly issued the “Announcement on Prohibiting the Production, Circulation, Use and Import and Export of Persistent Organic Pollutants such as Lindane”, which stipulated that “from March 26, 2019, the ban on perfluorooctane Production, distribution, use and import and export of sulfonic acid and its salts and perfluorooctane sulfonyl fluoride other than for acceptable purposes”. The laboratory immediately carried out the research and establishment of the detection method of PFOS, and on this basis, the database of perfluorinated compounds was expanded. See Table 5 for the data information of perfluorinated compounds.
1.2.5 Prostaglandin database establishment p>
Bimaprost, as a special drug for the treatment of glaucoma[9], has the side effect of promoting the growth of eyelashes. After making profits, the laboratory developed the detection methods for prostaglandin compounds and their analogs, and continuously expanded to the cosmetic risk substance screening database. Table 6 is the data information of expanded prostaglandin compounds.
In addition, prostaglandin compounds that are banned in cosmetics under the new EU regulations are also For the construction of screening data, see Table 7.
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1.2.6 Application examples of screening risk compounds in routine monitoring work in laboratories
Screening in routine monitoring work in laboratories See Figure 6 for an example of the application of the extracted compounds. Through the accurate mass number of the primary mass spectrometer, the comparison of fragment ions in the secondary spectrum, the retention time, the isotope abundance ratio and other information, the information in the database is compared with the information in the database to screen the samples contained in the sample. Risk compounds.
Figure 6 is an application example. The suspected risk substances screened by primary mass spectrometry are analyzed by primary isotope distribution and accurate mass matching of precursor ions. The abundance ratio and exact mass of the secondary fragments were matched, and the main fragments were all completely matched with the structure, and the accurate mass deviation of the fragments was less than 0.001Da, and the sample contained clotrimazole based on the matching score value.
2 Cosmetic Non-targeted Risk Substance Screening and Identification Technology
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As the illegal addition of cosmetics develops towards derivative modification and synthesis, the screening and identification technology of non-targeted risk substances in cosmetics is urgently needed introduced. In the successful completion of the detection and identification task of illegally adding unknown substances, the Central Inspection Institute has initially established a screening and identification process for unknown risk substances in cosmetics. First, by looking up relevant literature and public opinion reports, analyze the types of suspected risk substances that may be added. Next, suspicious compound scanning analysis is performed, and the suspicious compounds are screened by restricting conditions. Molecular element composition, degree of unsaturation, and exact molecular mass error must be satisfied when the molecular formula is assigned. Reduce the number of potential molecular ion peaks by limiting conditions. The possible molecular structure of the compound was deduced by analyzing the secondary mass spectrum. Relevant parent compound information was obtained by secondary structure searches and molecular formula searches in online web databases. The efficacy and public opinion information of the parent compound were searched to further verify the possibility of the deduced structure. Using different chromatographic separation systems, the samples were preliminarily purified and enriched, and prepared by medium and high pressure preparative chromatography to obtain monomer compounds. The monomer compound was determined by high-resolution mass spectrometry, and its exact molecular formula was obtained. The monomer compounds were determined by one-dimensional and two-dimensional nuclear magnetic resonance spectroscopy, and the chemical structures of the compounds were obtained [10-13]. Figure 7 shows the flow of the test method established by the laboratory in the successful case of illegal addition of non-targeted compounds.
3 Conclusion
Potential risk substances in cosmetics include hormone compounds, anti-infective drugs and other illegally added risk substances and exogenous risk substances. At present, in order to evade supervision, structural analogs and derivatives of risk substances have been added. In order to comprehensively screen risk substances and ensure the safety of consumers’ use of makeup, the CCIC has established a cosmetic risk substance screening platform. , allergenic flavors and fragrances, prostaglandin compounds and other illegal additives have been effectively screened and analyzed, which has played a role in the extensive screening of targeted risk substances. For non-targeted risk substances, the Central Inspection Institute has also initially established a corresponding screening and identification process, and it has been well applied in the incident of illegal addition of Benvimod in children’s cosmetics. While improving the targeted risk substance screening database, the CCIC has continuously improved the non-targeted risk substance screening technology, providing technical support for ensuring the safety of cosmetics.
First Author Profile
China Institute of Food and Drug Control, Dr. Haiyan Wang researcher. Major: Food and Cosmetics Inspection and Research
Introduction to the corresponding author
Lu Yong, Ph.D., Vice President of China Academy of Food and Drug Control/Director of Key Laboratory of Cosmetics Research and Evaluation of State Drug Administration. Supervision
Please scan the QR code for the long abstract in Chinese and English
Chinese Food Drug Regulation
0
International Standard Serial Number:
ISSN 1673-5390
National Unified Serial Number:
CN 11-5362/D
Journal level: National
Journal: Monthly
《Chinese Food and Drugs Supervision, as the official journal of the State Drug Administration, was founded in 199 For 8 years, it is currently a scientific and professional core journal under the supervision of the State Drug Administration and affiliated to China Health Media Group; it also studies and publicizes China’s food and drug regulatory policies, establishes scientific supervision concepts, improves supervision levels, and serves my country’s food and drug products. An important platform for the healthy development of the pharmaceutical industry.
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