1、raising standards worldwideNO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAWBSI Standards PublicationBS EN 16343:2013Cosmetics Analysisof cosmetic products Determination of 3-iodo-2-propynyl butylcarbamate (IPBC)in cosmetic preparations, LC-MS methodsBS EN 16343:2013 BRITISH STA
2、NDARDNational forewordThis British Standard is the UK implementation of EN 16343:2013.The UK participation in its preparation was entrusted to TechnicalCommittee CW/217, Cosmetics.A list of organizations represented on this committee can beobtained on request to its secretary.This publication does n
3、ot purport to include all the necessaryprovisions of a contract. Users are responsible for its correctapplication. The British Standards Institution 2013. Published by BSI StandardsLimited 2013ISBN 978 0 580 76543 8ICS 71.100.70Compliance with a British Standard cannot confer immunity fromlegal obli
4、gations.This British Standard was published under the authority of theStandards Policy and Strategy Committee on 31 May 2013.Amendments issued since publicationDate Text affectedBS EN 16343:2013EUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN 16343 May 2013 ICS 71.100.70 English Version Cosmetic
5、s - Analysis of cosmetic products - Determination of 3-iodo-2-propynyl butylcarbamate (IPBC) in cosmetic preparations, LC-MS methods Cosmtiques - Analyse des produits cosmtiques - Dtermination quantitative du carbamate 3-iodo-2-propynylbutyle (IPBC), mthodes CL-SM Kosmetische Mittel - Untersuchung v
6、on kosmetischen Mitteln - Bestimmung von 3-Iod-2-propinylbutylcarbamat (IPBC) in kosmetischen Mitteln, LC-MS-Verfahren This European Standard was approved by CEN on 25 April 2013. CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this
7、 European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN member. This European Standard exists in three official ve
8、rsions (English, French, German). A version in any other language made by translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management Centre has the same status as the official versions. CEN members are the national standards bodies of Austri
9、a, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden,
10、 Switzerland, Turkey and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG Management Centre: Avenue Marnix 17, B-1000 Brussels 2013 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Memb
11、ers. Ref. No. EN 16343:2013: EBS EN 16343:2013EN 16343:2013 (E) 2 Contents Page Foreword . 3 1 Scope 4 2 Principle 4 3 Reagents . 4 4 Apparatus and equipment . 5 5 Procedure . 5 6 Evaluation . 7 7 Test report 7 Annex A (informative) Results of the inter-laboratory test . 8 Bibliography 9 BS EN 16343
12、:2013EN 16343:2013 (E) 3 Foreword This document (EN 16343:2013) has been prepared by Technical Committee CEN/TC 392 “Cosmetics”, the secretariat of which is held by AFNOR. This European Standard shall be given the status of a national standard, either by publication of an identical text or by endors
13、ement, at the latest by November 2013, and conflicting national standards shall be withdrawn at the latest by November 2013. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. CEN and/or CENELEC shall not be held responsible for iden
14、tifying any or all such patent rights. According to the CEN-CENELEC Internal Regulations, the national standards organizations of the following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugo
15、slav Republic of Macedonia, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom. BS EN 16343:2013EN 16343:2013 (E) 4 1 Scope This
16、 European Standard specifies a method for the quantitative determination of 3-iodo-2-propynyl butylcarbamate (IPBC) in the concentration range from 0,005 g/100 g to 0,1 g/100 g - Annex V No. 56 in Regulation (EC) No 1223/2009 on cosmetic products. 2 Principle IPBC is extracted from the cosmetic prep
17、aration using methanol. IPBC present in the sample extract is separated using reverse phase HPLC with mass specific detection (LC-MS or LC-MS/MS). Quantitative determination of IPBC is made using the external standard method of calibration or standard addition. 3 Reagents 3.1 General If not otherwis
18、e specified, analytical-grade chemicals shall be used; water shall be distilled or of a suitable purity. “Solution” shall be understood as an aqueous solution unless otherwise specified. 3.2 Iodopropynyl butylcarbamat, CAS number: 55406-53-6 (Supplier : Sigma-Aldrich1)(521949), Dr. Ehrenstorfer GmbH
19、1)(C 14335000). 3.3 Methanol, HPLC grade, CAS number: 67-56-1. 3.4 Formic acid, CAS number: 64-18-6. 3.5 Tetrahydrofuran (THF), CAS number: 109-99-9. 3.6 Propan-2-ol, CAS number: 67-63-0. 3.7 Eluents 3.7.1 Eluent A, 1 ml of formic acid (3.4) is mixed with 1 000 ml of water. 3.7.2 Eluent B, Methanol
20、(3.3). 3.8 IPBC stock solution, = 1 mg/ml. Weigh approximately 0,05 g of IPBC (3.2) into a 50-ml-volumetric flask. Firstly, dissolve in a small amount of methanol (3.3) and then fill up to the calibration mark with methanol. This solution has a shelf life of 8 weeks if it is stored in a refrigerator
21、. 3.9 Calibration solutions (standard solutions) 5,0 ml of the stock solution (3.8) is transferred into a 50-ml-volumetric flask and filled to the calibration mark with methanol (3.3), ( = 0,1 mg/ml or 100 g/ml). From this solution, at least 5 solutions are prepared by dilution to obtain IPBC concen
22、trations of = 0,05 g/ml to = 1,0 g/ml. These solutions have a shelf life of 8 weeks if they are stored in a cool place. Examples of dilutions are given in Table 1. 1) This is an example of a suitable product available commercially. This information is given for the convenience of users of this docum
23、ent and does not constitute an endorsement by CEN of this product. Equivalent products may be used if they can be shown to lead to the same results. BS EN 16343:2013EN 16343:2013 (E) 5 Table 1 Calibration solutions No. Preparation Dilution Concentration g/ml 3.9 5 ml 3.8 filled up to 50 ml 1:10 100
24、3.10 10 ml 3.9 filled up to 100 ml 1:100 10 3.11 10 ml 3.10 filled up to 100 ml 1:1000 1 3.12 4 ml 3.10 filled up to 50 ml 1:1250 0,8 3.13 5 ml 3.10 filled up to 100 ml 1:2000 0,5 3.14 10 ml 3.11 filled up to 50 ml 1:5000 0,2 3.15 5 ml 3.11 filled up to 50 ml 1:10000 0,1 4 Apparatus and equipment 4.
25、1 Standard laboratory equipment 4.2 Membrane filter, in the form of a disposable syringe filter, pore width: 0,2 m2). 4.3 High-performance liquid chromatograph, suitable for gradient elution with mass-specific detector. 4.4 Analytical separation column, the following parameters have proved useful: R
26、P 18 Phase, 5 m, 150 mm 2 mm, e.g. Zorbax1), Spherisorb1), Phenomenex-Luna1)or equivalent. If a precolumn is used, it shall have the same analytical properties as the separation column. 5 Procedure 5.1 Sample preparation Weigh approximately 200 mg of the sample to the nearest 0,1 mg into a 20-ml-vol
27、umetric flask (alternatively a 50-ml-volumetric flask). Add 1,5 ml of THF (3.5) and shake. Add 10 ml of methanol (3.3), and allow to dissolve or suspend for 5 min in the ultrasonic bath at room temperature. Allow to cool to room temperature and fill to the calibration mark with methanol (3.3). The s
28、ample solution is diluted with methanol (3.3) to a ratio of 1:10, filtered through a membrane filter (4.2) and then analysed using LC-MS or LC-MS/MS. For poorly soluble or suspendable matrices it is recommended to partially dissolve the sample by adding 2 ml of 2-propanol (3.6) instead of THF (3.5)
29、or to stir it with a magnetic stirrer for 30 min prior to the treatment in the ultrasonic bath. 5.2 Liquid chromatography (LC) conditions When using the apparatus (4.3) and column (4.4), the following conditions have shown to be useful (see Table 2): 2) The ring test was performed using 0,2 m filter
30、. BS EN 16343:2013EN 16343:2013 (E) 6 Table 2 Gradient programme Time min Fraction eluent A % Fraction eluent B % 0 85 15 8 10 90 12 10 90 13 85 15 25 85 15 Column: RP 18 Phase, 5 m, 150 mm 2 mm Injection volume: 1 l to 10 l Flow rate: 0,2 ml/min Column oven temperature: 25 C 5.3 Detection 5.3.1 Gen
31、eral The detection and quantitative determination can be performed by evaluating the mass traces of IPBC or by evaluating the fragment ions. To avoid false low results by adduct formation occurring in the MRM mode of the Electrospray (ESI) method, the APCI (Atmospheric Pressure Chemical Ionisation)
32、method should be used for ionisation. 5.3.2 MS detection in Selected Ion Monitoring (SIM) mode Mass traces: m/z 282 M+H+and m/z 304 M+Na+. Evaluation is based on the total ion current (from the sum of the two masses). 5.3.3 MS detection in Multiple Reaction Monitoring (MRM) mode Protonated molecule
33、ion: 282 M+H+Fragment ion 1: 57 Fragment ion 2: 165 Evaluation is based on the most sensitive fragment ion. Since the analyte may form adducts with sodium ions, false low results can be obtained if larger amounts of sodium ions are present in the sample. If ESI ionisation is used, standard addition
34、should therefore be performed using the MRM mode so as to ensure reliable quantification results. When performing the standard addition, the content of the added IPBC should not exceed the content expected in the sample. BS EN 16343:2013EN 16343:2013 (E) 7 6 Evaluation 6.1 Identification and quantit
35、ative determination The IPBC is identified by comparing the retention times of the sample with those of calibration solutions. The quantitative determination of the analyte is performed based on a calibration function or the standard addition. The calibration solutions are chromatographed in accorda
36、nce with the conditions given in 5.2. The IPBC concentration is calculated from the calibration by linear regression on the basis of the obtained peak area. 6.2 Calculation The preservative content w in g/100 g, with respect to the sample, is calculated using the following formula: 10001000100=mFVcw
37、 (1) where w is the IPBC content, in g/100 g; c is the IPBC concentration in the sample solution, in g/ml, determined from the calibration function; m is the initial weight of the sample, in g; V is the volume of the sample measurement solution, in ml; F is the dilution factor, if required. The resu
38、lt is given in g/100 g, rounded to three decimal places. 7 Test report The test report shall contain the following data: a) information necessary for the identification of the sample (type, origin and designation of the sample); b) a reference to this European Standard; c) name of the laboratory per
39、forming the test; d) the date and type of sampling procedure (if known); e) the date of receipt and date of analysis; f) the date of test; g) the test results and the units in which they have been expressed; h) justification of any deviations from this official method; i) operations not specified in
40、 the method or regarded as optional, which might have affected the results. BS EN 16343:2013EN 16343:2013 (E) 8 Annex A (informative) Results of the inter-laboratory test This method has been developed by the working group “Cosmetics” of the German Federal Office of Consumer Protection and Food Safe
41、ty (BVL) for the purpose of implementing Section 64 of the Food and Feed Code (LFGB). It has been tested in an inter-laboratory test with a total of 13 participants. The following statistical data presented in Table A.1 have been determined for shower gel and cream in an inter-laboratory test with 1
42、3 participating laboratories using external quantification. Table A.1 Statistical data of the inter-laboratory test Parameter LC-MS method IPBC content, in g/100 g Shower gel Cream Number of participating laboratories 13 13 Number of outliers 3 2 Number of laboratories after elimination of the outli
43、ers 10 11 Mean value x , g/100 g 0,013 0,019 Repeatability r, g/100 g 0,003 0,003 Repeatability standard deviation sr, g/100 g 0,001 0,001 Reproducibility R, g/100 g 0,006 0,005 Reproducibility standard deviation sR, g/100 g 0,002 0,002 Recovery, % 101,4 103,7 BS EN 16343:2013EN 16343:2013 (E) 9 Bib
44、liography 1 Frauen M, Steinhart H, Rapp C, Hintze U. (2001): Rapid quantification of iodopropynyl butylcarbamate as the preservative in cosmetic formulations using high-performance liquid chromatography-electrospray mass spectrometry. J Pharm Biomed Anal. 25 (5-6):965-70 This page deliberately left
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