1、July 2014 Translation by DIN-Sprachendienst.English price group 10No part of this translation may be reproduced without prior permission ofDIN Deutsches Institut fr Normung e. V., Berlin. Beuth Verlag GmbH, 10772 Berlin, Germany,has the exclusive right of sale for German Standards (DIN-Normen).ICS 7
2、1.100.70!%7?“2202898www.din.deDDIN EN 16521Cosmetics Analytical methods GC/MS method for the identification and assay of 12 phthalates incosmetic samples ready for analytical injection;English version EN 16521:2014,English translation of DIN EN 16521:2014-07Kosmetische Mittel Analysenmethoden GC/MS-
3、Methode fr die Identifizierung und die Quantifizierung von 12 Phthalaten in zurdirekten Injektion geeigneten Proben kosmetischer Mittel;Englische Fassung EN 16521:2014,Englische bersetzung von DIN EN 16521:2014-07Cosmtiques Mthodes analytiques Mthode CG-SM pour lidentification et lanalyse de 12 phta
4、lates dans des chantillons deproduits cosmtiques prts tre injects dans un systme analytique;Version anglaise EN 16521:2014,Traduction anglaise de DIN EN 16521:2014-07www.beuth.deDocument comprises 15 pagesIn case of doubt, the German-language original shall be considered authoritative.06.14 DIN EN 1
5、6521:2014-07 2 A comma is used as the decimal marker. National foreword This document (EN 16521:2014) has been prepared by Technical Committee CEN/TC 392 “Cosmetics” (Secretariat: AFNOR, France). The responsible German body involved in its preparation was the Normenausschuss Lebensmittel und landwir
6、tschaftliche Produkte (Food and Agricultural Products Standards Committee), Working Committee NA 057-07-01 AA Kosmetische Mittel. EUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN 16521 May 2014 ICS 71.100.70 English Version Cosmetics - Analytical methods - GC/MS method for the identification and
7、 assay of 12 phthalates in cosmetic samples ready for analytical injection Cosmtiques - Mthodes analytiques - Mthode CG-SM pour lidentification et lanalyse de 12 phtalates dans des chantillons de produits cosmtiques prts tre injects dans un systme analytique Kosmetische Mittel - Analysenmethoden - G
8、C/MS-Methode fr die Identifizierung und die Quantifizierung von 12 Phthalaten in zur direkten Injektion geeigneten Proben kosmetischer Mittel This European Standard was approved by CEN on 10 April 2014. CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the con
9、ditions for giving this 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 exis
10、ts in three official versions (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 sta
11、ndards bodies of Austria, 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, S
12、lovenia, Spain, Sweden, Switzerland, Turkey and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG CEN-CENELEC Management Centre: Avenue Marnix 17, B-1000 Brussels 2014 CEN All rights of exploitation in any form and by any means reser
13、ved worldwide for CEN national Members. Ref. No. EN 16521:2014 EEN 16521:2014 (E) 2 Contents Page Foreword 3 Introduction .3 1 Scope 5 2 Reagents .5 3 Apparatus and equipment 6 4 Procedure .6 4.1 Standard purity 6 4.2 Sample preparation .7 4.3 Gas chromatography (GC) measurement conditions 7 4.4 Det
14、ection .8 4.4.1 General 8 4.4.2 MS detection in Selected Ion Monitoring (SIM) mode 8 5 Evaluation .9 5.1 Identification and quantitative determination .9 5.2 Calculation . 10 6 Test report . 10 Annex A (informative) Example of Chromatograms . 11 Bibliography . 12 DIN EN 16521:2014-07 EN 16521:2014 (
15、E) 3 Foreword This document (EN 16521:2014) 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 endorsement, at the latest
16、 by November 2014 and conflicting national standards shall be withdrawn at the latest by November 2014. 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 identifying any or all su
17、ch 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 Yugoslav Republic of Mace
18、donia, 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. DIN EN 16521:2014-07 EN 16521:2014 (E) 4 Introduction Phthalates are
19、esters of phthalic acid (Figure 1). About 80 % of all phthalates manufactured are used as “plasticizers” to make plastics flexible without sacrificing strength or durability. These compounds are present in cosmetic products like perfumes and toiletries. Some phthalates, particularly those of low mol
20、ecular weight, are introduced into cosmetics as ingredients, for examples DEP and DMP are used as solvents and perfume fixatives 1-3 or DEP can be used as alcohol denaturing 2, 4. Their presence in such products may come from their use as ingredients during the manufacturing process or may come from
21、 the migration of phthalates from packaging when plastic is used. Their presence as contaminant could also be due to the manufacturing process or raw materials used. Some analytical methods are proposed in the literature for the determination of phthalates in cosmetic products 1, 4-12. Figure 1 Este
22、rs of phthalic acid This standard proposes a GC/MS method for a simultaneous assay in cosmetic samples ready for analytical injection of 12 phthalates listed in Table 1. These chromatographic conditions are not suitable for the quantification of di-isononyl phthalate (DiNP) or di-isodecyl phthalate
23、(DiDP). According to SCCP 13, the possible presence of DiNP or DiDP in cosmetics does not seem to be a problem for human health. A GC/MS method using positive chemical ionisation with ammonia as collision gas is proposed in literature for the determinations of those compounds in cosmetic products 14
24、. Analyses are carried out on a GC/MS system with electron impact ionization mode (Ei). The separation of phthalates is obtained on a cross-linked 5 %-phenyl/95 %-dimethylpolysiloxane capillary column 30 m 0,25 mm (i. d.) 0,25 m film thickness using a temperature gradient. Phthalate quantification i
25、s performed by external calibration using an internal standard or by the standard addition. Cosmetic samples are analyzed directly or after a previous dilution in ethanol 15. DIN EN 16521:2014-07 EN 16521:2014 (E) 5 1 Scope This European Standard describes a GC/MS method for the assay of 12 phthalat
26、es, amongst which the 8 phthalates regulated by the European cosmetic regulation 1223/2009 16. This method is given for the analysis of samples ready for analytical injection from cosmetic products or raw materials used in cosmetic products. Samples should be compatible with GC analysis possibly aft
27、er dilution. This method does not include requirements for the preparation of samples in cosmetic matrices for which direct injection in GC is not feasible. 2 Reagents If not otherwise specified, analytical-grade chemicals shall be used. 2.1 Phthalates considered Table 1 Phthalates considered Phthal
28、ates CAS ManufacturerbQuality DBP a(dibutyl phthalate) 84742 ALDRICH 97,0 % DEHP a(diethylhexyl phthalate) 117817 ALDRICH 99,8 % BBP a(butylbenzyl phthalate) 85687 ALDRICH 97,0 % DMEP a(di(2-methoxyethyl) phthalate) 117828 ALDRICH 97,0 % DnPP a(di-n-pentyl phthalate) 131180 CIL CLUZEAU 99,0 % DiPP a
29、(diisopentyl phthalate) 605505 CIL CLUZEAU 95,0 % DPP a(n-pentyl isopentyl phthalate) 84777060 CIL CLUZEAU 95,0 %* DiBP a(diisobutyl phthalate) 84695 ACROS 98,0 % DCHP (dicyclohexyl phthalate) 84617 ALDRICH 98,0 % DEP (diethyl phthalate) 84662 ACROS 98,0 % DMP (dimethyl phthalate) 131113 ACROS 98,0
30、% DnOP (di-n-octyl phthalate) 117840 ALDRICH 98,0 % aRegulated phthalates. bThis is an example of a suitable product available commercially. This information is given for the convenience of users of this document and does not constitute an endorsement by CEN of this product. * Mixed isomers (DiPP, D
31、nPP and DPP) 2.2 Ethanol 2.3 Internal standard, 4,4-Dibromodiphenyl from Fluka 1)97,0 % was used as internal standard (ISTD). 2.4 Internal standard stock solution (SM-ISTD), c = 1 000 g/ml. 1) This is an example of a suitable product available commercially. This information is given for the convenie
32、nce of users of this document and does not constitute an endorsement by CEN of this product. DIN EN 16521:2014-07 EN 16521:2014 (E) 6 Weigh approximately 10 mg of 4,4-Dibromodiphenyl (2.3) into a 10 ml volumetric flask. Firstly, dissolve in a small amount of ethanol (2.2) and then fill to the calibr
33、ation mark with ethanol. This stock solution is daily prepared. 2.5 Phthalates stock solution, c = 500 g/ml (SM). Weigh approximately 10 mg of each phthalate (2.1) into a 20 ml volumetric flask. Firstly, dissolve in a small amount of ethanol (2.2) and then fill to the calibration mark with ethanol.
34、This stock solution has a shelf life of at least 2 weeks if stored in a refrigerator. NOTE Due to the low amount of DIPP standard (sold in quantities of 10,0 mg), a stock solution of this phthalate is prepared independently from other phthalates. In this way, two intermediate stock solutions SM-1 an
35、d SM-2 at 1 000 g/ml are prepared. SM-1 is prepared weighting approximately 10 mg of each phthalate (2.1), except DiPP, into a 10,0 ml volumetric flask, whereas SM-2 is prepared weighting approximately 5,0 mg of DiPP (2.1) into a 5,0 ml volumetric flask. The final stock solution SM is obtained mixin
36、g equal volumes of both standard solutions (c = 500 g/ml). All these stock solutions have a shelf life of at least 2 weeks if stored in a refrigerator. 2.6 Calibration solutions (standard solutions) 1,0 ml of the phthalate stock solution SM (2.5) is transferred into a 10,0 ml volumetric flask and fi
37、lled with ethanol (2.2) up to the calibration mark (c = 50 g/ml). From this intermediate solution (S1), at least 5 calibration solutions are prepared by dilution in ethanol (2.2) after the addition of 100 l of the internal standard stock solution (SM-ISTD). Phthalates concentrations on these calibra
38、tion solutions ranges from 0,25 g/ml to 5,0 g/ml with an ISTD concentration fixed at 10,0 g/ml. These calibration solutions are prepared extemporaneously and injected. If cosmetic samples (perfume) are directly prepared in a 1,5 ml GC vial, according to 4.2, the preparation of calibration solutions
39、have to be adapted: Calibration solutions, ranging from 0,25 g/ml to 5,0 g/ml, are prepared without internal standard and 10 l of the internal standard stock solution (SM-ISTD) are added to 1,0 ml of each calibration solution directly in a 1,5 ml GC vial. These calibration solutions are prepared ext
40、emporaneously. Vials are shaken and the solution injected. 3 Apparatus and equipment 3.1 Standard laboratory equipment. 3.2 Gas chromatography/mass spectrometry apparatus. 3.3 Gas chromatography/FID (for standard purity). 3.4 Analytical separation column. The following parameters have proved useful:
41、 GC column, low bleeding phase: 5 %-Phenyl-95 %-dimethylpolysiloxane, 30 m 0,25 mm (i. d.) 0,25 m or equivalent material. 4 Procedure 4.1 Standard purity The purity of each standard and the internal standard and the respective percentages for geometrical isomers (n-pentyl isopentyl phthalate - DPP)
42、shall be determined by GC-FID for further calculations. DIN EN 16521:2014-07 EN 16521:2014 (E) 7 4.2 Sample preparation 1,0 g of the cosmetic sample ready for analytical injection is transferred into a 10,0 ml volumetric flask and filled with ethanol (2.2) up to the calibration mark after the additi
43、on of 100 l of the internal standard stock solution (SM-ISTD). This solution is injected. In case of excessive concentration of phthalates, an appropriate previous dilution of the sample is performed. The limit of quantification using this sample preparation was set at 5 ppm. An alternative sample p
44、reparation consists to prepare the cosmetic sample directly in a 1,5 ml GC vial. 10 l of the internal standard stock solution (SM-ISTD) are added to 1,0 ml of cosmetic samples (perfume). Vials are shaken and the solution injected. The limit of quantification obtained using this sample preparation wa
45、s set at 0,5 ppm. Calibration solutions are prepared according to 2.6. 4.3 Gas chromatography (GC) measurement conditions When using the apparatus (3.2) and column (3.4), the following conditions have shown to be useful: Table 2 GC/MS programme Apparatus Gas chromatography with mass selective detect
46、or (GC/MS) Autosampler Column GC column, low bleeding phase: 5 %-Phenyl-95 %-dimethylpolysiloxane, 30 m 0,25 mm 0,25 m (or equivalent) Oven programme Ramp Temperature Time 100 C 0 min 30 C/min 200 C 0 min 3 C/min 260 C 0 min 30 C/min 320 C 5 min Injector T 300 C Interface T 250 C Source T 230 C Inje
47、ction time 30 min Gas / flow rate He/1 ml/min Injection port split/splitless Injection parameters : 1 l/Constant pressure, Split 1/20 Detection mode Quadrupole Ionisation mode EI (70 eV) Mass detection (Full/SIM) Identification: full-scan (m/z 40 to 350) Quantification: SIM using 3 specific ions Int
48、ernal standard 4,4-Dibromodiphenyl Calibration 0,25 g/ml to 5,0 g/ml Solvent used Ethanol (injection) DIN EN 16521:2014-07 EN 16521:2014 (E) 8 Examples of chromatogram, obtained using the GC/MS programme described in Table 2, are given in Annex A (Figure A.1). 4.4 Detection 4.4.1 General The detection and quantitative determination can be performed by evaluating the mass traces of each phthalate. Relative Retention (RR), m/z ions used for qua
