DIN EN ISO 12966-3-2016 Animal and vegetable fats and oils - Gas chromatography of fatty acid methyl esters - Part 3 Preparation of methyl esters using trimethylsulfonium hydroxide.pdf

1、November 2016 English price group 8No 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 67.200.10!%YT“2585449www.din.deD

2、IN EN ISO 12966-3Animal and vegetable fats and oils Gas chromatography of fatty acid methyl esters Part 3: Preparation of methyl esters using trimethylsulfonium hydroxide (TMSH) (ISO 129663:2016);English version EN ISO 129663:2016,English translation of DIN EN ISO 12966-3:2016-11Tierische und pflanz

3、liche Fette und le Gaschromatographie von Fettsuremethylestern Teil 3: Herstellung von Methylestern mittels Trimethylsulfoniumhydroxid (TMSH) (ISO 129663:2016);Englische Fassung EN ISO 129663:2016,Englische bersetzung von DIN EN ISO 12966-3:2016-11Corps gras dorigines animale et vgtale Chromatograph

4、ie en phase gazeuse des esters mthyliques dacides gras Partie 3: Prparation des esters mthyliques laide dhydroxyde de trimthylsulfonium (TMSH) (ISO 129663:2016);Version anglaise EN ISO 129663:2016,Traduction anglaise de DIN EN ISO 12966-3:2016-11SupersedesDIN EN ISO 129663:201004www.beuth.deDocument

5、 comprises 12 pagesDTranslation by DIN-Sprachendienst.In case of doubt, the German-language original shall be considered authoritative.10.16 DIN EN ISO 12966-3:2016-11 2 A comma is used as the decimal marker. National foreword This document (EN ISO 12966-3:2016) has been prepared by Technical Commit

6、tee ISO/TC 34 “Food products” (Secretariat: AFNOR, France) in collaboration with Technical Committee CEN/TC 307 “Oil seeds, vegetable and animal fats and oils and their by-products Methods of sampling and analysis” (Secretariat: AFNOR, France). The responsible German bodies involved in its preparati

7、on were DIN-Normenausschuss Lebensmittel und landwirtschaftliche Produkte (DIN Standards Committee Food and Agricultural Products) and the Deutsche Gesellschaft fr Fettwissenschaft (German Fat Research Society), Joint Committee NA 057-05-05 AA Gemeinschaftsausschuss fr die Analytik von Fetten, len,

8、Fettprodukten, verwandten Stoffen und Rohstoffen. ISO 12966 consists of the following parts, under the general title Animal and vegetable fats and oils Gas chromatography of fatty acid methyl esters: Part 1: Guidelines on modern gas chromatography of fatty acid methyl esters Part 2: Preparation of m

9、ethyl esters of fatty acids Part 3: Preparation of methyl esters using trimethylsulfonium hydroxide (TMSH) Part 4: Determination by capillary gas chromatography The DIN Standards corresponding to the International Standards referred to in this document are as follows: ISO 661 DIN EN ISO 661 ISO 835

10、DIN EN ISO 835 ISO 1042 DIN EN ISO 1042 ISO 5555 DIN EN ISO 5555 Amendments This standard differs from DIN EN ISO 12966-3:2010-04 as follows: a) fats coming from milk and milk products are now excluded from the scope of the standard; b) the standard has been editorially revised and brought in line w

11、ith the current rules of presentation. Previous editions DIN EN ISO 12966-3: 2010-04 DIN EN ISO 12966-3:2016-11 3 National Annex NA (informative) Bibliography DIN EN ISO 661, Animal and vegetable fats and oils Preparation of test sample DIN EN ISO 835, Laboratory glassware Graduated pipettes DIN EN

12、ISO 1042, Laboratory glassware One-mark volumetric flasks DIN EN ISO 5555, Animal and vegetable fats and oils Sampling DIN EN ISO 12966-3:2016-11 4 This page is intentionally blank EUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN ISO 12966-3 June 2016 ICS 67.200.10 Supersedes EN ISO 12966-3:2009

13、English Version Animal and vegetable fats and oils - Gas chromatography of fatty acid methyl esters - Part 3: Preparation of methyl esters using trimethylsulfonium hydroxide (TMSH)(ISO12966-3:2016) Corps gras dorigines animale et vgtale - Chromatographie en phase gazeuse des esters mthyliques dacide

14、s gras - Partie 3: Prparation des esters mthyliques laide dhydroxyde de trimthylsulfonium (TMSH) (ISO 12966-3:2016) Tierische und pflanzliche Fette und le - Gaschromatographie von Fettsuremethylestern -Teil 3: Herstellung von Methylestern mittels Trimethylsulfoniumhydroxid (TMSH)(ISO 12966-3:2016) T

15、his European Standard was approved by CEN on 12 May 2016. CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical referenc

16、es 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 versions (English, French, German). A version in any other language made by translation under the responsibility of a CEN me

17、mber 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 Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, F

18、rance, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES

19、 KOMITEE FR NORMUNG CEN-CENELEC Management Centre: Avenue Marnix 17, B-1000 Brussels 2016 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN ISO 12966-3:2016 EEuropean foreword 31 Scope . 52 Normative references 53 Principle 54 Reagen

20、ts 55 Apparatus . 66 Sampling 67 Preparation of the test sample . 68 Procedure. 69 Test report . 7Bibliography 8Contents PageDIN EN ISO 12966-3:2016-11 EN ISO 12966-3:2016 (E) 2Foreword 4European foreword This document (EN ISO 12966-3:2016) has been prepared by Technical Committee ISO/TC 34 “Food pr

21、oducts“ in collaboration with Technical Committee CEN/TC 307 “Oilseeds, vegetable and animal fats and oils and their by-products - Methods of sampling and analysis” the secretariat of which is held by AFNOR. This European Standard shall be given the status of a national standard, either by publicati

22、on of an identical text or by endorsement, at the latest by December 2016, and conflicting national standards shall be withdrawn at the latest by December 2016. 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 sh

23、all not be held responsible for identifying any or all such patent rights. This document supersedes EN ISO 12966-3:2009. According to the CEN-CENELEC Internal Regulations, the national standards organizations of the following countries are bound to implement this European Standard: Austria, Belgium,

24、 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, Switzerlan

25、d, Turkey and the United Kingdom. Endorsement notice The text of ISO 12966-3:2016 has been approved by CEN as EN ISO 12966-3:2016 without any modification. DIN EN ISO 12966-3:2016-11 EN ISO 12966-3:2016 (E) 3 ForewordISO (the International Organization for Standardization) is a worldwide federation

26、of national standards bodies (ISO member bodies). The work of preparing International Standards is normally carried out through ISO technical committees. Each member body interested in a subject for which a technical committee has been established has the right to be represented on that committee. I

27、nternational organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.The procedures used to develop this document and those

28、intended for its further maintenance are described in the ISO/IEC Directives, Part 1. In particular the different approval criteria needed for the different types of ISO documents should be noted. This document was drafted in accordance with the editorial rules of the ISO/IEC Directives, Part 2 (see

29、 www.iso.org/directives).Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of any patent rights identified during the development of the docume

30、nt will be in the Introduction and/or on the ISO list of patent declarations received (see www.iso.org/patents). Any trade name used in this document is information given for the convenience of users and does not constitute an endorsement.For an explanation on the meaning of ISO specific terms and e

31、xpressions related to conformity assessment, as well as information about ISOs adherence to the WTO principles in the Technical Barriers to Trade (TBT) see the following URL: Foreword - Supplementary informationThe committee responsible for this document is ISO/TC 34, Food products, Subcommittee SC

32、11, Animal and vegetable fats and oils.This second edition cancels and replaces the first edition (ISO 12966-3:2009), of which it constitutes a minor revision. The scope has been revised to state that the document is not applicable to milk and milk fat products.ISO 12966 consists of the following pa

33、rt, under the general title Animal and vegetable fats and oils Gas chromatography of fatty acid methyl esters: Part 1: Guidelines on modern gas chromatography of fatty acid methyl esters Part 2: Preparation of methyl esters of fatty acids Part 3: Preparation of methyl esters using trimethylsulfonium

34、 hydroxide (TMSH) Part 4: Determination by capillary gas chromatographyDIN EN ISO 12966-3:2016-11 EN ISO 12966-3:2016 (E) 4 1 ScopeThis part of ISO 12966 specifies a rapid base-catalysed transesterification method for fats and oils with trimethylsulfonium hydroxide (TMSH) to prepare fatty acid methy

35、l esters. The method is exclusively applicable to the preparation of methyl esters of fats and oils for gas liquid chromatographic (GLC) analysis. It is applicable to all fats and oils, but excluding those coming from milk and milk products. Isomerization of unsaturated fatty acids only occurs to a

36、minor extent and isomerized fatty acids are only present at the determination limit. As isomerization takes place, the procedure is not recommended for conjugated linoleic acid (CLA).Only about 70 % to 80 % of the free fatty acids are esterified. In the case of conjugated cyclopropyl and cyclopropen

37、yl fatty acids, side reactions may occur, but these do not interfere with the determination of the fatty acids.NOTE This part of ISO 12966 is based upon German Standard Method C-VI 11e (98) (see Reference 8).2 Normative referencesThe following documents, in whole or in part, are normatively referenc

38、ed in this document and indispensable for its application. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies.ISO 661, Animal and vegetable fats and oils Preparation of test sample3 PrincipleT

39、he sample is dissolved in tert-butyl methyl ether (TBME) and mixed with a methanolic solution of trimethylsulfonium hydroxide. Glycerides are base-catalysed transesterified and fatty acid methyl esters are formed (see References 4 to 8). Free fatty acids are converted to salts which are pyrolysed to

40、 methyl esters and dimethylsulfide in the injector. Excess reagent is also pyrolysed into methanol and dimethylsulfide. To obtain a complete pyrolytic reaction, a hot injector (split injection) of at least 250 C is necessary.For the determination of short-chain fatty acids (C4to C8), valeric acid me

41、thyl ester is used as an internal standard. Lipids containing hydroxy groups are partially converted to the corresponding O-methyl ether derivatives which may interfere with fatty acid methyl esters in the GLC separation (Reference 9). In the early part of the chromatogram (region of C4), peaks may

42、occur, which are from the reagent. These peaks are not taken into account.4 ReagentsWARNING Attention is drawn to the regulations which specify the handling of hazardous substances. Technical, organizational, and personal safety measures shall be followed.DIN EN ISO 12966-3:2016-11 EN ISO 12966-3:20

43、16 (E) 5 During the analysis, unless otherwise stated, use only reagents and solvents of recognized analytical grade.4.1 tert-Butyl methyl ether (TBME).4.2 Trimethylsulfonium hydroxide (TMSH), methanolic solution, amount of substance concentration c(Me3SOH) = 0,2 mol/l.The content of the solution ca

44、n be determined by acidimetry: dilute 5,0 ml of the solution with 10 ml of methanol, add two drops of phenolphthalein and titrate against 0,1 mol/l HCl. The concentration should be at least 0,15 mol/l.NOTE The solution remains stable for at least two months when stored at 4 C in small quantities in

45、closed tubes.5 ApparatusUsual laboratory equipment, and in particular:5.1 Test tubes, of capacity 2 ml (autosampler vials).5.2 Graduated pipettes, of capacities 250 l, 500 l, and 1 000 l, ISO 8351class A.6 SamplingA representative sample should have been sent to the laboratory. It should not have be

46、en damaged or changed during transport or storage.Sampling is not part of the method specified in this part of ISO 12966. A recommended sampling method is given in ISO 55553.7 Preparation of the test sampleThe test sample shall be liquid, dry and clear. Proceed in accordance with ISO 661, but heat t

47、he sample to just above the melting point.Solid samples are carefully melted at a temperature of maximum 10 C above their melting point and mixed. Avoid overheating.Samples containing water are dissolved in petroleum ether and dried for 30 min by addition of anhydrous sodium sulfate. The drying agen

48、t is removed by filtration through a fluted filter paper and the residue is carefully washed with petroleum ether. The solvent is subsequently removed with the aid of a rotary evaporator.8 Procedure8.1 Weigh, into a test tube (5.1), (10 2) mg of the test sample.8.2 Pipette (5.2) 500 l of TBME (4.1) into the test tube and dissolve the sample, warming gently if necessary.NOTE For the determination of short-chain fatty acids (C4to C8), valeric acid methyl ester i