1、BSI Standards PublicationBS ISO 16958:2015Milk, milk products, infantformula and adult nutritionals Determination of fatty acidscomposition Capillary gaschromatographic methodBS ISO 16958:2015 BRITISH STANDARDNational forewordThis British Standard is the UK implementation of ISO 16958:2015.The UK pa
2、rticipation in its preparation was entrusted to TechnicalCommittee AW/5, Chemical analysis of milk and milk products.A list of organizations represented on this committee can beobtained on request to its secretary.This publication does not purport to include all the necessaryprovisions of a contract
3、. Users are responsible for its correctapplication. The British Standards Institution 2015.Published by BSI Standards Limited 2015ISBN 978 0 580 87478 9ICS 67.100.10Compliance with a British Standard cannot confer immunity fromlegal obligations.This British Standard was published under the authority
4、 of theStandards Policy and Strategy Committee on 30 November 2015.Amendments/corrigenda issued since publicationDate T e x t a f f e c t e dBS ISO 16958:2015Milk, milk products, infant formula and adult nutritionals Determination of fatty acids composition Capillary gas chromatographic methodLait,
5、produits laitiers, formules infantiles et produits nutritionnels pour adultes Dtermination de la composition en acides gras Mthode de chromatographie en phase gazeuse sur colonne capillaireINTERNATIONAL STANDARDFirst edition2015-11-01Reference numbersISO 16958:2015(E)IDF 231:2015(E)ISO16958IDF231 IS
6、O and IDF 2015BS ISO 16958:2015ISO 16958:2015(E)IDF 231:2015(E)ii ISO and IDF 2015 All rights reservedCOPYRIGHT PROTECTED DOCUMENT ISO and IDF 2015, Published in SwitzerlandAll rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized otherwise in any for
7、m or by any means, electronic or mechanical, including photocopying, or posting on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address below or ISOs member body in the country of the requester.ISO copyright office International Da
8、iry FederationCh. de Blandonnet 8 CP 401 Silver Building Bd Auguste Reyers 70/B B-1030 BrusselsCH-1214 Vernier, Geneva, SwitzerlandTel. +41 22 749 01 11 Tel. + 32 2 325 67 40Fax +41 22 749 09 47 Fax + 32 2 325 67 41copyrightiso.org infofil-idf.orgwww.iso.org www.fil-idf.orgBS ISO 16958:2015ISO 16958
9、:2015(E)IDF 231:2015(E)Forewords .iv1 Scope . 12 Normative references 13 Terms and definitions . 14 Principle 25 Reagents 26 Apparatus . 67 Sampling 88 Preparation of test sample . 98.1 Liquid and powder milk and infant formula with a fat content 1,5 % m/m . 98.2 Liquid and powder milk and infant fo
10、rmula with a fat content 102,0 % or 98,0 % the origin of the problem could be the following: incomplete transesterification (i.e. problem with reagent/chemical); partial degradation of internal standard(s), or problem with their purity/stability; sample matrix effect problem.NOTE The analysis of a r
11、eference sample can help to determine if the problem originates from reagents and/or chemicals or with the analysed sample.10.2 Expression of resultsExpress to three decimal places the results expressed in g / 100 g and to one decimal place the results expressed in mg / 100 g.NOTE Fatty acids result
12、s expressed in g (or mg)/100 g product can be converted in other results expression format g (or mg) fatty acids /100 g fat: g (or mg) fatty acids / 100 g reconstituted powder (i.e. 25 g powder into 200 g water), liquid product (i.e. ready-to-feed), or liquid concentrates diluted 1:1 by mass. g (or
13、mg) fatty acids / serving size (according to serving size mass).11 Precision11.1 Interlaboratory testDetails of the interlaboratory studies, organized and elaborated in accordance with ISO 5725-1 and ISO 5725-2 on the precision of the method, are summarized in Annex C.The values for repeatability an
14、d reproducibility limit are expressed for the 95 % probability level and may not be applicable to concentration ranges and matrices other than those given.11.2 RepeatabilityThe absolute difference between two independent single test results, obtained using the same method on identical test material
15、in the same laboratory by the same operator using the same equipment IDF 231:2015(E)14 ISO and IDF 2015 All rights reservedBS ISO 16958:2015ISO 16958:2015(E)within a short interval of time, shall in not more than 5 % of cases be greater than r, as given in Annex C, Tables C.1 and C.2.11.3 Reproducib
16、ilityThe absolute difference between two independent single test results, obtained using the same method on identical test material in different laboratories with different operators using different equipment, shall in not more than 5 % of cases be greater than R, as given in Annex C, Tables C.1 and
17、 C.2.11.4 Limit of detectionUnder the described conditions (detector sensitivity, noise, sample dilution, etc.), the estimated detection limit expressed as three times the standard deviation of the background signal (noise) lies around 0,0003 g/100 g of product.11.5 Limit of quantitationThe limit of
18、 quantification, for each fatty acid, is about 0,001 g/100 g of product. The limit of quantification corresponds to the lowest level where the robust repeatability was calculated with satisfactory results.12 Test reportThe test report shall include at least the following information:a) all informati
19、on necessary for the complete identification of the sample;b) the sampling method used, if known; with reference to this International Standard, i.e. ISO 16958 | IDF 231;c) the test method used, together with reference to this International Standard;d) all operating details not specified in this Int
20、ernational Standard, or regarded as optional, together with details of any incidents which may have influenced the test result(s);e) the test result(s) obtained.IDF 231:2015(E) ISO and IDF 2015 All rights reserved 15BS ISO 16958:2015ISO 16958:2015(E)Annex A (normative) Groups or classes of fatty aci
21、ds and individual fatty acidsA.1 Group or class of fatty acidsA.1.1 Trans fatty acidsTrans fatty acids (TFA) is the sum of fatty acids containing one or more non conjugated double bonds in trans configuration (only C18:1, C18:2 and C18:3 trans are included in the sum).NOTE Presence of other trans is
22、omers naturally present in milk fat have been reported in the literature (e.g. C16:1 trans), but their contributions have no significant influence on the total amount of trans fatty acids in milk products. In addition, their identification is rather complex because these isomers are frequently subje
23、ct to interferences from other isomers of fatty acids (e.g. cis, iso and anteiso) and thus need a preliminary separation or the use of specific chromatographic conditions.A.1.2 Conjugated linoleic acidsConjugated linoleic acids (CLA) is the sum of octadecadienoic acids containing conjugated double b
24、onds in cis or trans configuration; mainly cis-9, trans-11 octadecadienoic acid (i.e. rumenic acid). CLA are not included in the sum of TFAs.A.1.3 Saturated fatty acidsSaturated fatty acids (SFA) is the sum of all fatty acids without double bonds.A.1.4 Monounsaturated fatty acidsMonounsaturated fatt
25、y acids (MUFA) is the sum of all fatty acids containing one double bond in cis configuration.A.1.5 Polyunsaturated fatty acidsPolyunsaturated fatty acids (PUFA) is the sum of all fatty acids containing two or more double bonds in cis configuration.NOTE LC-PUFA is the generic name to describe the lon
26、g chain polyunsaturated fatty acids. These fatty acids are also included with PUFA (i.e. arachidonic, eicosapentaenoic and docosahexaenoic acids).A.1.6 Omega-3 fatty acidsOmega-3 fatty acids is the sum of cis polyunsaturated fatty acids having the first double bond at carbon n-3 (-3) from the termin
27、al methyl group.A.1.7 Omega-6 fatty acidsOmega-6 fatty acids is the sum of cis polyunsaturated fatty acids having the first double bond at carbon n-6 (-6) from the terminal methyl group.A.1.8 Omega-9 fatty acidsOmega-9 fatty acids is the sum of cis polyunsaturated fatty acids having the first double
28、 bond at carbon n-9 (-9) from the terminal methyl group.IDF 231:2015(E)16 ISO and IDF 2015 All rights reservedBS ISO 16958:2015ISO 16958:2015(E)A.2 Individual fatty acidsA.2.1 Linoleic acidLinoleic acid (LA) is an essential fatty acid, an 18 carbon fatty acid containing two double bonds at carbons 9
29、 and 12 (C18:2 all cis-9,12) also named C18:2 n-6 (-6).A.2.2 Linolenic acidLinolenic acid (ALA), also called -linolenic acid, is an essential fatty acid, an 18 carbon fatty acid containing three double bonds at carbons 9, 12, and 15 (C18:3 all cis-9,12,15) also named C18:3 n-3 (-3).A.2.3 Arachidonic
30、 acidArachidonic acid (ARA) is not an essential fatty acid, a 20 carbon fatty acid containing four double bonds at carbons 5, 8, 11, and 14 (C20:4 all cis-5,8,11,14) also named C20:4 n-6 (-6).A.2.4 Eicosapentaenoic acidEicosapentaenoic acid (EPA) is a semi-essential fatty acid (essential for pregnan
31、t women and infants), a 20 carbon fatty acid containing five double bonds at carbons 5, 8, 11, 14, and 17 (C20:5 all cis-5,8,11,14,17) also named C20:5 n-3 (-3).A.2.5 Docosahexaenoic acidDocosahexaenoic acid (DHA) is a semi-essential fatty acid (essential for pregnant women and infants), a 22 carbon
32、 fatty acid containing six double bonds at carbons 4, 7, 10, 13, 16, and 19 (C22:6 all cis-4,7,10,13,16,19) also named C22:6 n-3 (-3).Table A.1 Configuration and groups of fatty acidsChain lengthConfiguration and group Systematic name Trivial name AbbreviationC4:0 SFA Butanoic ButyricC6:0 SFA Hexano
33、ic CaproicC8:0 SFA Octanoic CaprylicC10:0 SFA Decanoic CapricC12:0 SFA Dodecanoic LauricC14:0 SFA Tetradecanoic MyristicC14:1 -5 (or n-5) cis MUFA 9-Tetradecenoic MyristoleicC15:0 SFA PentadecanoicC15:1 -5 (or n-5) cis MUFA 10-PentadecenoicC16:0 SFA Hexadecanoic PalmiticC16:1 -7 (or n-7) cis MUFA 9-
34、Hexadecenoic PalmitoleicC17:0 SFA HeptadecanoicC17:1 -7 (or n-7) cis MUFA 10-HeptadecenoicC18:0 SFA Octadecanoic StearicC18:1 TFA transaSum of C18:1 trans isomers All trans 4 to 16 octadecenoicC18:1 -9 (or n-9) cis MUFA 9-Octadecenoic OleicaDo not include trans fatty acids in MUFA and PUFA sums.IDF
35、231:2015(E) ISO and IDF 2015 All rights reserved 17BS ISO 16958:2015ISO 16958:2015(E)Chain lengthConfiguration and group Systematic name Trivial name AbbreviationC18:2 TFA transaSum of C18:2 trans isomers All trans 9,12 octadecadienoic in deodorized oils and trans originated from milk fat (i.e. C18:
36、2 cis-9,trans-13, C18:2 trans-8,cis-12 and C18:2 trans-11,cis-15)C18:2 -6 (or n-6) cis PUFA 9,12-Octadecadienoic Linoleic LAC18:2 CLA -7 (or n-7) cis/trans PUFA 9,11-Octadecadienoic Rumenic CLAC18:3 -6 (or n-6) cis PUFA 6,9,12-Octadecatrienoic Gamma-linolenicC18:3 TFA transaSum of C18:3 trans isomer
37、s All trans 9,12,15 Octadecatrie-noicC18:3 -3 (or n-3) cis PUFA 9,12,15-Octadecatrienoic Linolenic ALAC20:0 SFA Eicosanoic ArachidicC20:1 -9 (or n-9) cis MUFA 11-Eicosenoic GondoicC20:2 -6 (or n-6) cis PUFA 11,14-EicosadienoicC20:3 -6 (or n-6) cis PUFA 8,11,14-Eicosatrienoic Dihomo-gamma-linolenic (
38、DHGLA)C20:3 -3 (or n-3) cis PUFA 11,14,17-EicosatrienoicC20:4 -6 (or n-6) cis PUFA 5,8,11,14-Eicosatetrae-noicArachidonic ARAC20:5 -3 (or n-3) cis PUFA 5,8,11,14,17-Eicosapen-taenoicEicosapentaenoic EPAC21:0 SFA HeneicosanoicC22:0 SFA Docosanoic BehenicC22:1 -9 (or n-9) cis MUFA 13-Docosenoic Erucic
39、C22:2 -6 (or n-6) cis PUFA 13,16-DocosadienoicC22:6 -3 (or n-3) cis PUFA 4,7,10,13,16,19-Docosa-hexaenoicDocosahexaenoic DHAC24:0 SFA Tetracosanoic LignocericC24:1 -9 (or n-9) cis MUFA 15-Tetracosenoic NervonicaDo not include trans fatty acids in MUFA and PUFA sums.IDF 231:2015(E)Table A.1 (continue
40、d)18 ISO and IDF 2015 All rights reservedBS ISO 16958:2015ISO 16958:2015(E)Table A.2 AbbreviationsFAME Fatty acid methyl ester MUFA Monounsaturated fatty acidsFA Fatty acid PUFA Polyunsaturated fatty acidsGLC Gas liquid chromatography LC-PUFA Long chain polyunsaturated fatty acidsMTBE Tert-Butyl met
41、hyl ester -3 (or n-3) Omega-3 fatty acidsMeOH Methanol -6 (or n-6) Omega-6 fatty acidsR Resolution factor -9 (or n-9) Omega-9 fatty acidsRF Response factor LA Linolenic acid (C18:2 all cis-9,12 or n-6)RT Retention time ALA Linolenic acid (C18:3 all cis-9,12,15 or n-3) also called -linolenic acidTAG
42、Triacylglycerol ARA (AA) Arachidonic acid (C20:4 all cis-5,8,11,14,17 or n-6)TFA Trans fatty acid EPA Eicosapentaenoic acid (C20:9 all cis-5,8,11,14,17 or n-3)CLA Conjugated linoleic acid (C18:2 cis-9,trans-11, also called rumenic acidDHA Docosahexaenoic acid (C22:6 all cis-4,7,10,13,16,19 or n-3)SF
43、A Saturated fatty acids OFA Other fatty acids sum of unknowns (i.e. unidentified), less important, less abundant or not considered (i.e. branched) fatty acids. OFA are not included into TFA, SFA, MUFA and PUFA sums.IDF 231:2015(E) ISO and IDF 2015 All rights reserved 19BS ISO 16958:2015ISO 16958:201
44、5(E)Annex B (informative) Examples of the gasliquid chromatographic analysisFigure B.1 Example of GC chromatogram of milk product (enlarged view of C18:1 TFA, C18:2 TFA, C18:3 TFA and CLA) using split injectionIDF 231:2015(E)20 ISO and IDF 2015 All rights reservedBS ISO 16958:2015ISO 16958:2015(E)Fi
45、gure B.2 Example of GC chromatogram of milk product (enlarged view of C18:1 TFA, C18:2 TFA, C18:3 TFA and CLA) using on-column injectionIDF 231:2015(E) ISO and IDF 2015 All rights reserved 21BS ISO 16958:2015ISO 16958:2015(E)Figure B.3 Example of GC chromatogram (insufficient and sufficient resolution between C18:1 cis and trans isomers)IDF 231:2015(E)22 ISO and IDF 2015 All rights reserved
