BS DD ISO TS 27106-2010 Cheese - Determination of nisin A content by LC-MS and LC-MS-MS《乳酪 用LC-MS和LC-MS-MS对乳酸链球菌素A含量的测定》.pdf

1、DD ISO/TS27106:2009ICS 67.100.30NO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAWDRAFT FOR DEVELOPMENTCheese Determination of nisinA content by LC-MSand LC-MS-MSThis Draft for Developmentwas published under theauthority of the StandardsPolicy and StrategyCommittee on 31 March201

2、0 BSI 2010ISBN 978 0 580 64720 8Amendments/corrigenda issued since publicationDate CommentsDD ISO/TS 27106:2009National forewordThis Draft for Development is the UK implementation of ISO/TS27106:2009.This publication is not to be regarded as a British Standard.It is being issued in the Draft for Dev

3、elopment series of publications andis of a provisional nature. It should be applied on this provisional basis,so that information and experience of its practical application can beobtained.Comments arising from the use of this Draft for Development arerequested so that UK experience can be reported

4、to the internationalorganization responsible for its conversion to an international standard.A review of this publication will be initiated not later than 3 years afterits publication by the international organization so that a decision can betaken on its status. Notification of the start of the rev

5、iew period will bemade in an announcement in the appropriate issue of Update Standards.According to the replies received by the end of the review period,the responsible BSI Committee will decide whether to support theconversion into an international Standard, to extend the life of theTechnical Speci

6、fication or to withdraw it. Comments should be sent tothe Secretary of the responsible BSI Technical Committee at BritishStandards House, 389 Chiswick High Road, London W4 4AL.The UK participation in its preparation was entrusted to TechnicalCommittee AW/5, Chemical analysis of milk and milk product

7、s.A list of organizations represented on this committee can be obtained onrequest to its secretary.This publication does not purport to include all the necessary provisionsof a contract. Users are responsible for its correct application.Compliance with a British Standard cannot confer immunityfrom l

8、egal obligations.DD ISO/TS 27106:2009Reference numbersISO/TS 27106:2009(E)IDF/RM 217:2009(E)ISO and IDF 2009TECHNICAL SPECIFICATION ISO/TS27106IDF/RM217First edition2009-12-01Cheese Determination of nisin A content by LC-MS and LC-MS-MS Fromage Dtermination de la teneur en nisine A par CL-SM et CL-S

9、M-SM DD ISO/TS 27106:2009ISO/TS 27106:2009(E) IDF/RM 217:2009(E) PDF disclaimer This PDF file may contain embedded typefaces. In accordance with Adobes licensing policy, this file may be printed or viewed but shall not be edited unless the typefaces which are embedded are licensed to and installed o

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13、onic or mechanical, including photocopying and microfilm, without permission in writing from either ISO or IDF at the respective address below. ISO copyright office International Dairy Federation Case postale 56 CH-1211 Geneva 20 Diamant Building Boulevard Auguste Reyers 80 B-1030 Brussels Tel. + 41

14、 22 749 01 11 Tel. + 32 2 733 98 88 Fax + 41 22 749 09 47 Fax + 32 2 733 04 13 E-mail copyrightiso.org E-mail infofil-idf.org Web www.iso.org Web www.fil-idf.org Published in Switzerland ii ISO and IDF 2009 All rights reservedDD ISO/TS 27106:2009ISO/TS 27106:2009(E) IDF/RM 217:2009(E) ISO and IDF 20

15、09 All rights reserved iiiForeword ISO (the International Organization for Standardization) is a worldwide federation 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

16、in a subject for which a technical committee has been established has the right to be represented on that committee. International organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the International Electrotechnical Commi

17、ssion (IEC) on all matters of electrotechnical standardization. International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2. The main task of technical committees is to prepare International Standards. Draft International Standards adopted by the technica

18、l committees are circulated to the member bodies for voting. Publication as an International Standard requires approval by at least 75 % of the member bodies casting a vote. In other circumstances, particularly when there is an urgent market requirement for such documents, a technical committee may

19、decide to publish other types of document: an ISO Publicly Available Specification (ISO/PAS) represents an agreement between technical experts in an ISO working group and is accepted for publication if it is approved by more than 50 % of the members of the parent committee casting a vote; an ISO Tec

20、hnical Specification (ISO/TS) represents an agreement between the members of a technical committee and is accepted for publication if it is approved by 2/3 of the members of the committee casting a vote. An ISO/PAS or ISO/TS is reviewed after three years in order to decide whether it will be confirm

21、ed for a further three years, revised to become an International Standard, or withdrawn. If the ISO/PAS or ISO/TS is confirmed, it is reviewed again after a further three years, at which time it must either be transformed into an International Standard or be withdrawn. Attention is drawn to the poss

22、ibility 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. ISO/TS 27106IDF/RM 217 was prepared by Technical Committee ISO/TC 34, Food products, Subcommittee SC 5, Milk and milk products, and

23、 the International Dairy Federation (IDF). It is being published jointly by ISO and IDF. DD ISO/TS 27106:2009ISO/TS 27106:2009(E) IDF/RM 217:2009(E) iv ISO and IDF 2009 All rights reservedForeword IDF (the International Dairy Federation) is a non-profit organization representing the dairy sector wor

24、ldwide. IDF membership comprises National Committees in every member country as well as regional dairy associations having signed a formal agreement on cooperation with IDF. All members of IDF have the right to be represented on the IDF Standing Committees carrying out the technical work. IDF collab

25、orates with ISO in the development of standard methods of analysis and sampling for milk and milk products. The main task of Standing Committees is to prepare International Standards. Draft International Standards adopted by the Standing Committees are circulated to the National Committees for endor

26、sement prior to publication as an International Standard. Publication as an International Standard requires approval by at least 50% of IDF National Committees casting a vote. In other circumstances, particularly when there is an urgent market requirement for such documents, a Standing Committee may

27、 decide to publish an other type of normative document which is called by IDF: Reviewed method. Such a method represents an agreement between the members of a Standing Committee and is accepted for publication if it is approved by at least 50 % of the committee members casting a vote. A Reviewed met

28、hod is equal to an ISO/PAS or ISO/TS and will, therefore, also be published jointly under ISO conditions. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. IDF shall not be held responsible for identifying any or all such patent rig

29、hts. ISO/TS 27106IDF/RM 217 was prepared by the International Dairy Federation (IDF) and Technical Committee ISO/TC 34, Food products, Subcommittee SC 5, Milk and milk products. It is being published jointly by IDF and ISO. All work was carried out by the Joint ISO-IDF Action Team on Food additives

30、and vitamins of the Standing Committee on Analytical methods for additives and contaminants under the aegis of its project leader, Mr. T. Berger (CH). DD ISO/TS 27106:2009TECHNICAL SPECIFICATION ISO/TS 27106:2009(E)IDF/RM 217:2009(E) ISO and IDF 2009 All rights reserved 1Cheese Determination of nisi

31、n A content by LC-MS and LC-MS-MS 1 Scope This Technical Specification specifies a method for the quantitative determination of the nisin A content in cheese. The method is suitable for measuring low levels of nisin A with a quantification limit of 1 mg/kg. NOTE Nisin is a peptide produced by some b

32、acteria (e.g. Lactococcus lactis subsp. Lactis) inhibiting or destroying other microorganisms. It is widely used as a natural preservative for foods, e.g. vegetables, cheese, meat, and cacao. In cheese making, nisin is used to prevent late blowing. Its use is restricted to maximum levels in the fina

33、l product. Nisin appears in two forms, nisin A and nisin Z, which differ in one amino acid. This method is applicable to the determination of nisin A only. 2 Terms and definitions For the purposes of this document, the following terms and definitions apply. 2.1 nisin A content mass fraction of subst

34、ance determined by the procedure specified in this Technical Specification NOTE The nisin A content is expressed in milligrams per kilogram. 3 Principle The sample is grated and extracted with dilute formic acid at 80 C. After ultracentrifugation, interfering proteins are separated by means of filtr

35、ation through an ultrafiltration (UF) membrane. In the purified extract, nisin A is separated using a polymeric stationary phase and detected using mass spectrometry (with MS-MS as an option). 4 Reagents and reference substances Use only reagents of recognized analytical grade and distilled water or

36、 water of at least equivalent purity, unless otherwise specified. 4.1 Bovine serum albumin (BSA) stock solution. Dissolve 10 mg of BSA (purity 96 % mass fraction), in 10 ml water. 4.2 Bovine serum albumin (BSA) buffer solution. Mix 80 ml water with 20 ml of acetonitrile (4.6), 0,5 ml of formic acid

37、(4.3), 0,01 ml of trifluoracetic acid (4.5) and 1 ml of BSA stock solution (4.1). 4.3 Formic acid (HCOOH). 4.4 Formic acid solution, HCOOH= 5 g/l. Pipette 0,41 ml of formic acid (4.3) into a 100 ml one-mark volumetric flask (5.12). Make up to the mark with water and mix. DD ISO/TS 27106:2009ISO/TS 2

38、7106:2009(E) IDF/RM 217:2009(E) 2 ISO and IDF 2009 All rights reserved4.5 Trifluoracetic acid (CF3COOH). 4.6 Acetonitrile (CH3CN), “pure”. 4.7 Methanol (CH3OH). 4.8 Nisin A, of purity 95 % mass fraction1). 5 Apparatus Usual laboratory equipment and, in particular, the following. 5.1 Laboratory centr

39、ifuge, capable of producing a radial acceleration of at least 3 000g. 5.2 Ultracentrifuge, capable of producing a radial acceleration of 20 800g. 5.3 Ultrafiltration membrane, of pore size 30 kD2). 5.4 Membrane filter, of pore size 0,22 m3). 5.5 Balance, capable of weighing to the nearest 10 mg, wit

40、h a readability of 1 mg. 5.6 Analytical balance, capable of weighing to the nearest 0,1 mg, with a readability of 0,01 mg. 5.7 Water bath, capable of shaking and maintaining a temperature of 80 C 2 C. 5.8 Ultrasonic bath, capable of shaking and maintaining a temperature of 80 C 2 C. 5.9 Cheese grate

41、r, with openings of size approximately 2 mm. 5.10 LC-MS equipment. 5.10.1 Elution gradient pumping system, capable of operating at 0,25 ml/min. 5.10.2 Manual or automatic injector, capable of injecting volumes of 5 l. 5.10.3 Column heater, capable of maintaining a column temperature of 40 C 2 C. 5.1

42、0.4 Column, reversed phase, PLRP-S, 300 4), 3 m, 150 mm 2 mm. 5.10.5 Mass spectrometer detector, capable of operating in ion mode ESI+ at m/z 839,6. 5.11 LC-MS-MS equipment (optional). 1) Ambicin N is an example of a suitable product available commercially. This information is given for the convenie

43、nce of users of this document and does not constitute an endorsement by ISO or IDF of this product. 2) Millipore Centricon YM-30 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

44、by ISO or IDF of this product. 3) Millipore Millex-GV PVDF 0,22 m 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 ISO or IDF of this product. 4) PLRP-S, 300 is the trade name

45、 of a product supplied by Polymer Lab Ltd. This information is given for the convenience of users of this document and does not constitute an endorsement by ISO or IDF of the product named. Equivalent products may be used if they can be shown to lead to the same results. DD ISO/TS 27106:2009ISO/TS 2

46、7106:2009(E) IDF/RM 217:2009(E) ISO and IDF 2009 All rights reserved 35.11.1 Elution gradient pumping system, capable of delivering a flow rate of 0,2 ml/min. 5.11.2 Manual or automatic injector, capable of injecting volumes of 10 l. 5.11.3 Column heater, capable of maintaining a column temperature

47、of 30 C 2 C. 5.11.4 Column for reversed phase chromatography, PLRP-S, 300 4), 3 m, 150 2 mm. 5.11.5 Mass spectrometer detector, capable of operating in ion mode ESI+ MS-MS at m/z 672/672, 672/811, 672/649, 840/840. 5.12 One-mark volumetric flasks, of capacity 100 ml, ISO 10422class A. 6 Sampling A r

48、epresentative sample should have been sent to the laboratory. It should not have been damaged or changed during transport or storage. Sampling is not part of the method specified in this Technical Specification. A recommended sampling method is given in ISO 707IDF 501. 7 Procedure 7.1 Preparation of

49、 the nisin A standard solution 7.1.1 Nisin A standard stock solution, nA= 100 mg/l. Weigh, to the nearest 0,01 mg, 10,00 mg nisin A (4.8) into a 100 ml one-mark volumetric flask (5.12). Make up to the mark with formic acid solution (4.4) and mix. Prepare the nisin A standard stock solution daily. 7.1.2 Nisin A standard working solution, nA= 300 g/l. Pipette 300 l of nisin A standard stock solution (7.1.1) into a 100 ml one-mark volumetric flask (5.12). Make up to the mark with BSA buffer solution (4.2)