1、BSI Standards PublicationBS EN ISO 4833-1:2013Microbiology of the foodchain Horizontal methodfor the enumeration ofmicroorganismsPart 1: Colony count at 30 degrees C by thepour plate techniqueBS EN ISO 4833-1:2013 BRITISH STANDARDNational forewordThis British Standard is the UK implementation of EN
2、ISO 4833-1:2013. Together with BS EN ISO 4833-2:2013 it supersedes BS EN ISO 4833:2003, which is withdrawn.The UK participation in its preparation was entrusted to Technical Committee AW/9, Microbiology.A list of organizations represented on this committee can be obtained on request to its secretary
3、.This publication does not purport to include all the necessary provisions of a contract. Users are responsible for its correct application. The British Standards Institution 2013. Published by BSI Standards Limited 2013ISBN 978 0 580 71933 2 ICS 07.100.30 Compliance with a British Standard cannot c
4、onfer immunity from legal obligations.This British Standard was published under the authority of the Standards Policy and Strategy Committee on 31 October 2013.Amendments issued since publicationDate T e x t a f f e c t e dEUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN ISO 4833-1 September 201
5、3 ICS 07.100.30 Supersedes EN ISO 4833:2003English Version Microbiology of the food chain - Horizontal method for the enumeration of microorganisms - Part 1: Colony count at 30 degrees C by the pour plate technique (ISO 4833-1:2013) Microbiologie de la chane alimentaire - Mthode horizontale pour le
6、dnombrement des micro-organismes -Partie 1: Comptage des colonies 30 degrs C par la technique densemencement en profondeur (ISO 4833-1:2013) Mikrobiologie von Lebensmitteln und Futtermitteln - Horizontales Verfahren zur Zhlung von Mikroorganismen -Teil 1: Koloniezhlverfahren bei 30 C mittels Gusspla
7、ttenverfahren (ISO 4833-1:2013) This European Standard was approved by CEN on 26 July 2013. 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
8、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 versions (English, French, German). A version in any other language made by translation un
9、der 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 Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, Former
10、 Yugoslav 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 United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EU
11、ROPEN DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG CEN-CENELEC 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 Members. Ref. No. EN ISO 4833-1:2013: EBS EN ISO 4833-1:2013EN ISO 4833-1:2013 (
12、E) 3 Foreword This document (EN ISO 4833-1:2013) has been prepared by Technical Committee ISO/TC 34 “Foodproducts“ in collaboration with Technical Committee CEN/TC 275 “Food analysis - Horizontal methods” thesecretariat of which is held by DIN. This European Standard shall be given the status of a n
13、ational standard, either by publication of an identicaltext or by endorsement, at the latest by March 2014, and conflicting national standards shall be withdrawn atthe latest by March 2014. Attention is drawn to the possibility that some of the elements of this document may be the subject of patentr
14、ights. CEN and/or CENELEC shall not be held responsible for identifying any or all such patent rights. This document supersedes EN ISO 4833:2003, together with EN ISO 4833-2:2013. According to the CEN-CENELEC Internal Regulations, the national standards organizations of the followingcountries are bo
15、und to implement this European Standard: Austria, Belgium, Bulgaria, Croatia, Cyprus, CzechRepublic, Denmark, Estonia, Finland, Former Yugoslav Republic of Macedonia, France, Germany, Greece,Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal
16、,Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United Kingdom. Endorsement notice The text of ISO 4833-1:2013 has been approved by CEN as EN ISO 4833-1:2013 without any modification. BS EN ISO 4833-1:2013ISO 4833-1:2013(E) ISO 2013 All rights reserved iiiContents PageForewo
17、rd iv1 Scope . 12 Normative references 13 Terms and definitions . 14 Principle 25 Culture media and diluents . 25.1 General . 25.2 Diluents . 25.3 Agar medium: plate count agar (PCA) 25.4 Overlay medium (if necessary; see 9.2.7) 36 Apparatus . 47 Sampling 48 Preparation of test sample . 49 Procedure
18、. 49.1 Test portion, initial suspension and dilutions 49.2 Inoculation and incubation 49.3 Counting of colonies . 510 Expression of results 510.1 Method of calculation 510.2 Precision . 510.3 Interpretation of test results . 611 Test report . 7Annex A (informative) Use of the critical difference for
19、 the interpretation of results . 8Bibliography 9BS EN ISO 4833-1:2013ISO 4833-1:2013(E)ForewordISO (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 th
20、rough 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. International organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collab
21、orates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization.The procedures used to develop this document and those intended for its further maintenance are described in the ISO/IEC Directives, Part 1. In particular the different approval
22、 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, www.iso.org/directives.Attention is drawn to the possibility that some of the elements of this document may be the subject of
23、 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 document will be in the Introduction and/or on the ISO list of patent declarations received, www.iso.org/patents.Any trade name used
24、in this document is information given for the convenience of users and does not constitute an endorsement.The committee responsible for this document is ISO/TC 34, Food products, Subcommittee SC 9, Microbiology.This first edition, together with ISO 4833-2, cancels and replaces ISO 4833:2003.ISO 4833
25、 consists of the following parts, under the general title Microbiology of the food chain Horizontal method for the enumeration of microorganisms: Part 1: Colony count at 30 C by the pour plate technique Part 2: Colony count at 30 C by the surface plating techniqueiv ISO 2013 All rights reservedBS EN
26、 ISO 4833-1:2013INTERNATIONAL STANDARD ISO 4833-1:2013(E)Microbiology of the food chain Horizontal method for the enumeration of microorganisms Part 1: Colony count at 30 C by the pour plate technique1 ScopeThis part of ISO 4833 specifies a horizontal method for enumeration of microorganisms that ar
27、e able to grow and form colonies in a solid medium after aerobic incubation at 30 C. The method is applicable to:a) products intended for human consumption and for animal feed;b) environmental samples in the area of food and feed production and handling.This part of ISO 4833 is applicable to:1) prod
28、ucts that require a reliable count when a low limit of detection is specified (below 102/g or 102/ml for liquid samples or below 103/g for solid samples);2) products expected to contain spreading colonies that obscure colonies of other organisms, e.g. milk and milk products likely to contain spreadi
29、ng Bacillus spp.The applicability of this part of ISO 4833 to the examination of certain fermented food and animal feeds is limited and other media or incubation conditions can be more appropriate. However, this method can be applied to such products even though it is possible that the predominant m
30、icroorganisms in those products are not detected effectively.For some matrices, the method specified in this part of ISO 4833 can give different results to those obtained using the method specified in ISO 4833-2.2 Normative referencesThe following documents, in whole or in part, are normatively refe
31、renced in this document and are 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 6887 (all parts), Microbiology of food and animal feeding stuffs Prepa
32、ration of test samples, initial suspension and decimal dilutions for microbiological examinationISO 7218, Microbiology of food and animal feeding stuffs General requirements and guidance for microbiological examinationsISO 11133, Microbiology of food, animal feed and water Preparation, production, s
33、torage and performance testing of culture media3 Terms and definitionsFor the purposes of this document, the following terms and definitions apply. ISO 2013 All rights reserved 1BS EN ISO 4833-1:2013ISO 4833-1:2013(E)3.1microorganismentity of microscopic size, encompassing bacteria, fungi, protozoa
34、and virusesSOURCE: ISO/TS 11139:2006,32.26Note 1 to entry: For the purposes of this part of ISO 4833, microorganisms are bacteria, yeasts and moulds that are able to produce colonies under the conditions specified in this part of ISO 4833.4 PrincipleA specified quantity of the liquid test sample, or
35、 a specified quantity of an initial suspension in the case of other products, is dispensed into an empty Petri dish and mixed with a specified molten agar culture medium to form a poured plate.Other plates are prepared under the same conditions using decimal dilutions of the test sample or of the in
36、itial suspension.The plates are incubated under aerobic conditions at 30 C for 72 h.The number of microorganisms per gram or per millilitre of the test sample is calculated from the number of colonies obtained in the plates containing fewer than 300 colonies.5 Culture media and diluents5.1 GeneralFo
37、llow ISO 11133 for preparation, production and performance testing of culture media.5.2 DiluentsUse the diluent(s) specified in ISO 6887 for the product concerned or the specific International Standard dealing with the product under examination.5.3 Agar medium: plate count agar (PCA)5.3.1 Compositio
38、nEnzymatic digestion of casein 5,0 gYeast extract 2,5 gGlucose, anhydrous (C6H12O6) 1,0 gAgara9 g to 18 gWater 1 000 mlaDepending on the gel strength of the agar.When dairy products are examined, add skimmed milk powder at 1,0 g/l of the culture medium. The skimmed milk powder shall be free from inh
39、ibitory substances.5.3.2 PreparationDissolve the components or the dehydrated complete medium in the water, by heating if necessary. Mix thoroughly and leave to stand for several minutes.2 ISO 2013 All rights reservedBS EN ISO 4833-1:2013ISO 4833-1:2013(E)Adjust the pH (6.4), if necessary, so that a
40、fter sterilization it is 7,0 0,2 at 25 C.Dispense the medium into tubes, flasks or bottles (6.8) of suitable capacity. Sterilize in an autoclave (6.1) at 121 C for 15 min.lf the medium is to be used immediately, cool it to 44 C to 47 C in a water bath (6.3) before use. If not, store it in the dark a
41、t a temperature of (5 3) C for no longer than 3 months, under conditions which do not allow any change in its composition and properties.Before beginning the microbiological examination, completely melt the medium, then cool it to 44 C to 47 C in a water bath (6.3) before use. See ISO 11133.Use the
42、molten agar as soon as possible; it should not be retained for more than 4 h.5.3.3 Performance testing of the culture medium5.3.3.1 GeneralPlate count agar is a non-selective medium, used in this part of ISO 4833 as a pour plate. Productivity shall be tested according to ISO 11133.5.3.3.2 Productivi
43、tyIncubation (30 1) C for (72 3) h Control strains Escherichia coli WDCM 00013 or WDCM 00012aWorld Data Centre for Micro-organisms (WDCM)Bacillus subtilis subsp. spizizenii WDCM 00003aStaphylococcus aureus WDCM 00032 or WDCM 00034Reference medium Tryptone soya agarControl method QuantitativeCriterio
44、n Productivity ratio (PR) 0,7aThe strains to be used as a minimum by the user laboratory. See Reference 7 for information on culture collection strain numbers and contact details.5.4 Overlay medium (if necessary; see 9.2.7)5.4.1 CompositionAgara12 g to 18 gWater 1 000 mlaDepending on the gel strengt
45、h of the agar.5.4.2 PreparationAdd the agar to the water and heat to boiling, stirring frequently until the agar is completely dissolved, or steam for about 30 min.Adjust the pH (6.4), if necessary, so that after sterilization it is 7,0 0,2 at 25 C.Dispense the medium into tubes, flasks or bottles (
46、6.8) of appropriate capacity.Sterilize in an autoclave at 121 C for 15 min. ISO 2013 All rights reserved 3BS EN ISO 4833-1:2013ISO 4833-1:2013(E)lf the medium is to be used immediately, cool it to 44 C to 47 C in a water bath (6.3) before use. If not, store it in the dark at a temperature of (5 3) C
47、 for no longer than 3 months, under conditions which do not allow any change in its composition and properties.Before beginning the microbiological examination, completely melt the medium then cool it to 44 C to 47 C in a water bath (6.3) before use. See ISO 11133.6 ApparatusDisposable apparatus is
48、an acceptable alternative to re-usable glassware and plastic if it has suitable specifications.Usual microbiological laboratory equipment (see ISO 7218) and in particular the following.6.1 Oven for dry sterilization or autoclave for wet sterilization, used in accordance with ISO 7218.6.2 Incubator,
49、capable of being maintained at (30 1) C.6.3 Water bath, capable of being maintained at 44 C to 47 C.6.4 pH-meter, accurate to within 0,1 pH unit at 25 C.6.5 Petri dishes, made of glass or plastic, of diameter 90 mm to 100 mm.6.6 Total delivery graduated pipettes, of nominal capacity 1 ml, graduated in 0,1 ml divisions, ISO 8351class A, or automatic pipettes, ISO 8655-2,2with use of sterile tips.6.7 Colony-counting equipment (optional), consisting of an illuminated base and, optionally, a mech
