DIN EN 12353-2013 en 9586 Chemical disinfectants and antiseptics - Preservation of test organisms used for the determination of bactericidal (including Legionella) mycobactericidal.pdf

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1、April 2013 Translation by DIN-Sprachendienst.English price group 15No 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

2、11.080.20; 71.100.35!$,“1990904www.din.deDDIN EN 12353Chemical disinfectants and antiseptics Preservation of test organisms used for the determination ofbactericidal (including Legionella), mycobactericidal, sporicidal,fungicidal and virucidal (including bacteriophages) activity;English version EN 1

3、2353:2013,English translation of DIN EN 12353:2013-04Chemische Desinfektionsmittel und Antiseptika Aufbewahrung von Testorganismen fr die Prfung der bakteriziden (einschlielichLegionella), mykobakteriziden, sporiziden, fungiziden und viruziden (einschlielichBakteriophagen) Wirkung;Englische Fassung

4、EN 12353:2013,Englische bersetzung von DIN EN 12353:2013-04Antiseptiques et dsinfectants chimiques Conservation des organismes test utiliss pour la dtermination de lactivit bactricide(Legionella inclus), mycobactricide, sporicide, fongicide et virucide (bacteriophagesinclus);Version anglaise EN 1235

5、3:2013,Traduction anglaise de DIN EN 12353:2013-04SupersedesDIN EN 12353:2007-02www.beuth.deDocument comprises 35 pagesIn case of doubt, the German-language original shall be considered authoritative.04.13 DIN EN 12353:2013-04 2 A comma is used as the decimal marker. National foreword This document

6、(EN 12353:2013) has been prepared by Technical Committee CEN/TC 216 “Chemical disinfectants and antiseptics” (Secretariat: AFNOR, France). The responsible German body involved in its preparation was the Normenausschuss Medizin (Medical Standards Committee), Working Committee NA 063-04-07 AA Chemisch

7、e Desinfektionsmittel und Antiseptika in der Humanmedizin. Amendments This standard differs from DIN EN 12353:2007-02 as follows: a) the methods of preservation of Legionella, mycobacteria, bacteriophages and viruses are new and have been added. b) the standard has been brought in line with other CE

8、N/TC 216 standards dealing with quantitative suspension tests. Data obtained by using the former version of EN 12353 are still valid. Previous editions DIN EN 12353: 2007-02 EUROPEAN STANDARD NORME EUROPENNE EUROPISCHE NORM EN 12353 February 2013 ICS 11.080.20; 71.100.35 Supersedes EN 12353:2006Engl

9、ish Version Chemical disinfectants and antiseptics - Preservation of test organisms used for the determination of bactericidal (including Legionella), mycobactericidal, sporicidal, fungicidal and virucidal (including bacteriophages) activity Antiseptiques et dsinfectants chimiques - Conservation des

10、 organismes test utiliss pour la dtermination de lactivit bactricide (Legionella inclus), mycobactricide, sporicide, fongicide et virucide (bacteriophages inclus) Chemische Desinfektionsmittel und Antiseptika - Aufbewahrung von Testorganismen fr die Prfung der bakteriziden (einschlielich Legionella)

11、, mykobakteriziden, sporiziden, fungiziden und viruziden (einschlielich Bakteriophagen) Wirkung This European Standard was approved by CEN on 14 December 2012. CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European Standard t

12、he 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 exists in three official versions (English, Fre

13、nch, 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 standards bodies of Austria, Belgium, Bulgaria

14、, 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, Switzerland, Turkey

15、 and United Kingdom. EUROPEAN COMMITTEE FOR STANDARDIZATION COMIT EUROPEN DE NORMALISATION EUROPISCHES KOMITEE FR NORMUNG 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 123

16、53:2013: EEN 12353:2013 (E) 2 Contents Page Foreword 3 Introduction .4 1 Scope 5 2 Normative references 5 3 Terms and definitions .5 4 Requirements .5 5 Methods 5 5.1 Principle 5 5.2 Materials and reagents 6 5.3 Apparatus and glassware 15 5.4 Procedure for preservation of test organisms General . 16

17、 5.5 Procedure for preservation of bacteria (incl. Legionella, spore-forming bacteria, excl. mycobacteria and bacterial spores) and yeasts 17 5.6 Procedure for preservation of mycobacteria . 18 5.7 Procedure for preservation of moulds (e.g. Aspergillus brasiliensis) 19 5.8 Procedure for preservation

18、 of viruses (except dairy bacteriophages) . 20 5.9 Procedure for preservation of bacteriophages . 20 5.10 Verification of the purity and identity of test organisms 21 5.11 Documentation 21 Annex A (informative) Test organisms Culture collection references and relation to CEN/TC 216 standards and prE

19、Ns 23 A.1 Bacteria (except mycobacteria and spore-forming bacteria) . 23 A.2 Mycobacteria . 24 A.3 Spore-forming bacteria 24 A.4 Fungi (moulds and yeasts) 25 A.5 Viruses . 25 A.6 Bacteriophages . 26 Annex B (informative) Graphical representations 27 Bibliography . 32 DIN EN 12353:2013-04EN 12353:201

20、3 (E) 3 Foreword This document (EN 12353:2013) has been prepared by Technical Committee CEN/TC 216 “Chemical disinfectants and antiseptics”, 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

21、or by endorsement, at the latest by August 2013, and conflicting national standards shall be withdrawn at the latest by August 2013. 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

22、for identifying any or all such patent rights. This document supersedes EN 12353:2006. The document was revised to adapt it to the latest state of science, to correct errors and ambiguities. The following are the significant technical changes since the last edition: The methods of preservation of Le

23、gionella, mycobacteria, bacteriophages and viruses are new and were added. Data obtained by using the former version of EN 12353 are still valid. According to the CEN/CENELEC Internal Regulations, the national standards organisations of the following countries are bound to implement this European St

24、andard: 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, Slovenia,

25、Spain, Sweden, Switzerland, Turkey and the United Kingdom. DIN EN 12353:2013-04 EN 12353:2013 (E) 4 Introduction Standardized tests for the determination of bactericidal (incl. Legionella pneumophila), mycobactericidal, sporicidal, fungicidal and virucidal (incl. bacteriophages) activity of chemical

26、 disinfectants and antiseptics necessitate the use of test organisms whose purity and identity have been verified and whose biological behaviour remains stable. Therefore it is essential to specify the storage requirements. This European Standard aims at describing methods for preservation of test o

27、rganisms used for such purposes. DIN EN 12353:2013-04 EN 12353:2013 (E) 5 1 Scope This European Standard specifies methods for keeping test organisms used and defined in European Standards for the determination of bactericidal (incl. Legionella pneumophila), mycobactericidal, sporicidal, fungicidal

28、and virucidal (incl. bacteriophages) activity of chemical disinfectants and antiseptics drawn up by CEN/TC 216. These methods for keeping test organisms can only be carried out in connection with at least one of those standards where a reference to this European Standard is established. NOTE 1 Annex

29、 A (informative) contains a non-exhaustive list of test organisms for which this standard can be applied. NOTE 2 European Standards (EN and prEN) where this European Standard is referenced are listed in the Bibliography. NOTE 3 A specific part on the preservation of bacterial spores may be added onc

30、e the results of the ongoing ring trials are available. 2 Normative references The following documents, in whole or in part, are normatively referenced in this document and are indispensable for its application. For dated references, only the edition cited applies. For undated references, the latest

31、 edition of the referenced document (including any amendments) applies. EN 14885, Chemical disinfectants and antiseptics Application of European Standards for chemical disinfectants and antiseptics 3 Terms and definitions For the purposes of this document, the terms and definitions given in EN 14885

32、 apply. 4 Requirements Each test organism specified in a CEN/TC 216 European Standard and referred to in this European Standard shall be handled as described in this European Standard. The purity and identity of the preserved test organism shall be verified during the preparation and regularly durin

33、g the storage, except for viruses where only the identity is checked before the stock virus suspension is stored. The preserved test organism except viruses - should be checked at regular intervals (at least in the case of longer storage than 14 months) to ensure that its susceptibility to products

34、has not changed. As long as CEN/TC 216 has not developed specific tests for this purpose any suitable method can be used e.g. EN 1040 for bacteria, EN 1275 for fungi, EN 14348 for mycobacteria, EN 13623 for Legionella pneumophila, EN 14476 for viruses or EN 13610 for dairy bacteriophages. 5 Methods

35、5.1 Principle A sample of the test organism in general in freeze dried form - is obtained from a culture collection. This sample is cultured, prepared for storage, filled into storage vessels and placed in the deep freeze. DIN EN 12353:2013-04 EN 12353:2013 (E) 6 From this sample a stock culture is

36、prepared and subsequently used to prepare working cultures for the test procedure. In some cases the working cultures are directly prepared from the deep freeze samples. 5.2 Materials and reagents 5.2.1 Test organisms See Annex A for examples of test organisms. The origin (culture collection), taxon

37、omic name and reference number, date of receipt and batch number of the freeze dried test organisms shall be recorded (5.11.2). 5.2.2 Culture media and reagents 5.2.2.1 General The formulas of all media and reagents are given in case commercial ready-to-use material is not used. It is to be checked

38、that each commercial supplier has established an appropriate quality control system. All weights of chemical substances given in this European Standard refer to the anhydrous salts unless otherwise stated. Hydrated forms may be used as an alternative, but the weights required shall be adjusted to al

39、low for consequent molecular weight differences. The reagents shall be of analytical grade and/or appropriate for microbiological purposes. They shall be free from substances that are toxic or inhibitory to the test organisms. To improve reproducibility, it is recommended that whenever possible, com

40、mercially available dehydrated material is used for the preparation of culture media. The manufacturers instructions relating to the preparation of these products should be rigorously followed. All specified pH values are measured at (20 1) C. For each culture medium, cell culture and reagent a limi

41、tation for use should be fixed. 5.2.2.2 Water The water shall be freshly glass distilled water and not demineralized water. If distilled water of adequate quality is not available, water for injections (see bibliographic reference 1) can be used. Sterilize in the autoclave (5.3.2.1a). Sterilization

42、is not necessary if the water is used for e.g. preparation of culture media and subsequently sterilized. 5.2.2.3 Tryptone Soya Broth (TSB) for bacteria, except Legionella Tryptone soya broth, consisting of: Tryptone, pancreatic digest of casein 17,0 g Soya peptone, papaic digest of soybean meal 3,0

43、g Sodium chloride (NaCl) 5,0 g Water (5.2.2.2) 800,0 ml Dipotassium phosphate (K2HPO4) 2,5 g Glucose 2,5 g Water (5.2.2.2) to 1 000,0 ml Sterilize in the autoclave (5.3.2.1a). After sterilization the pH of the medium shall be equivalent to 7,2 0,2. DIN EN 12353:2013-04 EN 12353:2013 (E) 7 5.2.2.4 Ma

44、lt Extract Broth (MEB) for fungi Malt extract broth, consisting of: Malt extract (food grade, e.g. Christomalt powder from Difal or equivalent that is not highly purified and not only based on maltose, e.g. malt extract from OXOID)120,0 g Water (5.2.2.2) to 1 000,0 ml Sterilize in the autoclave (5.3

45、.2.1a). After sterilization the pH of the medium shall be equivalent to 5,6 0,2. 5.2.2.5 Cryoprotectant solution for bacteria, spore-forming bacteria, fungi Cryoprotectant solution, consisting of: Beef extract 3,0 g Tryptone, pancreatic digest of casein 5,0 g Glycerol (C3H8O3) 2 150,0 g Water (5.2.2

46、.2) to 1 000,0 ml Dissolve the constituents in boiling water. Sterilize in the autoclave (5.3.2.1a). After sterilization the pH of the solution shall be equivalent to 6,9 0,2. Any commercially available cryoprotectant containing glycerol for preservation of test organisms equivalent to the solution

47、described above may be used. If justified, any other equivalent cryoprotectant solution may be used, e.g. for Legionella (5.5.2). 5.2.2.6 Middlebrook 7 H 9 broth with 10 % ADC enrichment and glycerol as reconstituent and cryoprotectant solution for mycobacteria (MADC) Middlebrook 7 H 9 broth, consis

48、ting of: Middlebrook 7 H 9 broth powder 4,7 g Glycerol (C3H8O3) 2 100,0 ml Water (5.2.2.2) 800,0 ml Treat in the autoclave (5.3.2.1a) for a holding time of only 10 min and cool to 45 C. Add under aseptic conditions 100 ml Middlebrook ADC enrichment to obtain approximately 1 000,0 ml. The pH of the medium shall be equivalent to 6,6 0,2. 5.2.2.7 Polysorbate 80 solution Polysorbate 80 solution, consisting of: Polysorbate 80 0,5 g Water (5.2.2.2) to 1 000,0 ml Steriliz