SIST EN 13697:2015
(Main)Chemical disinfectants and antiseptics - Quantitative non-porous surface test for the evaluation of bactericidal and/or fungicidal activity of chemical disinfectants used in food, industrial, domestic and institutional areas - Test method and requirements without mechanical action (phase 2, step 2)
Chemical disinfectants and antiseptics - Quantitative non-porous surface test for the evaluation of bactericidal and/or fungicidal activity of chemical disinfectants used in food, industrial, domestic and institutional areas - Test method and requirements without mechanical action (phase 2, step 2)
This European Standard specifies a test method (phase 2/step 2) and the minimum requirements for bactericidal and/or fungicidal activity of chemical disinfectants that form a homogeneous physically stable preparation in hard water and that are used in food, industrial, domestic and institutional areas, excluding areas and situations where disinfection is medically indicated and excluding products used on living tissues.
The scope of this European Standard applies at least to the following :
a) Processing, distribution and retailing of :
1) Food of animal origin :
milk and milk products ;
meat and meat products ;
fish, seafood and products ;
eggs and egg products ;
animal feeds ;
etc.
2) Food of vegetable origin :
beverages ;
fruits, vegetables and derivatives (including sugar distillery) ;
flour, milling and backing ;
animal feeds ;
etc.
b) Institutional and domestic areas :
catering establishments ;
public areas ;
public transports ;
schools ;
nurseries ;
shops ;
sports rooms ;
waste container (bins) ;
hotels ;
dwellings ;
clinically non sensitive areas of hospitals ;
offices ;
etc.
c) Other industrial areas :
packaging material ;
biotechnology (yeast, proteins, enzymes...) ;
pharmaceutical ;
cosmetics and toiletries ;
textiles ;
space industry, computer industry ;
etc.
Using this European Standard, it is possible to determine the bactericidal or fungicidal activity of the undiluted product. As three concentrations have to be tested, in the active to non active range, dilution of the product is required and, therefore, the product shall form a homogeneous stable preparation in hard water.
NOTE 1 The method described is intended to determine the activity of commercial formulations or active substances on bacteria and/or fungi in the conditions in which they are used.
NOTE 2 This method corresponds to a phase 2/step 2 test (see annex E).
Chemische Desinfektionsmittel und Antiseptika - Quantitativer Oberflächen-Versuch nicht poröser Oberflächen zur Bestimmung der bakteriziden und/oder fungiziden Wirkung chemischer Desinfektionsmittel in den Bereichen Lebensmittel, Industrie, Haushalt und öffentliche Einrichtungen - Prüfverfahren und Anforderungen ohne mechanische Behandlung (Phase 2, Stufe 2)
Diese Europäische Norm legt ein Prüfverfahren (Phase 2/Stufe 2) und die Mindestanforderungen an die bakterizide und/oder fungizide oder levurozide Wirkung von chemischen Desinfektionsmitteln in den Bereichen Lebensmittel, Industrie, Haushalt und in öffentlichen Einrichtungen fest, die mit Wasser standardisierter Härte oder, im Fall von gebrauchsfertigen Produkten mit Wasser eine homogene, physikalisch stabile Lösung ergeben. Ausgenommen davon sind Bereiche und Situationen mit medizinisch indizierter Desinfektion und Produkte, die auf lebenden Geweben benutzt werden.
Diese Europäische Norm gilt grundsätzlich für den folgenden Anwendungsbereich:
a) Verarbeitung, Vertrieb und Verkauf von:
1) Lebensmitteln tierischer Herkunft:
- Milch und Milchprodukte;
- Fleisch und Fleischprodukte;
- Fisch, Meerestiere und daraus hergestellte Zubereitungen;
- Eier und Eiprodukte;
- Tiernahrung;
- usw.
2) Lebensmitteln pflanzlicher Herkunft:
- Getränke;
- Früchte, Gemüse und daraus hergestellte Zubereitungen (einschließlich Zucker, Destillations-produkte);
- Mehl, gemahlene und gebackene Produkte;
- Tiernahrung;
- usw.
b) öffentliche Einrichtungen und Haushaltsbereiche:
- Verpflegungseinrichtungen;
- öffentliche Bereiche;
- öffentliche Verkehrsmittel;
- Schulen;
- Kindergärten;
- Geschäfte;
- Sportstätten;
- Abfallbehälter (Mülltonnen);
- Hotels;
- Wohngebäude;
- klinisch nicht relevante Bereiche von Krankenhäusern;
- Büroräume;
- usw.
c) andere industrielle Bereiche:
- Verpackungsmaterial;
- Biotechnologie (Hefe, Proteine, Enzyme usw.);
- Hersteller von Pharmazeutika;
- Hersteller von Kosmetika und Toilettenartikeln;
- Hersteller von Textilien;
- Weltraumforschungsindustrie und Computerindustrie;
- usw.
Bei Anwendung dieser Europäischen Norm ist die Bestimmung der bakteriziden oder fungiziden oder levuroziden Wirkung des unverdünnten Produkts nicht möglich. Da vom aktiven bis zum nicht aktiven Bereich drei Konzentrationen zu prüfen sind, ist eine Verdünnung des Produkts erforderlich, deshalb muss das Produkt eine homogene stabile Lösung mit hartem Wasser bilden.
In EN 14885 ist die Beziehung zwischen den einzelnen Prüfungen untereinander und zu den Anwen-dungsempfehlungen detailliert festgelegt.
ANMERKUNG 1 Das beschriebene Verfahren bestimmt die Wirksamkeit gewerblich hergestellter Zubereitungen oder anderer aktiver Substanzen auf Bakterien und/oder Pilze unter Anwendungsbedingungen.
ANMERKUNG 2 Dieses Verfahren kann nicht zur Bestimmung der Wirkung der Produkte gegenüber Mykobakterien angewendet werden.
Antiseptiques et désinfectants chimiques - Essai quantitatif de surface non-poreuse pour l'évaluation de l'activité bactéricide et/ou fongicide des désinfectants chimiques utilisés dans le domaine de l'agro-alimentaire, dans l'industrie, dans les domaines domestiques et en collectivité - Méthode d'essai sans action mécanique et prescriptions (phase 2/étape 2)
La présente Norme européenne décrit une méthode d’essai (phase 2, étape 2) et les prescriptions minimales relatives à l’activité bactéricide et/ou fongicide ou levuricide des désinfectants chimiques qui forment une préparation homogène, physiquement stable dans l’eau dure ou - dans le cas des produits prêts à l'emploi, avec de l'eau - et qui sont utilisés dans le domaine de l’agro-alimentaire, dans l’industrie, dans le domaine domestique et en collectivités, à l’exception des domaines et des situations où la désinfection est médicalement prescrite et à l’exclusion des produits utilisés sur les tissus vivants.
Le domaine d’application de la présente Norme européenne couvre au moins les secteurs suivants :
a) fabrication, distribution et vente au détail de :
1) produits alimentaires d’origine animale :
- lait et produits laitiers ;
- viande et produits dérivés de la viande ;
- poisson, fruits de mer et leurs produits dérivés ;
- oeufs et produits dérivés ;
- alimentation animale ;
- etc.
2) produits alimentaires d’origine végétale :
- boissons ;
- fruits, légumes et leurs dérivés (y compris le sucre, les produits de distillation) ;
- farine, minoterie et boulangerie ;
- produits alimentaires pour animaux ;
- etc.
b) domaines domestiques et collectivité :
- établissements de restauration ;
- lieux publics ;
- transports publics ;
- écoles ;
- crèches ;
- magasins ;
- salles de sport ;
- conteneurs pour déchets (poubelles) ;
- hôtels ;
- locaux d’habitation ;
- zones cliniquement non sensibles des hôpitaux ;
- bureaux ;
- etc.
c) autres secteurs industriels :
- matériaux d’emballage ;
- biotechnologie (levure, protéines, enzymes.) ;
- industrie pharmaceutique ;
- cosmétiques et produits d’hygiène corporelle ;
- textiles ;
- industrie spatiale, secteur informatique ;
- etc.
Il est possible d’utiliser la présente Norme européenne pour déterminer l’activité bactéricide ou fongicide ou levuricide des produits prêts à l’emploi. Trois concentrations, dont une active et l’autre non active, devant être soumises à essai, une dilution du produit est nécessaire et ce dernier doit donc former une préparation homogène et stable dans l’eau dure, pour un produit à diluer et en eau distillée pour un produit prêt à l’emploi.
L'EN 14885 spécifie en détail la relation entre les différents essais et les « recommandations d'emploi.
NOTE 1 La méthode décrite est destinée à déterminer l’efficacité des formulations commerciales ou des substances actives sur les bactéries et/ou les champignons dans les conditions dans lesquelles elles doivent être utilisées.
NOTE 2 Cette méthode ne peut pas être utilisée pour évaluer l'activité des produits vis-à-vis des mycobactéries.
Kemična razkužila in antiseptiki - Kvantitativni preskus na neporoznih površinah za vrednotenje baktericidnega in/ali fungicidnega delovanja kemičnih razkužil v živilski in drugih industrijah, gospodinjstvu in javnih ustanovah - Preskusna metoda in zahteve brez mehanskega delovanja (faza 2, stopnja 2)
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2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.Chemische Desinfektionsmittel und Antiseptika - Quantitativer Oberflächen-Versuch nicht poröser Oberflächen zur Bestimmung der bakteriziden und/oder fungiziden Wirkung chemischer Desinfektionsmittel in den Bereichen Lebensmittel, Industrie, Haushalt und öffentliche Einrichtungen - Prüfverfahren und Anforderungen ohne mechanische Behandlung (Phase 2, Stufe 2)Antiseptiques et désinfectants chimiques - Essai quantitatif de surface non-poreuse pour l'évaluation de l'activité bactéricide et/ou fongicide des désinfectants chimiques utilisés dans le domaine de l'agro-alimentaire, dans l'industrie, dans les domaines domestiques et en collectivité - Méthode d'essai sans action mécanique et prescriptions (phase 2/étape 2)Chemical disinfectants and antiseptics - Quantitative non-porous surface test for the evaluation of bactericidal and/or fungicidal activity of chemical disinfectants used in food, industrial, domestic and institutional areas - Test method and requirements without mechanical action (phase 2, step 2)71.100.35Kemikalije za dezinfekcijo v industriji in domaChemicals for industrial and domestic disinfection purposesICS:Ta slovenski standard je istoveten z:EN 13697:2015SIST EN 13697:2015en,fr,de01-junij-2015SIST EN 13697:2015SLOVENSKI
STANDARDSIST EN 13697:20021DGRPHãþD
SIST EN 13697:2015
EUROPEAN STANDARD NORME EUROPÉENNE EUROPÄISCHE NORM
EN 13697
April 2015 ICS 11.080.20; 71.100.35 Supersedes EN 13697:2001English Version
Chemical disinfectants and antiseptics - Quantitative non-porous surface test for the evaluation of bactericidal and/or fungicidal activity of chemical disinfectants used in food, industrial, domestic and institutional areas - Test method and requirements without mechanical action (phase 2, step 2)
Antiseptiques et désinfectants chimiques - Essai quantitatif de surface non-poreuse pour l'évaluation de l'activité bactéricide et/ou fongicide des désinfectants chimiques utilisés dans le domaine de l'agro-alimentaire, dans l'industrie, dans les domaines domestiques et en collectivité - Méthode d'essai sans action mécanique et prescriptions (phase 2/étape 2)
Chemische Desinfektionsmittel und Antiseptika - Quantitativer Oberflächen-Versuch nicht poröser Oberflächen zur Bestimmung der bakteriziden und/oder fungiziden Wirkung chemischer Desinfektionsmittel in den Bereichen Lebensmittel, Industrie, Haushalt und öffentliche Einrichtungen - Prüfverfahren und Anforderungen ohne mechanische Behandlung (Phase 2, Stufe 2) This European Standard was approved by CEN on 20 January 2015.
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 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 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, 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 and United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION EUROPÄISCHES KOMITEE FÜR NORMUNG
CEN-CENELEC Management Centre:
Avenue Marnix 17,
B-1000 Brussels © 2015 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN 13697:2015 ESIST EN 13697:2015
EN 13697:2015 (E) 2 Contents Page Foreword .3 Introduction .4 1 Scope .5 2 Normative references .6 3 Terms and definitions .6 4 Requirements .7 5 Test methods .8 5.1 Principle .8 5.2 Materials and reagents .8 5.2.1 Test organisms .8 5.2.2 Culture media and reagents .9 5.2.3 Test surface . 12 5.3 Apparatus and glassware . 13 5.4 Preparation of test organism suspensions and product test solutions . 14 5.4.1 Test organism suspensions . 14 5.4.2 Product test solutions . 17 5.5 Procedure . 17 5.5.1 Choice of experimental conditions . 17 5.5.2 Test procedure . 18 5.5.3 Counting of the test mixtures . 20 5.6 Calculation and expression of results . 21 5.6.1 Elaboration of data: counting of weighed average values . 21 5.6.2 Verification of methodology . 22 5.6.3 Expression of results . 23 5.6.4 Conclusion . 23 5.7 Test report . 24 Annex A (informative)
Corresponding referenced strains . 26 Annex B (informative)
Neutralizers . 28 Annex C (informative)
Expression of results with the dilution-neutralization method . 30 Annex D (informative)
Bactericidal activity in surfaces in general use conditions (for clean conditions) . 32 Annex E (informative)
Precision of the test result . 33 Bibliography . 34
SIST EN 13697:2015
EN 13697:2015 (E) 3 Foreword This document (EN 13697:2015) 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 or by endorsement, at the latest by October 2015 and conflicting national standards shall be withdrawn at the latest by October 2015. 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 for identifying any or all such patent rights. This document supersedes EN 13697:2001. The changes between this edition and EN 13697:2001 are the following: — interfering substance has been changed from 0,03 % bovine albumin to 0,85 % skimmed milk (see Clause 4, Table 1) for Pseudomonas aeruginosa under clean conditions only; — A. brasiliensis (ex A. niger) spore preparation has been updated in order to harmonize this step with the QST fungicidal test method amendment issued in 2012 (see 5.4.1.3 b)); — Calculations of the weighed means and of the results have been modified in order to be harmonized with new CEN TC 216 standards (see 5.4.1.5, 5.5.2, 5.5.3 and 5.6); — Other paragraphs have been harmonized to new CEN TC 216 standards (e.g. preparation of hard water, 5.2.2.7). Results obtained from the previous standard for Aspergillus niger need to be repeated to take into account the new spore morphology requirement and the change in interfering substance. According to the CEN-CENELEC Internal Regulations, the national standards organizations of the following countries are bound to implement this European Standard: 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, Spain, Sweden, Switzerland, Turkey and the United Kingdom. SIST EN 13697:2015
EN 13697:2015 (E) 4 Introduction This European Standard describes a surface test method for establishing whether a product proposed as a disinfectant in the fields described in Clause 1 has or does not have bactericidal and/or fungicidal or yeasticidal activity on non-porous surfaces. This European Standard has been revised in order to modify the interfering substance under “clean conditions” adopted for P. aeruginosa; in order to modify the calculation of N, NC, NT, Nc, Na and consequently the final results and to harmonize the standard with the other recent CEN TC 216 standards. The laboratory test closely simulates practical conditions of application. Chosen conditions (contact time, temperature, organisms on surfaces .) reflect parameters which are found in practical situations including conditions which may influence the action of disinfectants. Each use concentration found from this test corresponds to defined experimental conditions. The conditions are intended to cover general purposes and to allow reference between laboratories and product types. However, for some applications the recommendations of use of a product can differ and therefore additional test conditions need to be used. SIST EN 13697:2015
EN 13697:2015 (E) 5 1 Scope This European Standard specifies a test method (phase 2/step 2) and the minimum requirements for bactericidal and/or fungicidal or yeasticidal activity of chemical disinfectants that form a homogeneous physically stable preparation in hard water or – in the case of ready-to-use products – with water in food, industrial, domestic and institutional areas, excluding areas and situations where disinfection is medically indicated and excluding products used on living tissues. The scope of this European Standard applies at least to the following: a) Processing, distribution and retailing of: 1) Food of animal origin: i) milk and milk products; ii) meat and meat products; iii) fish, seafood and products; iv) eggs and egg products; v) animal feeds; vi) etc. 2) Food of vegetable origin: i) beverages; ii) fruits, vegetables and derivatives (including sugar distillery); iii) flour, milling and backing; iv) animal feeds; v) etc. b) Institutional and domestic areas: 1) catering establishments; 2) public areas; 3) public transports; 4) schools; 5) nurseries; 6) shops; 7) sports rooms; 8) waste container (bins); SIST EN 13697:2015
EN 13697:2015 (E) 6 9) hotels; 10) dwellings; 11) clinically non sensitive areas of hospitals; 12) offices; 13) etc. c) Other industrial areas: 1) packaging material; 2) biotechnology (yeast, proteins, enzymes.); 3) pharmaceutical; 4) cosmetics and toiletries; 5) textiles; 6) space industry, computer industry; 7) etc. Using this European Standard, it is possible to determine the bactericidal or fungicidal or yeasticidal activity of the undiluted product. As three concentrations are tested, in the active to non active range, dilution of the product is required and, therefore, the product forms a homogeneous stable preparation in hard water. EN 14885 specifies in detail the relationship of the various tests to one another and to use recommendations. NOTE 1 The method described is intended to determine the activity of commercial formulations or active substances on bacteria and/or fungi in the conditions in which they are used. NOTE 2 This method cannot be used to evaluate the activity of products against mycobacteria. 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 edition of the referenced document (including any amendments) applies. EN 12353, Chemical disinfectants and antiseptics - Preservation of test organisms used for the determination of bactericidal (including Legionella), mycobactericidal, sporicidal, fungicidal and virucidal (including bacteriophages) activity EN 14885, Chemical disinfectants and antiseptics - Application of European Standards for chemical disinfectants and antiseptics ISO 4793, Laboratory sintered (fritted) filters - Porosity grading, classification and designation 3 Terms and definitions For the purposes of this document, the terms and definitions given in EN 14885 apply. SIST EN 13697:2015
EN 13697:2015 (E) 7 4 Requirements The product shall demonstrate at least a 4 decimal log (lg) reduction for bacteria and at least a 3 decimal log (lg) reduction for fungi, when tested in accordance with Table 1 and 5.5.1. Table 1 — Obligatory and additional conditions Test Conditions Bactericidal activity on non-porous surfaces without mechanical action Yeasticidal activity on non-porous surfaces without mechanical action Fungicidal activity on non-porous surfaces without mechanical action Test organism
(see 5.2.1) obligatory Enterococcus hirae Escherichia coli Pseudomonas aeruginosa Staphylococcus aureus Candida albicans Candida albicans Aspergillus brasiliensis (ex A. niger) example Salmonella typhimurium Lactobacillus brevis Enterobacter cloacae Saccharomyces cerevisiae (for breweries) Saccharomyces cerevisiae var. diastaticus (for breweries) any relevant test organism Test temperature obligatory Between 18 °C ± 1 °C and 25 °C ± 1 °C Between 18 °C ± 1 °C and 25 °C ± 1 °C Between 18 °C ± 1 °C and 25 °C ± 1 °C additional 4 °C ± 1 °C; 10 °C ± 1 °C; 40 °C ± 1 °C 4 °C ± 1 °C; 10 °C ± 1 °C; 40 °C ± 1 °C 4 °C ± 1 °C; 10 °C ± 1 °C; 40 °C ± 1 °C Contact time obligatory 5 min ± 10 s 15 min ± 10 s 15 min ± 10 s additional 1 min ± 5 s; 15 min ± 10 s; 30 min ± 10 s; 60 min ± 10 s 1 min ± 5 s; 5 min ± 10 s; 30 min ± 10 s; 60 min ± 10 s 1 min ± 5 s; 5 min ± 10 s; 30 min ± 10 s; 60 min ± 10 s Interfering substance obligatory clean conditions 0,3 g/l Bovine Albumin for Staphylococcus aureus, Enterococcus hirae and Escherichia coli ; 8,5 g/l skim milk for Pseudomonas aeruginosa 0,3 g/l Bovine Albumin for C. albicans 0,3 g/l Bovine Albumin for C. albicans and A. brasiliensis dirty conditions 3,0 g/l Bovine Albumin for Staphylococcus aureus, Enterococcus hirae, Pseudomonas aeruginosa and Escherichia coli 3,0 g/l Bovine Albumin for C. albicans 3,0 g/l Bovine Albumin for C. albicans and A. brasiliensis additional any relevant substance any relevant substance any relevant substance Log reduction from a water control (decimal log) ≥ 4Log ≥ 3Log ≥ 3Log The obligatory contact times for surface disinfectants stated in Table 1 were chosen to enable comparison of standard conditions. The referenced test conditions are by no means intended as requirements for the use of a product, nor as requirements for the evaluation and acceptance of products by regulatory authorities. The recommended contact time for the use of the product is within the responsibility of the manufacturer. Where appropriate (specific purposes), additional specific bactericidal/yeasticidal/fungicidal activity shall be determined under other conditions of time, temperature, additional strains and interfering substances in order to take into account intended specific use conditions. NOTE For the additional conditions, the concentration defined as a result can be lower than the one obtained under the obligatory test conditions. SIST EN 13697:2015
EN 13697:2015 (E) 8 5 Test methods 5.1 Principle A test suspension of bacteria or fungi in a solution of interfering substances is inoculated onto a test stainless steel surface and dried. A prepared sample of the product under test is applied in a manner which covers the dried film. The surface is maintained at a specified temperature for a defined period of time. The surface is transferred to a previously validated neutralization medium so that the action of the disinfectant is immediately neutralized. The number of surviving organisms which can be recovered from the surface is determined quantitatively. The number of bacteria or fungi on a surface treated with hard water in place of the disinfectant is also determined and the reduction in viable counts attributed to the product is calculated by difference. 5.2 Materials and reagents 5.2.1 Test organisms The bactericidal activity shall be evaluated using the following four strains: — Pseudomonas aeruginosa ATCC 15 4421); — Staphylococcus aureus ATCC 6 538; — Enterococcus hirae ATCC 10 541; — Escherichia coli ATCC 10 536. The fungicidal or yeasticidal activity shall be evaluated using the following two strains: — Candida albicans ATCC 10 231; — Aspergillus brasiliensis (ex A. niger) ATCC 16 404. If required for specific applications, additional strains may be chosen from, for example: — Salmonella typhimurium ATCC 13 311; — Lactobacillus brevis DSM 6 235; — Enterobacter cloacae DSM 6 234; — Saccharomyces cerevisiae (for breweries) or ATCC 9 763 or DSM 1 333; — Saccharomyces cerevisiae var. diastaticus (for breweries) DSM 70 487. NOTE See Annex A for corresponding strain numbers in some other culture collections. If additional strains are used, they shall be incubated under optimum growth conditions (temperature, time, atmosphere) and noted in the test report. If the additional strains selected do not correspond to the specified strains, their suitability for supplying inocula of sufficient concentration shall be verified. If the additional strains tested are not classified at a
1) ATCC 15 442, ATCC 6 538, ATCC 10 541, ATCC 10 536, ATCC 10 231, ATCC 16 404 and ATCC 13311 are the collection numbers of strains supplied by the American Type Culture Collections. This information is given for the convenience of users of this standard and does not constitute an endorsement by CEN of the product named. Equivalent products can be used if they can be shown to lead to the same results. SIST EN 13697:2015
EN 13697:2015 (E) 9 reference centre, their identification characteristics shall be stated. In addition, they shall be held by the testing laboratory or national culture under a reference for 5 years. 5.2.2 Culture media and reagents 5.2.2.1 General The reagents shall be of analytical grade and/or appropriate for microbiological purposes. 5.2.2.2 Water The water shall be free from substances that are toxic or inhibiting to bacteria and fungi. It shall be freshly glass distilled and not demineralized water. Sterilize in the autoclave (see 5.3.2.1). NOTE 1 If the water is sterilized during sterilization of the reagents, this is not necessary. NOTE 2 If distilled water of adequate quality is not available, water for injectable preparation (see European Pharmacopoeia) can be used. 5.2.2.3 Tryptone Soya Agar (TSA) For maintenance of bacterial strains and performance of viable counts. Tryptone, pancreatic digest of casein 15,0 g Soya peptone, papaic digest of Soybean meal 5,0 g NaCl 5,0 g Agar 15,0 g Water (see 5.2.2.2) 1 000,0 ml Sterilize in the autoclave (see 5.3.2.1). After sterilization, the pH of the medium shall be equivalent to 7,2 ± 0,2 when measured at 20 ° C. 5.2.2.4 Malt extract agar (MEA) For maintenance of fungal strains, sporulation and performance of viable counts. Malt extract (food grade, e.g. Cristomalt powder from Difal) 30,0 g Agar 15,0 g Water (see 5.2.2.2) 1 000,0 ml The malt extract should be food grade (e.g. Cristomalt powder from Difal) or equivalent that is not highly purified and not only based on maltose (e.g. Malt extract from OXOID) 2). However, if there are problems producing at least 75 % spiny spores see 5.4.1.4.2.
2) This information is given for the convenience of users of this European Standard and does not constitute an endorsement by CEN of the product named. Equivalent products may be used if they can be shown to lead to the same results. SIST EN 13697:2015
EN 13697:2015 (E) 10 Sterilize in the autoclave [5.3.2.1a)]. After sterilization, the pH of the medium shall be equivalent to 5,6 ± 0,2 when measured at (20 ± 1) °C. In case of encountering problems with neutralization (5.5.2.3 and 5.5.2.4), it may be necessary to add neutralizer to the MEA. Annex B gives guidance on the neutralizers that may be used. 5.2.2.5 Diluent Tryptone sodium chloride solution: Tryptone, pancreatic digest of casein 1,0 g NaCl 8,5 g Water (see 5.2.2.2) 1 000,0 ml Sterilize in the autoclave (see 5.3.2.1). After sterilization the pH shall be equivalent to 7,0 ± 0,2 when measured at 20 °C. 5.2.2.6 Neutralizer The neutralizer shall be validated for the product under test in accordance with 5.5.2.3 and 5.5.2.4. The neutralizer shall be sterile. NOTE Information on neutralizers that have been found to be suitable for some categories of products is given in Annex B. 5.2.2.7 Hard water for dilution of the products Hard water for dilution of products shall be prepared as follows: — solution A: Dissolve 19,84 g anhydrous MgCl2 and 46,24 g anhydrous CaCl2 in water (see 5.2.2.2) and dilute to 1 000 ml. Sterilize by membrane filtration (5.3.2.19) or in the autoclave (5.3.2.1 a). Autoclaving – if used - may cause a loss of liquid. In this case, make up to 1 000 ml with water (5.2.2.2) under aseptic conditions. Store the solution in the refrigerator (5.3.2.15) for no longer than one month. — solution B: Dissolve 35,02 g NaHC03 in water (see 5.2.2.2) and dilute to 1 000 ml. 1 000 ml. Sterilize by membrane filtration (5.3.2.19). Store the solution in the refrigerator (5.3.2.15) for no longer than one week. Add at least 600 ml water (see 5.2.2.2) to 6,0 ml of solution A in a 1 000 ml volumetric flask, then add 8,0 ml solution B. Mix and dilute to 1 000 ml with water (see 5.2.2.2). Sterilize by passing through a filter with a maximum effective pore size of 0,45 µm. The pH of the hard water shall be 7,0 ± 0,2, when measured at (20 ± 1) °C (5.3.2.6). If necessary, adjust the pH by using a solution of approximately 40 g/l (about 1 mol/l) of sodium hydroxide (NaOH) or approximately 36,5 g/l (about 1 mol/l) of hydrochloric acid (HCl). The hard water shall be freshly prepared under aseptic conditions and used within 12 h. NOTE When preparing the test product solutions (5.4.2), the addition of the product to the hard water produces a different final water hardness expressed as calcium carbonate (CaCO3) in each test tube. In any case, the final hardness is lower than 375 mg/l of calcium carbonate. SIST EN 13697:2015
EN 13697:2015 (E) 11 5.2.2.8 Interfering substances 5.2.2.8.1 General The interfering substance shall be chosen according to the conditions of use laid down for the product. The interfering substance shall be sterile and prepared at 2 times its final concentration in the test. For the additional interfering substances, the ionic composition (e.g. pH, calcium and/or magnesium hardness) and chemical composition (e.g. mineral substances, protein, carbohydrates, lipids, detergents) shall be fully defined. NOTE The term “interfering substance” is used even if it contains more than one substance. The method of preparation and sterilization together with the composition shall be noted in the test report (see 5.7). 5.2.2.8.2 Bovine albumin and skimmed solutions Bovine albumin solutions for the test conditions shall be prepared as follows: — Preparation for clean conditions: — dissolve 0,06 g of bovine albumin (Cohn fraction V for Dubos medium) in 100 ml of water (see 5.2.2.2); — sterilize by 0,45 µm membrane filtration (see 5.3.2.19), keep in the refrigerator and use within one month. The final concentration of bovine albumin in the test procedure (see 5.5.2) is 0,3 g/l. Skim milk solution for the test conditions shall be prepared as follows: — Skimmed milk (clean conditions for Pseudomonas aeruginosa only), guaranteed free of antibiotics or additives, shall be reconstituted at a rate of 100 g powder per litre of water (see 5.2.2.2). The working solution shall be prepared as follows: — prepare a solution of 1,70 % (V/V) in water (see 5.2.2.2) by adding 17 parts of the 10 % stock solution (100 g skimmed milk in 1 l water) to 83 parts of water; — sterilize for 30 min at 105 °C ± 3 °C (or 5 min at 121 °C ± 3 °C). The final concentration of milk in the test procedure (see 5.5.2) is 8,5 g/l of reconstituted milk. — Preparation for dirty conditions; — dissolve 0,60 g of bovine albumin (Cohn fraction V for Dubos medium) in 100 ml of water (see 5.2.2.2); — sterilize by 0,45 µm membrane filtration (see 5.3.2.19), keep in the refrigerator and use within one month. The final concentration of bovine albumin in the test procedure (see 5.5.2) is 3,0 g/l. In addition, other interfering substances for chemical disinfectants with detergent properties (therefore simulating additional dirty conditions for specific uses) can be chosen from: SIST EN 13697:2015
EN 13697:2015 (E) 12 5.2.2.8.3 Milk (dairies) Skimmed milk, guaranteed free of antibiotics or additives, shall be reconstituted at a rate of 100 g powder per litre of water (see 5.2.2.2). The working solution shall be prepared as follows: — prepare a solution of 2,0 % (V/V) in water (see 5.2.2.2) by adding 2 parts of reconstituted milk to 98 parts of water; — sterilize for 30 min at 105 °C ± 3 °C (or 5 min at 121 °C ± 3 °C). The final concentration of milk in the test procedure (see 5.5.2) shall be 1,0 % (V/V) of reconstituted milk. 5.2.2.8.4 Yeast extract (breweries) Dehydrated yeast extract for bacteriology, shall be prepared as follows: — prepare a 20 g/l solution in water (see 5.2.2.2), adjust to pH 7,0 ± 0,2 with sodium hydroxide; — sterilize in the autoclave (see 5.3.1). The final concentration of yeast extract in the test procedure (see 5.5.2) shall be 10 g/l. 5.2.2.8.5 Sucrose (beverage, soft drink industries) Prepare a 20 g/l solution in water (see 5.2.2.2), sterilize by membrane filtration. The final concentration of sucrose in the test procedure (see 5.5.2) is 10 g/l. 5.2.2.8.6 pH 5,0 and pH 9,0 buffer solutions (clean-in-place) The buffer solution shall be described in the test report and pH values shall be recorded. The final pH in the test shall be equal to 5,0 ± 0,2 or 9,0 ± 0,2. 5.2.2.9 Sodium lauryl sulphate (cosmetics or cosmetic industries) Prepare a 10 g/l solution of sodium lauryl sulphate in water (see 5.2.2.2). Sterilize in the autoclave (see 5.3.1). The final concentration of sodium lauryl sulphate in the test procedure (see 5.5.2) is 5 g/l. 5.2.3 Test surface Stainless steel discs (2 cm diameter discs) 304 with grade 2b finish on both sides. The surfaces should be flat. The surfaces should be used only once. Prior to use the surfaces should be placed in a beaker (minimum size: 50 ml) containing not less than 20 ml of 5 % (V/V) Decon® 3) for 60 min. Immediately rinse the discs with running freshly distilled water for 10 s. The surface shall not be allowed to dry to any extent. The discs shall only be handled with forceps. Rinse the discs with water (see 5.2.2.2) for a further 10 s to ensure complete removal of the surfactant. To supply a satisfactory flow of water, a sterilized flui
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