1、BRITISH STANDARD BS EN 13610:2002 Chemical disinfectants Quantitative suspension test for the evaluation of virucidal activity against bacteriophages of chemical disinfectants used in food and industrial areas Test method and requirements (phase 2, step 1) The European Standard EN 13610:2002 has the
2、 status of a British Standard ICS 11.080.20; 67.050; 71.100.35 BS EN 13610:2002 This British Standard, having been prepared under the direction of the Health and Environment Sector Policy and Strategy Committee, was published under the authority of the Standards Policy and Strategy Committee on 10 J
3、anuary 2003 BSI 10 January 2003 ISBN 0 580 41070 6 National foreword This British Standard is the official English language version of EN 13610:2002. The UK participation in its preparation was entrusted to Technical Committee CH/216, Chemical disinfectants and antiseptics, which has the responsibil
4、ity to: A list of organizations represented on this committee can be obtained on request to its secretary. Cross-references The British Standards which implement international or European publications referred to in this document may be found in the BSI Catalogue under the section entitled “Internat
5、ional Standards Correspondence Index”, or by using the “Search” facility of the BSI Electronic Catalogue or of British Standards Online. This publication does not purport to include all the necessary provisions of a contract. Users are responsible for its correct application. Compliance with a Briti
6、sh Standard does not of itself confer immunity from legal obligations. aid enquirers to understand the text; present to the responsible international/European committee any enquiries on the interpretation, or proposals for change, and keep the UK interests informed; monitor related international and
7、 European developments and promulgate them in the UK. Summary of pages This document comprises a front cover, an inside front cover, the EN title page, pages 2 to 44, an inside back cover and a back cover. The BSI copyright date displayed in this document indicates when the document was last issued.
8、 Amendments issued since publication Amd. No. Date CommentsEUROPEANSTANDARD NORMEEUROPENNE EUROPISCHENORM EN13610 December2002 ICS11.080.20;67.050;71.100.35 Englishversion ChemicaldisinfectantsQuantitativesuspensiontestforthe evaluationofvirucidalactivityagainstbacteriophagesof chemicaldisinfectants
9、usedinfoodandindustrialareasTest methodandrequirements(phase2,step1) DsinfectantschimiquesEssaiquantitatifdesuspension pourlvaluationdelactivitvirucidecontreles bactriophagesdesdsinfectantschimiquesutilissdans ledomainedelagroalimentaireetdanslindustrie Mthodedessaietexigences(phase2,tape1) Chemisch
10、eDesinfektionsmittelQuantitativer SuspensionsversuchzurBestimmungderviruziden WirkunggegenberBakteriophagenvonchemischen DesinfektionsmittelnindenBereichenLebensmittel,und IndustriePrfverfahrenundAnforderung(Phase2,Stufe 1) ThisEuropeanStandardwasapprovedbyCENon23October2002. CENmembersareboundtocom
11、plywiththeCEN/CENELECInternalRegulationswhichstipulatetheconditionsforgivingthisEuropean Standardthestatusofanationalstandardwithoutanyalteration.Uptodatelistsandbibliographicalreferencesconcerningsuchnational standardsmaybeobtainedonapplicationtotheManagementCentreortoanyCENmember. ThisEuropeanStan
12、dardexistsinthreeofficialversions(English,French,German).Aversioninanyotherlanguagemadebytranslation undertheresponsibilityofaCENmemberintoitsownlanguageandnotifiedtotheManagementCentrehasthesamestatusastheofficial versions. CENmembersarethenationalstandardsbodiesofAustria,Belgium,CzechRepublic,Denm
13、ark,Finland,France,Germany,Greece, Iceland,Ireland,Italy,Luxembourg,Malta,Netherlands,Norway,Portugal,Spain,Sweden,SwitzerlandandUnitedKingdom. EUROPEANCOMMITTEEFORSTANDARDIZATION COMITEUROPENDENORMALISATION EUROPISCHESKOMITEEFRNORMUNG ManagementCentre:ruedeStassart,36B1050Brussels 2002CEN Allrights
14、ofexploitationinanyformandbyanymeansreserved worldwideforCENnationalMembers. Ref.No.EN13610:2002EEN13610:2002(E) 2 Contents page Foreword3 Introduction .4 1 Scope 5 2 Normativereferences 6 3 Termsanddefinitions. .6 4 Requirements .7 5 Testmethods7 5.1 Principle7 5.2 Materialandreagents . .7 5.3 Appa
15、ratusandglassware . 12 5.4 Preparationofhostbacteriasuspensions 13 5.5 Preparationofbacteriophagesuspension. .14 5.6 Producttestsolution . 16 5.7 Procedure .16 5.8 Calculationandexpressionofresults. 19 5.9 Testreport 23 AnnexA (normative) Testforvalidationofdilutionneutralizationandmolecularsievingm
16、ethods.25 AnnexB (informative) Neutralizers. .33 AnnexC (informative) Exampleofatypicaltestreport.35 AnnexD (informative) InformationontheapplicationandinterpretationofEuropeanstandardson chemicaldisinfectantsandantiseptics . 39 AnnexE (informative) ExampleofplaquesfromlysatesofphagesP001andP00841 B
17、ibliography 44EN13610:2002(E) 3 Foreword Thisdocument(EN13610:2002)hasbeenpreparedbyTechnicalCommitteeCEN/TC216,“Antisepticsand chemicaldisinfectants“thesecretariatofwhichisheldbyAFNOR. ThisEuropeanStandardshallbegiventhestatusofanationalstandard,eitherbypublicationofanidenticaltextor byendorsement,
18、atthelatestbyJune2003,andconflictingnationalstandardsshallbewithdrawnatthelatestby June2003. InthisEuropeanStandardtheAnnexAisnormativeandtheAnnexesB,C,DandEareinformative. AccordingtotheCEN/CENELECInternalRegulations,thenationalstandardsorganizationsofthefollowing countriesareboundtoimplementthisEu
19、ropeanStandard:Austria,Belgium,CzechRepublic,Denmark,Finland, France,Germany,Greece,Iceland,Ireland,Italy,Luxembourg,Malta,Netherlands,Norway,Portugal,Spain, Sweden,SwitzerlandandtheUnitedKingdom.EN13610:2002(E) 4 Introduction ThisEuropeanStandarddescribesasuspensiontestmethodintendedtoestablishwhet
20、heraproductproposedas adisinfectantinthefieldsdescribedinclause1hasordoesnothavevirucidalactivityagainstbacteriophages. NOTE Virulentbacteriophages(phages)lyticfor(i.e.virulentfor)starterculturesof Lactococcuslactissubsp. lactisused fortheproductionofcheeseandotherfermentedmilkproductsareusedasmodel
21、virusestotestthevirucidalactivityofa product. Thelaboratorytestcloselysimulatespracticalconditionsofapplication.Chosenconditions(contacttime, temperature,virusesi.e.bacteriophagesinsuspension,.)reflectparameterswhicharefoundinpractical situationsincludingconditionswhichmayinfluencetheactionofdisinfe
22、ctants. Eachutilizationconcentrationfoundfromthistestcorrespondstodefinedexperimentalconditions. Theconditionsareintendedtocovergeneralpurposesandtoallowreferencebetweenlaboratoriesandproduct types. Howeverforsomeapplicationstherecommendationsofusemaydifferandthereforeadditionaltestconditions needto
23、beused.EN13610:2002(E) 5 1Scope ThisEuropeanStandardspecifiesatestmethod(phase2,step1)andrequirementsfortheminimumvirucidal activityagainstbacteriophagesofchemicaldisinfectantsthatformahomogeneous,physicallystablepreparationin hardwaterandthatareusedinfoodandindustrialareas,excludingareasandsituatio
24、nswheredisinfectionis medicallyindicatedandexcludingproductsusedonlivingtissues. ThisEuropeanStandardappliesatleasttothefollowing: a) processing,distributionandretailingof: 1) foodofanimalorigin: milkandmilkproducts; meatandmeatproducts; fish,seafood,andtheirproducts; eggsandeggproducts; animal feed
25、s; etc.; 2) foodofvegetableorigin: beverages; fruits,vegetablesandtheirderivatives(includingsugar,distillery.); flour,millingandbaking; animal feeds; .; b)other industrial areas: biotechnology(yeast,proteins,enzymes,.). UsingthisEuropeanStandard,itisnotpossibletodeterminethevirucidalactivityagainstb
26、acteriophagesof undilutedproductassomedilutionisalwaysproducedbyaddingtheinoculumandinterferingsubstance. Forchemicaldisinfectantsthatcanbeusedwithoutdilutionitisnotpossibletodeterminewhethertheseproducts, ataconcentrationabove80%haveavirucidalactivityagainstbacteriophages. NOTE Themethoddescribedis
27、intendedtodeterminetheactivityofcommercialformulationsoractivesubstanceson viruses(bacteriophages)intheconditionsinwhichtheyareused. ThisEuropeanStandardisapplicableonlytodisinfectants,complyingwiththevalidationtest(seeannexA).EN13610:2002(E) 6 2 Normativereferences Notapplicable. 3 Termsanddefiniti
28、ons ForthepurposesofthisEuropeanStandard,thefollowingtermsanddefinitionsapply. 3.1 product(forchemicaldisinfectionand/orantisepsis) chemicalagentorformulationusedasachemicaldisinfectantorantiseptic EN1040:1997 3.2 virucideagainstbacteriophages productwhichinactivatesthebacteriophages NOTE Theadjecti
29、vederivedfrom“virucide“is“virucidal“. 3.3 virucidalactivityagainstbacteriophages capabilityofaproducttoreducetheinfectivityofintactbacteriophagesbelongingtoreferencebacteriophage strainsP001andP008foratleast4lgundertheconditionsdefinedbythisstandard 3.4 infectivityofbacteriophages theabilityofabacte
30、riophagetopropagateinasuitablehostbacterialcellresultinginthereleaseofbacteriophage progeny 3.5 inactivationofbacteriophage thereductionofinfectivityofabacteriophagebyaproductspecifiedasachemicaldisinfectant 3.6 referencebacteriophagesuspension bacteriophagesuspensionofadefinedvirusstrainmaintainedi
31、nreferencecentersandwhichshouldnotbe passagedmorethan10times 3.7 stockbacteriophagesuspension bacteriophagesuspensionofadefinedstrainthathasbeenmultipliedonalargescaletoobtainabacteriophage suspensionrevealingidenticalcharacteristicsasthereferencebacteriophagesuspension NOTE Thisstockbacteriophagesu
32、spensionisusedtoprepareahightitertestbacteriophagesuspensionforthe bacteriophageinactivationtest. 3.8 hightiterbacteriophagesuspension thehightiterbacteriophagessuspensionobtainedfromagarplatesrevealingconfluentlysisinthebacteriallawn thatisusedtopreparethebacteriophagetestsuspensioninthevirucidalte
33、stingofthedisinfectant 3.9 bacteriophagetestsuspension thebacteriophagesuspensionofadefinedtiterthatisusedinthevirucidaltestingofthedisinfectantEN13610:2002(E) 7 4Requirements Theproductdilutedinhardwaterwhentestedinaccordancewithclause5shalldemonstrateatleasta4log 10 reductionofinfectivityofbacteri
34、ophageswhentestedinthepresenceofavolumefractionof1%acidicwhey (preparedfromacidifiedlowfatmilk)oroptionallyinthepresenceofavolumefractionof1%skimmilkasthe interferingsubstanceaccordingtoitspracticalapplicationsandundertherequiredtestconditions(20C,15min,2 referencebacteriophagestrains). Thevirucidal
35、activityagainstbacteriophagesshallbeevaluatedusingthetwovirulentbacteriophages Lactococcus lactissubsp. lactisbacteriophageP001and Lactococcuslactis subsp. lactis bacteriophageP008. Bothphagesshallbepropagatedonthehoststrain Lactococcuslactis subsp. lactisF7/2. Thedeterminedvirucidalconcentrationoft
36、hetestedproductissuggestedasbeingsuitableforpracticalsituations ofuse. Whereappropriate,additionalspecificvirucidalactivityagainstbacteriophagesshallbedeterminedunderother conditionsoftime,temperature,additionalstrainsandinterferingsubstancesinaccordancewith(see5.7.1)in ordertotakeintoaccountintende
37、dspecificuseconditions. NOTE Fortheseadditionalconditions,theconcentrationdefinedasaresultcanbelowerthantheoneobtainedunderthe initialtestconditionsof20C,15min,2selectedbacteriophagereferencestrains. 5Test methods 5.1Principle 5.1.1 Atestsuspensionofbacteriophagesinasolutionofinterferingsubstancesis
38、addedtoapreparedsample oftheproductundertestdilutedinhardwater. Themixtureismaintainedat20C1Cfor15min10s(requiredobligatorytestconditions). Attheendofthecontacttime,aliquotsaretakenandthevirucidalactivityagainstbacteriophagesinthisportionis immediatelyneutralizedorsuppressedbyavalidatedmethod.Themet
39、hodofchoiceisdilutionneutralizationwitha validatedneutralizer.Ifasuitableneutralizerisnotavailableforaspecificproduct,removaloftheproductby molecularsieving(i.e.gelfiltration)shallbeused. Thenumberofsurvivingbacteriophageparticlesandthenumberofbacteriophageparticlesinthetestsuspension aredeterminedf
40、romappropriatedilutionserieswithafactorof10preparedinmediumintesttubes. 5.1.2 Additionalandoptionalexposuretimes,temperaturesandinterferingsubstancesarespecified(see5.7.1). NOTE1 Thetestdescribedisbasedonanassessment(underspecificconditions)whichgivesareductionofatleast 99,99%(4lg)oftheinfectivityof
41、thedifferentphagesafterdifferentcontacttimes. NOTE2 Forprincipalreasons,theresultofaninactivationappliedtoaviralpopulationisnotnecessarilyequalto100%:i.e ., onecannotconcludethatthereisa100%inactivationwhenonconductingtheexperimentnoinfectiousphagearefoundwithin alimitnumberofsampling. 5.2Materialan
42、dreagents 5.2.1 Testorganisms Thevirucidalactivityagainstbacteriophagesshallbeevaluatedusingthetwofollowingbacteriophagestrains:EN13610:2002(E) 8 Lactococcuslactis subsp. lactisbacteriophageP001 DSM4262 1) ; Lactococcuslactis subsp. lactisbacteriophageP008 DSM10567. Bothphagesshallbepropagatedonthef
43、ollowinghoststrain: Lactococcuslactis subsp. lactisF7/2 DSM4366. 5.2.2 Culturemediaandreagents 5.2.2.1General Thereagentsshallbeofanalyticalgradeand/orappropriateformicrobiologicalpurposes.Theyshallbefreefrom substancesthatcausetoxicorinactivatingeffectseithertothebacteriophagesortothehostbacteriaus
44、edfor countingphagederivedplaques. NOTE Toimprovereproducibility,itisrecommendedthatcommerciallyavailabledehydratedmaterialisusedforthe preparationofculturemedia.Themanufacturersinstructionsrelatingtothepreparationoftheseproductsshouldberigorously followed. 5.2.2.2Water Thewatershallbefreshlyglassdistilledwaterandno
