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/CENELECInternalRegulationswhichstipulatetheconditionsforgivingthisEurope an Standardthestatusofanationalstandardwithoutanyalteration.Uptodatelistsandbibliographicalreferencesconcernings uchnational standardsmaybeobtainedonapplicationtotheManagementCentreortoanyCENmember. ThisEuropeanSt
12、andardexistsinthreeofficialversions(English,French,German).Aversioninanyotherlanguagemadebytra nslation undertheresponsibilityofaCENmemberintoitsownlanguageandnotifiedtotheManagementCentrehasthesamestatusasthe official versions. CENmembersarethenationalstandardsbodiesofAustria,Belgium,CzechRepublic,
13、Denmark,Finland,France,Germany,Greece, Iceland,Ireland,Italy,Luxembourg,Malta,Netherlands,Norway,Portugal,Spain,Sweden,SwitzerlandandUnitedKingdom. EUROPEANCOMMITTEEFORSTANDARDIZATION COMITEUROPENDENORMALISATION EUROPISCHESKOMITEEFRNORMUNG ManagementCentre:ruedeStassart,36B1050Brussels 2002CEN Allri
14、ghtsofexploitationinanyformandbyanymeansreserved 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 Testmethods 7 5.1 Principle7 5.2 Materialandreagents . .7 5.3
15、 Apparatusandglassware . 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) Testforvalidationofdilutionneutralizationandmolecular
16、sievingmethods.25 AnnexB (informative) Neutralizers. .33 AnnexC (informative) Exampleofatypicaltestreport.35 AnnexD (informative) InformationontheapplicationandinterpretationofEuropeanstandardson chemicaldisinfectantsandantiseptics . 39 AnnexE (informative) ExampleofplaquesfromlysatesofphagesP001and
17、P00841 Bibliography 44EN13610:2002(E) 3 Foreword Thisdocument(EN13610:2002)hasbeenpreparedbyTechnicalCommitteeCEN /TC 216,“Antisepticsand chemicaldisinfectants“thesecretariatofwhichisheldbyAFNOR. ThisEuropeanStandardshallbegiventhestatusofanationalstandard,eitherbypublicationofanidenticaltextor byen
18、dorsement,atthelatestbyJune2003,andconflictingnationalstandardsshallbewithdrawnatthelatestby June2003. InthisEuropeanStandardtheAnnexAisnormativeandtheAnnexesB,C,DandEareinformative. AccordingtotheCEN/CENELECInternalRegulations,thenationalstandardsorganizationsofthefollowing countriesareboundtoimple
19、mentthisEuropeanStandard:Austria,Belgium,CzechRepublic,Denmark,Finland, France,Germany,Greece,Iceland,Ireland,Italy,Luxembourg,Malta,Netherlands,Norway,Portugal,Spain, Sweden,SwitzerlandandtheUnitedKingdom.EN13610:2002(E) 4 Introduction ThisEuropeanStandarddescribesasuspensiontestmethodintendedtoest
20、ablishwhetheraproductproposedas adisinfectantinthefieldsdescribedinclause1hasordoesnothavevirucidalactivityagainstbacteriophages. NOTE Virulentbacteriophages(phages)lyticfor(i.e.virulentfor)starterculturesof Lactococcuslactissubsp. lactisused fortheproductionofcheeseandotherfermentedmilkproductsareu
21、sedasmodelvirusestotestthevirucidalactivityofa product. Thelaboratorytestcloselysimulatespracticalconditionsofapplication.Chosenconditions(contacttime, temperature,virusesi.e.bacteriophagesinsuspension,.)reflectparameterswhicharefoundinpractical situationsincludingconditionswhichmayinfluencetheactio
22、nofdisinfectants. Eachutilizationconcentrationfoundfromthistestcorrespondstodefinedexperimentalconditions. Theconditionsareintendedtocovergeneralpurposesandtoallowreferencebetweenlaboratoriesandproduct types. Howeverforsomeapplicationstherecommendationsofusemaydifferandthereforeadditionaltestconditi
23、ons needtobeused.EN13610:2002(E) 5 1Scope ThisEuropeanStandardspecifiesatestmethod(phase2,step1)andrequirementsfortheminimumvirucidal activityagainstbacteriophagesofchemicaldisinfectantsthatformahomogeneous,physicallystablepreparationin hardwaterandthatareusedinfoodandindustrialareas,excludingareasa
24、ndsituationswheredisinfectionis medicallyindicatedandexcludingproductsusedonlivingtissues. ThisEuropeanStandardappliesatleasttothefollowing: a) processing,distributionandretailingof: 1) foodofanimalorigin: milkandmilkproducts; meatandmeatproducts; fish,seafood,andtheirproducts; eggsandeggproducts; a
25、nimal feeds; etc.; 2) foodofvegetableorigin: beverages; fruits,vegetablesandtheirderivatives(includingsugar,distillery.); flour,millingandbaking; animal feeds; .; b)other industrial areas: biotechnology(yeast,proteins,enzymes,.). UsingthisEuropeanStandard,itisnotpossibletodeterminethevirucidalactivi
26、tyagainstbacteriophagesof undilutedproductassomedilutionisalwaysproducedbyaddingtheinoculumandinterferingsubstance. Forchemicaldisinfectantsthatcanbeusedwithoutdilutionitisnotpossibletodeterminewhethertheseproducts, ataconcentrationabove80%haveavirucidalactivityagainstbacteriophages. NOTE Themethodd
27、escribedisintendedtodeterminetheactivityofcommercialformulationsoractivesubstanceson viruses(bacteriophages)intheconditionsinwhichtheyareused. ThisEuropeanStandardisapplicableonlytodisinfectants,complyingwiththevalidationtest(seeannexA).EN13610:2002(E) 6 2 Normativereferences Notapplicable. 3 Termsa
28、nddefinitions ForthepurposesofthisEuropeanStandard,thefollowingtermsanddefinitionsapply. 3.1 product(forchemicaldisinfectionand/orantisepsis) chemicalagentorformulationusedasachemicaldisinfectantorantiseptic EN1040:1997 3.2 virucideagainstbacteriophages productwhichinactivatesthebacteriophages NOTE
29、Theadjectivederivedfrom“virucide“is“virucidal“. 3.3 virucidalactivityagainstbacteriophages capabilityofaproducttoreducetheinfectivityofintactbacteriophagesbelongingtoreferencebacteriophage strainsP001andP008foratleast4lgundertheconditionsdefinedbythisstandard 3.4 infectivityofbacteriophages theabili
30、tyofabacteriophagetopropagateinasuitablehostbacterialcellresultinginthereleaseofbacteriophage progeny 3.5 inactivationofbacteriophage thereductionofinfectivityofabacteriophagebyaproductspecifiedasachemicaldisinfectant 3.6 referencebacteriophagesuspension bacteriophagesuspensionofadefinedvirusstrainm
31、aintainedinreferencecentersandwhichshouldnotbe passagedmorethan10times 3.7 stockbacteriophagesuspension bacteriophagesuspensionofadefinedstrainthathasbeenmultipliedonalargescaletoobtainabacteriophage suspensionrevealingidenticalcharacteristicsasthereferencebacteriophagesuspension NOTE Thisstockbacte
32、riophagesuspensionisusedtoprepareahightitertestbacteriophagesuspensionforthe bacteriophageinactivationtest. 3.8 hightiterbacteriophagesuspension thehightiterbacteriophagessuspensionobtainedfromagarplatesrevealingconfluentlysisinthebacteriallawn thatisusedtopreparethebacteriophagetestsuspensioninthev
33、irucidaltestingofthedisinfectant 3.9 bacteriophagetestsuspension thebacteriophagesuspensionofadefinedtiterthatisusedinthevirucidaltestingofthedisinfectantEN13610:2002(E) 7 4Requirements Theproductdilutedinhardwaterwhentestedinaccordancewithclause5shalldemonstrateatleasta4log 10 reductionofinfectivit
34、yofbacteriophageswhentestedinthepresenceofavolumefractionof1%acidicwhey (preparedfromacidifiedlowfatmilk)oroptionallyinthepresenceofavolumefractionof1%skimmilkasthe interferingsubstanceaccordingtoitspracticalapplicationsandundertherequiredtestconditions(20C,15min,2 referencebacteriophagestrains). Th
35、evirucidalactivityagainstbacteriophagesshallbeevaluatedusingthetwovirulentbacteriophages Lactococcus lactissubsp. lactisbacteriophageP001and Lactococcuslactis subsp. lactis bacteriophageP008. Bothphagesshallbepropagatedonthehoststrain Lactococcuslactis subsp. lactisF7/2. Thedeterminedvirucidalconcen
36、trationofthetestedproductissuggestedasbeingsuitableforpracticalsituations ofuse. Whereappropriate,additionalspecificvirucidalactivityagainstbacteriophagesshallbedeterminedunderother conditionsoftime,temperature,additionalstrainsandinterferingsubstancesinaccordancewith(see5.7.1)in ordertotakeintoacco
37、untintendedspecificuseconditions. NOTE Fortheseadditionalconditions,theconcentrationdefinedasaresultcanbelowerthantheoneobtainedunderthe initialtestconditionsof20C,15min,2selectedbacteriophagereferencestrains. 5Test methods 5.1Principle 5.1.1 Atestsuspensionofbacteriophagesinasolutionofinterferingsu
38、bstancesisaddedtoapreparedsample oftheproductundertestdilutedinhardwater. Themixtureismaintainedat20C1Cfor15min10s(requiredobligatorytestconditions). Attheendofthecontacttime,aliquotsaretakenandthevirucidalactivityagainstbacteriophagesinthisportionis immediatelyneutralizedorsuppressedbyavalidatedmet
39、hod.Themethodofchoiceisdilutionneutralizationwitha validatedneutralizer.Ifasuitableneutralizerisnotavailableforaspecificproduct,removaloftheproductby molecularsieving(i.e.gelfiltration)shallbeused. Thenumberofsurvivingbacteriophageparticlesandthenumberofbacteriophageparticlesinthetestsuspension ared
40、eterminedfromappropriatedilutionserieswithafactorof10preparedinmediumintesttubes. 5.1.2 Additionalandoptionalexposuretimes,temperaturesandinterferingsubstancesarespecified(see5.7.1). NOTE1 Thetestdescribedisbasedonanassessment(underspecificconditions)whichgivesareductionofatleast 99,99%(4lg)oftheinf
41、ectivityofthedifferentphagesafterdifferentcontacttimes. NOTE2 Forprincipalreasons,theresultofaninactivationappliedtoaviralpopulationisnotnecessarilyequalto100%:i.e ., onecannotconcludethatthereisa100%inactivationwhenonconductingtheexperimentnoinfectiousphagearefoundwithin alimitnumberofsampling. 5.2
42、Materialandreagents 5.2.1 Testorganisms Thevirucidalactivityagainstbacteriophagesshallbeevaluatedusingthetwofollowingbacteriophagestrains:EN13610:2002(E) 8 Lactococcuslactis subsp. lactisbacteriophageP001 DSM4262 1) ; Lactococcuslactis subsp. lactisbacteriophageP008 DSM10567. Bothphagesshallbepropag
43、atedonthefollowinghoststrain: Lactococcuslactis subsp. lactisF7/2 DSM4366. 5.2.2 Culturemediaandreagents 5.2.2.1General Thereagentsshallbeofanalyticalgradeand/orappropriateformicrobiologicalpurposes.Theyshallbefreefrom substancesthatcausetoxicorinactivatingeffectseithertothebacteriophagesortothehost
44、bacteriausedfor countingphagederivedplaques. NOTE Toimprovereproducibility,itisrecommendedthatcommerciallyavailabledehydratedmaterialisusedforthe preparationofculturemedia.Themanufacturersinstructionsrelatingtothepreparationoftheseproductsshouldberigorously followed. 5.2.2.2Water Thewatershallbefreshlyglassdistilledwa
