短链氯化石蜡分析方法-3

短链氯化石蜡分析方法-3

EcotoxicologyandEnvironmentalSafety67(2007)206–211

http://www.77cn.com.cn/locate/ecoenv

Highlightedarticle

Effectsofshort-chainchlorinatedparaf nsonsoilorganisms

ernohla,JitkaCvkova,KlaraKobeticova,JanLana,JitkaBezchlebova

,JakubHofmanÃIvanaSochova

FacultyofScience,MasarykUniversity,ResearchCentreforEnvironmentalChemistryandEcotoxicology,Kamenice126/3,Brno,62500CzechRepublic

Received8June2006;receivedinrevisedform8December2006;accepted23December2006

Availableonline26March2007

Abstract

Despitethefactthatchlorinatedparaf nshavebeenproducedinrelativelylargeamounts,andhighconcentrationshavebeenfoundinsewagesludgeappliedtosoils,thereislittleinformationontheirconcentrationsinsoilsandtheeffectonsoilorganisms.Theaimofthisstudywastoinvestigatethetoxicityofchlorinatedparaf nsinsoils.Theeffectsofshort-chainchlorinatedparaf ns(64%chlorinecontent)oninvertebrates(Eiseniafetida,Folsomiacandida,Enchytraeusalbidus,Enchytraeuscrypticus,Caenorhabditiselegans)andsubstrate-inducedrespirationofindigenousmicroorganismswerestudied.Differenceswerefoundinthesensitivityofthetestedorganismstoshort-chainchlorinatedparaf ns.F.candidawasidenti edasthemostsensitiveorganismwithLC50andEC50valuesof5733and1230mg/kg,respectively.Toxicityresultswerecomparedwithavailablestudiesandthepredictednoeffectconcentration(PNEC)of5.28mg/kgwasestimatedforthesoilenvironment,basedonourdata.r2007PublishedbyElsevierInc.

Keywords:Soilinvertebrates;Soilmicroorganisms;Short-chainchlorinatedparaf ns;Soilpollution;Riskassessment

1.Introduction

Chlorinatedparaf ns(CPs;chlorinatedn-alkanes)areindustrialchemicalscomprisingofchlorinatedstraightchainhydrocarbons.Thesechemicalshavebeenproducedsincethe1930sandusuallyexistasmixtures.Theycanbesubdividedintothreemaingroupsaccordingtothelength(thenumberofcarbons)ofthealkanechain:short(SCCPs;C10–13),medium(MCCPs;C14–17)andlong(LCCPs;C417;EuropeanCommission,1999).Chlorinatedparaf nsarewidelyusedasadditivesinmetalworking uids, ameretardantsinrubbers,additivesinpaints,coatings,sealantsandadhesives.Duetotheirhighchemicalandthermalstability,theywereusedtoreplacepolychlorinatedbiphenylsinthemid1980s.Inthelastdecade,theannualworld-wideproductionofchlorinatedparaf nswasesti-matedtobe300ktons(EuropeanCommission,1999).DuetothewidespreaddistributionintheenvironmentandhighlogKowvalues(5–12.6;Tomyetal.,1998),CPsexertahigh

Correspondingauthor.Fax:+420549492840.

E-mailaddress:hofman@recetox.muni.cz(J.Hofman).0147-6513/$-seefrontmatterr2007PublishedbyElsevierInc.doi:10.1016/j.ecoenv.2006.12.015

potentialforbioaccumulation,withstrongsorptiononsewagesludge,soilsandsedimentsandverylowmobility.Levelsofchlorinatedparaf nsinsomeenvironmentalmatricesarerelativelywellknown.Levelsinwaterhavebeendetectedwithintherange0.05–6mg/l(Tomyetal.,1998).Concentrationsof0.0005–1000mg/kg(samplingarearemotefromanindustrializedarea)and1.5–18mg/kg(nearanindustrialarea)weredetectedinriversediments(Tomyetal.,1998).Additionally,concentrationsofshort-andmedium-chainchlorinatedparaf nsof69–431mg/kgwerefoundinsedimentsfromtheNorthandBalticseas(HuttigandOehme,2005).Tomyetal.(1998)summarizedtheavailabledataonthelevelsofshort-andmedium-chainchlorinatedparaf nsinsewagesludgefromindustrializedareas(0.76–65mg/kg).Concentrationsupto93mg/kgwerefoundinUKsewagesludgefromanindustrialareabyNichollsetal.(2001).Itiswellknownthattheapplicationofsewagesludgetoagriculturalsoilsimprovessoilqualitybut,ontheotherhand,itmayalsoincreasetheconcentrationofcontaminantsinsoils.However,Nichollsetal.(2001)reportedthatconcentrationsofSCCPsinfarmsoilsaftertheapplicationofdigestedsewagewithhighlevelsofchlorinatedparaf nswerebelowthelimitofthe

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detection(0.1mg/kg).Nevertheless,detectableamounts

(upto1.7mg/kgwetwt)werefoundinearthwormscollectedinthesesoils,implyingthatthebioconcentrationofchlorinatedparaf nsmayoccurinsewagesludge-treatedsoils.Nootherstudiesareavailableonchlorinatedparaf nlevelsinsoilsundersewagesludgeapplication.

Despiteahighbioconcentrationpotentialandhighlevelsofchlorinatedparaf nsinsewagesludgeappliedtosoils,thereislimiteddataonthetoxicityofCPstosoilorganisms.Hence,inthisstudy,thetoxicityofanSCCPmixture(64%chlorinecontent)wasstudiedusingabatteryofsoiltoxicitytests.EffectsonsurvivalandreproductionofFolsomiacandida,Eiseniafetida,Enchytraeusalbidus,Enchytraeuscrypticus,Caenorhabditiselegansandonsubstrate-inducedrespirationofmicroorganismsweredeterminedandcomparedwiththeresultsfromSverdrupetal.(2006).Thepredictednoeffectconcentration(PNEC)wascharacterizedaccordingtotheEuropeanCommission(2003)guidelinebasedonourresultsandavailabledatafromtheliterature.

2.Materialsandmethods2.1.Testorganisms

Allinvertebrates(F.candida,E.fetida,E.crypticus,E.albidus,andC.elegans)havebeenculturedatRECETOXlaboratories(Brno,CzechRepublic).F.candida(Insecta:Collembola)waskeptonplasterofParisandpulverizedactivatedcharcoalinratioof9:1(w:w)at18721Cindarkness.Onceaweek,drybaker’syeastswereaddedasfood.E.fetida(Annelida:Oligochaeta)wasculturedinthemixtureofsphagnumpeatandhorsemanureintheratioof1:1(w:w)thatwasadjustedtopHof6–7withcalciumcarbonate(CaCO3).Thewatercontentwas$80%ofthemaximumwaterholdingcapacity(WHCmax).Additionalfeedingwasnotnecessary.Theculturewasmaintainedat20721Cinthedark.E.crypticus(Annelida:Oligochaeta)wasculturedinOECDarti cialsoil(OECD,2000a;50%ofWHCmax)andE.albidus(Annelida:Oligochaeta)waskeptincommercialgardensubstrate(80%ofWHCmax;1%CaCO3).Bothspeciesweremaintainedat18721Cindarknessandwerefedwithoat akesweekly.C.elegans(Nematoda),wildtypestrainN2,var.Bristol,culturewaskeptonNGMagarwithbacteriallawnofuracil-de cientstrainofEscherichiacoli(OP50)asafoodsourceandwasmaintainedat20721Cindarkness.Theindigenousmicrobialcommunityofanaturalsoilwasusedinmicrobialtest.

2.2.Preparationofsoil

OECDarti cialsoilwaspreparedforF.candida,E.fetida,E.albidusandE.crypticustests.Thesoilcompositionwas70%sand,20%kaolinclayand10% nelygroundsphagnumpeat(OECD,2000a).Theorganicmattercontentwas4.7%.ThepHKClwassetto6.070.5withCaCO3atthebeginningoftestsandwasfoundtoincreaseto6.570.5attheendofthetests.Thearti cialsoilwasfoundtobeanunsuitablesubstrateintheC.eleganstestbecausethe rmpeat oatedonthesurfaceduringtheextractionprocedureanddisabledcountingofsurvivingworms.Hence,thenaturalsoilwasusedinthistestandinthemicrobialtest.Thissoilwascollectedfromthetoplayerofa eldnearBrno(CzechRepublic).Thesoilwasaloamysandcambisolwiththefollowingparticlesizedistribution:sand(450mm)64.4%,silt(2–50mm)29.1%,clay(o2mm)6.5%.Thetotalcationexchangecapacitywas20meq/http://www.77cn.com.cnaniccarbonandtotalnitrogencontentswere2.35%and0.27%,http://www.77cn.com.cnanicpollutantsandheavymetalscontentswerecompar-abletothebackgroundlevelsaccordingtotheCzechRepublicguideline

(MinistryofEnvironmentofCzechRepublic,1996).ForC.eleganstest,thesoilwasair-driedatroomtemperatureandthensieved(o2mm),defaunated(deepfreezing)andstoredunderdryconditionsanddarkness.Forthemicrobialtest,thefreshsoilwassieved(o2mm)aftersamplingandstoredin41Cindarkness.Thewatercontentwasadjustedto50%ofWHCmax(arti cialsoil)and$80%ofWHCmax(C.eleganstest).Thewaterdecreaseininvertebratetestswascheckedweeklybyweighingandwaterwasreplenishedifnecessary.Thewatercontentinthemicrobialtestisdescribedthereinafter.

2.3.Preparationoftestsubstanceandspikingprocedure

Chlorinatedparaf ns(labeledasC12,64%chlorinecontentbyweight;

aviscoushoney-likeliquid)wereprovidedbyNova

ckezavodyInc.(Slovakia)asanindustrialproduct.BasedonchemicalanalysisbySCGC/LRMS-ECNI(HP5890SeriesIIgaschromatograph)describedinStejnarovaetal.(2005),thismixtureincludedallshort-chainparaf nfractions(C10–13)withacompositionofC106%,C1137%,C1232%andC1325%.

Stocksolutionswerepreparedbydissolvingparaf nsinacetone(HPLCpurity,Merck,CzechRepublic)fortestswithinvertebratesandcyclohexane(HPLCpurity,Merck,CzechRepublic)fortestswithmicroorganisms.Stocksolutionscorrespondingtothehighestconcentra-tionsweremixedinalltestsseparately.Dilutionsfromthestocksolutionweremadeusingacetoneorcyclohexane.Range- ndingtestsfoundlowSCCPtoxicityandthemaximumconcentrationrequiredbytherespectiveguidelines(1000mg/kg)didnotaffectmostoftheorganisms.Hence,thetestedconcentrationseriesinthetestsweresetfrom100to10000mg/kg.Thesoilsurfaceineachcontainerwassprinkledwiththeacetone/cyclohexanesolutiontoreachtheappropriateconcentrationinthesoil.Solutionswereappliedtodryormoistsoil(40%ofWHCmax)forinvertebrateormicrobialtests,respectively.Thevolumeofsolutionscorrespondedto10%ofsoildrywtinalltests.Allcontainerswerekeptinthefumehood(24h)toevaporatethesolvent.Aftertheevaporation,soilsweremixedthoroughly,distilledwaterwasaddedatanappropriatevolumeandsoilsweremixedagain.Pureacetone/cyclohexanecontrolstochecksolventtoxicityandwatercontrols(withoutsolventapplication)werepreparedineachtest.VolumesofthechemicalinsoilswereanalyzedatthebeginningofthetestaccordingtoStejnarovaetal.(2005)bySCGC/LRMS-ECNI(HP5890SeriesIIgaschromatograph)andinitialsoilconcentrationscorrespondedwelltonominalconcentrations.Allcon-centrationswereexpressedonasoildryweightbasisasnominal(initiallyadded)concentrations.

2.4.Folsomiacandidatest

ThetestwasperformedaccordingtoISO11267(1999).Eachtestcontainercontained30gdrywtofthesoil.Exposureconcentrationswere78.125,156.25,312.5,625,1250,2500,5000,10000mg/kg.Fivereplicateswereusedforeachconcentration.Tensynchronized,10–12dold,organismswereintroducedintoeachtestcontainer.Theexposureperiodwas28dandcontainersweremaintainedat201Cin16/8light–darkcycle.Drybaker’http://www.77cn.com.cnanismswereextractedby otationandanimals oatedonthesurfaceweremanuallycounted.

2.5.Eiseniafetidatest

ThetoxicitytestwasperformedaccordingtoOECDguideline222(OECD,2000a).Fiveexposureconcentrations(100,320,1000,3200,10000mg/kg)wereused.Threecontainers(1lvolume)pereachconcentrationwith500gdrywtofthesoilwerepreparedand10adultwormswithclitellum(300–400mg)wereintroducedintoeachcontainer.Food(5gofdrygroundhorsemanure)wasaddedweeklyunderthesoilsurface.Thesurvivalandreproduction(thenumberofcocoons)were

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evaluatedafter4weeksbymanualcounting.Thetestwaskeptin20721C

andunder16/8light–darkcycle.

2.9.Dataanalysis

AllstatisticalanalysesweredonewithSTATISTICA6.0software(StatSoft,Inc.,2004).Theconcentrationatwhich50%ofadultsurvivalwasobserved(LC50)and50%effectconcentrationforthereproductiveoutput(EC10andEC50)werecalculatedaccordingtoHaanstraetal.(1985)usinglogisticregressionanalysis.Thestandarderrorwasusedfortheexpressionofthedatavariation.Noobservedeffectconcentrations(NOECs)andlowesteffectconcentrations(LOECs)weredeterminedbyanalysisofvariance(ANOVA)andDunnett’sprocedureata5%signi cancelevel.

2.6.Enchytraeidtests

TestswithbothenchytraeidspecieswereperformedaccordingtoOECDguideline220(OECD,2000b).Tenand20gdrywtofthesoilwaspreparedfortestswithE.crypticusandE.albidus,respectively.Exposureconcentrationswereequalinbothtests(500,1000,3000,6000,10000mg/kg).Fivereplicatesfortreatmentandeightreplicatesforthecontrolswereprepared.Tenadultwormswithclitellumwereaddedtoeachcontainerinbothtests.Animalswereexposedfor42d(E.albidus)and28d(E.crypticus).Testswerekeptin20721Candunder16/8light–darkcycle.Approximately50mgofdrygroundoat akespercontainerwereaddedeveryweek.Attheendoftheexposureperiods,thewormswerekilledby5mlofethanolappliedtothesoilanddyedbyBengalred.Thesurvivalofadultsandthenumberofjuvenilesweremanuallycounted.

3.Results

Resultsobtainedfromallthetestswerevalidaccordingtothecriteriade nedinthetestguidelines.Nosigni cantdifferenceswereobservedbetweensolventandwatercontrols(ANOVA;po0.05).Nosigni cantmortalityofE.fetidaandE.albidusadults(ANOVA;Dunnett’stest;po0.05)occurredatanyexposureconcentration.Short-chainchlorinatedparaf nshadsigni canteffect(ANOVA;Dunnett’stest;po0.05)onE.crypticusmortalityatthehighestexposureconcentration(70%survivalincompar-isonwithsolventcontrol),however,adose–responsecurvecouldnotbeestablishedandonlyNOECandLOECvaluesweredeterminedforthisendpoint.Dose–responsecurvesofadultmortalitywereestablishedforF.candidaandC.elegansandLC50valueswerecalculated.Thereproduc-tionprovedtobeamoresensitiveendpointthatallowedthecreationofdose–responsecurves,andthecalculationofEC50valuesforallspecies,exceptC.eleganswherethereproductionwasnotevaluated.EC10valueswerecalcu-latedonlyforthereproductionofF.candidaandE.fetida.ResultsofinvertebratetestsaresummarizedinTable1.Effectsonsubstrate-inducedrespirationofmicroorganismswereevaluatedafter2and4weeks.Thisendpointwasnotaffectedat14doftheincubationatanyexposureconcentration.Hence,thehighestexposureconcentration(10000mg/kg)wasestablishedastheNOECvalue.After28doftheincubation,asigni cantdecreaseofmicrobial

2.7.Caenorhabditiseleganstest

ThetestwasperformedaccordingtoASTM(2001)guideline.Threereplicateswith2.33gdrywtofthesoilwereprepared.Fiveexposureconcentrations(500,1000,3000,6000and10000mg/kg)wereusedforthetest.Tenage-synchronizedadults(3–4d)wereintroducedintoeachtestcontainer.Thetesttook48handwasmaintainedat201Cindarkness.Aftertheexposureperiod,theadultsurvivalwasevaluatedbyLUDOXextractionandcountsofanimalsweredeterminedundermicroscope.

2.8.Microbialtest

ThetestwasperformedaccordingtoOECD(1999)guidelinedraft217.Tengramsofthesoiladjustedto40%ofWHCmaxwerepreparedandthreereplicatesweredesignedforalltreatedlevels.Thepreincubationwascarriedoutduring4d.Afterthat,exposureconcentrations(500,1000,5000,10000mg/kg)werepreparedasdescribedabove.Waterlossaftertheevaporationwasreplenishedandtotalwatercontentwasadjustedto60%ofWHCmax.Bottleswithsoilwerecoveredbyrubbercapsandincubatedat221Cindarkness.Soilsampleswereanalyzedforglucose-inducedrespirationrateafter14and28doftheincubation.TherespirationwasmeasuredasCO2productionwithinthe rst6haftertheadditionofglucose(5mgCglucose/gdrywtsoil)(ISO14240-1,1997).Gassamples(1ml)werewithdrawnfromthebottlesbyasyringe,andCO2wasquanti edbyGCwithH2mobilephase,PorapackQstationaryphase,andthermalconductivitydetector.

Table1

SummaryoftoxicitytestresultsofSCCPs(64%chlorinecontent)

Adultsurvival

TestorganismFolsomiacandida*Eiseniafetida*

Enchytraeusalbidus*Enchytraeuscrypticus*Caenorhabditiselegans**

Testduration2828422848ddddhrs

NOEC(mg/kg)125010000b10000b60001000

LOEC(mg/kg)2500NDND10000b3000

LC50(95%CI)a(mg/kg)

5733(4483–7149)NDNDND

8836(6003–11668)

ReproductionNOEC(mg/kg)625100030006000

LOEC(mg/kg)12503200600010000b

EC50(95%CI)a(mg/kg)1230284960277809

(1009–1451)(2170–3529)(3576–8478)(4381–11237)

EC10(95%CI)a(mg/kg)660(341–870)1158(336–1980)NDND

*Arti cialsoil.

**Loamysandsoil.

a,CIcon denceinterval;b,thehighesttestedconcentration;ND,couldnotbeestimated.

Estimatedvaluesfor50%effectonadultsurvival(LC50),10%and50%effectonreproduction(EC10andEC50)andnoobservedeffectconcentration(NOEC)andlowestobservedeffectconcentration(LOEC)forbothendpointsarereportedforF.candida,E.fetida,E.albidus,E.crypticusandC.elegans.

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respiration(ANOVA;Dunnett’stest;po0.05)wasfound

atthehighestexposureconcentration(72%ofsolventcontrol),andconcentrationsof10000and5000mg/kgweredesignatedasNOECandLOECvalues,respectively.Allresultsanddose–responserelationshipsarepresentedinFig.1.4.Discussion

4.1.Sensitivityofspecies

Differencesinecologicalstrategy,lifecyclehistoryand/orexposureroutemakethecomparisonoforganismsdif cult.Moreover,experimentaldesignswerenotidenticalforalltests.SinceF.candida,E.fetida,E.albidusandE.crypticusweretestedundersimilarconditions(arti cialsoil,moisturecontent,temperature,testduration,end-points),resultsofthesetestsmaybecompared.F.candidawasshowntobethemostsensitiveorganisminbothevaluatedendpoints.This ndingisinaccordancewithpreviouslypublishedstudies(Sverdrupetal.,2002;LockandJanssen,2002).ThelethalityendpointshowedsimilarlylowsensitivityinallOligochaetatests.However,earthwormswereshowntoberelativelysensitivetoSCCPswhenthereproductionwastakenastheendpoint.Enchytraeidspeciesaretaxonomicallyclosetoearthwormsandsomestudiesreportedsimilarsensitivityofthesespecies(RombkeandMoser,2002).However,resultsobtainedinourstudyareinaccordancewithotherstudiesreportinglowersensitivityofenchytraeidspeciestosomechemicals(e.g.PAHsandmercuryreportedbySverdrupetal.,2002andLockandJanssen,2001).Withinthisgroup

Fig.1.

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oftestswithsimilartestconditions,wecandeterminethe

followingorderofspeciessensitivitytoSCCPsrankedfromthemosttoleastsensitive:F.candida4E.fetida4E.albidus$E.crypticus.

NematodaC.eleganswasnotcomparedtootherorganismswithregardtodifferenttestconditions.Despitethisfact,C.elegansvaluesoftheNOECandLC50didnotdifferfromothersmarkedly.Substrate-inducedrespirationofmicroorganismswasrelativelyinsensitivetoSCCPtoxicity.Thisendpointdescribestheactivityofwholesoilmicrobialcommunityasa‘‘blackbox’’andpossiblechangesinthestructureofmicrobialcommunityinducedbycontaminantmightbenotrecognized.Hence,otherparameters(e.g.nitri cation;vanBeelenandDoelman,1997;Sverdrupetal.,2006)describingasensitivepartofthemicrobialcommunity,shouldbeselectedforthetoxicitytesting.

Consideringallofthesefacts,F.candidamayberecommendedforthetoxicitytestingoforganicchemicalsforitshighsensitivityinbothendpointsandlowtestdemands.ThereproductionofE.fetidaisalsosensitiveinourstudyandmayberecommended.However,thelowsensitivityofthemortalityofadultsaswellasthehightestdemandsmakethisorganismlesssuitable.Otherorganismsprovedthelowsensitivityandhighdatavariabilityinourstudy.However,sinceotherchemicalsmaydifferfromSCCPsinbioavailability,exposureroutes,mechanismsofaction,etc.,thisrankingofsensitivityoforganismscannotbegeneralizedforstudyofotherchemicals.Therefore,abatteryoforganismsshouldbeusedfortestingandobtainingdataforriskassessmentasrecommendedinmanystudies(e.g.Cortetetal.,1999;Bierkensetal.,1998;HundandTraunspurger,1994)http://www.77cn.com.cnparisonwithavailablestudies

WhereasinformationonthetoxicityofSCCPstoaquaticinvertebratesand shwerewellgatheredintheRiskAssessmentReport(EuropeanCommission,1999)andeffectsonmammalsarealsoknown(Duncanetal.,1980),informationontoxicityinsoilsislimited.Sverdrupetal.(2006)publisheddataoneffectsofSCCPs(C10–13;60%chlorinecontent)onnitrifyingmicroorganisms,E.crypticusandredclovers(Trifoliumpratense).Thetestwascarriedoutinagriculturalsoilandexposurelevelsdidnotexceedtheconcentrationof1000mg/kg.Neitherenchytraeidreproductionnorweightofredcloverseedswereaffectedbytheseconcentrations,whichcorrespondedwithourresults.FornitrifyingmicroorganismsEC10wasdeterminedas570mg/kg.ThisvalueissimilartotheEC10value(660mg/kg)forF.candidareproductioninourstudy,neverthelessveryfarfrom10000mg/kgwhichcausedonlycca10%inhibitionofsubstrateinducedrespirationinourstudy.

Inanotherstudy(Thomsonetal.,2001),effectsofMCCPs(C14–17;54%chlorinecontent)toE.fetidaweredeterminedusingthearti cialsoiltest.Theadultmortality,

effectsonthegrowthandreproduction(thenumberofjuveniles)wasevaluatedwithNOECvaluesof3200,1000and320mg/kg(nominalconcentrations),respectively.Identicalconcentrationswereusedinourstudytomakethecomparisonofresultspossible.MCCPsprovedthehighertoxicitytoE.fetidamortality.Thereproductionresultscannotbecomparedtodatafromourstudyduetodifferencesinevaluatedreproductionendpoints(numbersofjuvenilesandcocoons).SinceSCCPsaregenerallyregardedtobemoretoxicthanthosewithmedium-orlong-chains(Tomyetal.,1998),itwassurprisingthattheeffectofSCCPstoE.fetidamortalitygatheredinourstudywaslowerthaneffectofMCCPsreportedbyThomsonetal.(2001).Thissupportsourabove-discussedopinionconcerninglowsensitivityofthisendpointinourwork.4.3.Ecologicalriskassessment

EcologicalriskassessmentofSCCPswaselaboratedbyEuropeanUnion(EuropeanCommission,1999).Thesigni cantriskfortheaquaticenvironmentwasindicatedforsomelocalsources(EuropeanCommission,1999).ThisevaluationwasnotdonefortheterrestrialcompartmentduetolimitedknowledgeonSCCPsinsoils.Inthatreport,thePNECvalueforterrestrialcompartmentwassuggestedtobe0.8mg/kgbasedontheequilibriumpartitioningmodelandlevelsinsoilsweremodeledusingtheEUSESmodel.Recently,Sverdrupetal.(2006)estimatedthePNECas57mg/kgwheretheEC10valueforeffectonnitrifyingmicroorganisms(570mg/kg)andassessmentfactorof10(threetrophicleveltoxicitydata)accordingtoguidelineEuropeanCommission(2003)wereusedforPNECcalculation.Inourstudy,theEC10valueofF.candidareproduction(660mg/kg)wasappliedtothePNECcalculation.Datawasextrapolatedaccordingtoguideline(EuropeanCommission,2003)tostandardsoilorganiccarboncontent(2%;inthepresentstudy,soilorganiccarboninF.candidatestwas5%)andassessmentfactorof50wereused(forlong-termtoxicitytestsoftwotrophiclevels).Afterdataextrapolation,the nalPNECvalueof5.28mg/kgwasestimatedbasedonourdata.ThisstudytogetherwiththatofSverdrupetal.(2006)providedPNECvaluesbasedonalargedatasetincludingthreetrophiclevels.However,moreinformationonenviron-mentallevels,bioavailabilityandfateintheenvironmentshouldbefoundtoestimatetheriskforterrestrialenvironment.5.Conclusion

Withinthetestbattery,F.candidaprovedtobethemostsensitiveorganismstoSCCPs(64%chlorinecontent).EstimationsofEC10,EC50andLC50valuesforalltestedorganismswereintheconcentrationrangeof600–9000mg/kg.ThelowestEC10valueof660mg/kgwasfoundfortheF.candidareproduction.Theleastsensitivewassubstrate-inducedrespirationofsoilmicroorganisms.Thecompar-

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isonofourdatawithavailablestudiesonCPsshowed

relativelysimilarresults,despitethedif cultyofaninter-laboratorycomparison.ThePNECforsoilorganismswasestimatedbasedonourdatawithresultingvalueof5.28mg/kgforSCCPs(64%chlorinecontent)buttheriskcannotbeevaluatedbecauseareliablePECismissing.Acknowledgments

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