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ELSEVIERMarineChemistry51(1996)325-346CharacterizationofmarineandterrestrialDOMinseawaterusingexcitation-emissionmatrixspectroscopyPaulaG.CobleUniversityofSouthFloridaatSt.Petersburg,DepartmentofMarineScience,140SeventhAwe.S.,St.Petersburg,FL33701,USAReceived1March1995;accepted23June1995AbstractHigh-resolutionfluorescencespectroscopywasusedtocharacterizedissolvedorganicmatter(DOM)inconcentratedandunconcentratedwatersamplesfromawidevarietyoffreshwater,coastalandmarineenvironments.Severaltypesoffluorescentsignalswereobserved,includinghumic-like,tyrosine-like,andtryptophan-like.Humic-likefluorescenceconsistedoftwopeaks,onestimulatedbyUVexcitation(peakA)andonebyvisibleexcitation(peakC).Forallsamples,thepositionsofbothexcitationandemissionmaximaforpeakCweredependentuponwavelengthofobservation,withashifttowardslongerwavelengthemissionmaximumatlongerexcitationwavelengthandlongerwavelengthexcitationmaximumatlongeremissionwavelength.Atrendwasobservedinthepositionofwavelength-independentmaximumfluorescence(Exmax/Emmax)forpeakC,withmaximumatshorterexcitationandemissionwavelengthsformarinesamplesthanforfreshwatersamples.Meanpositionsofthesemaximawere:riversEx,,,/Em,,,=340/448nm;coastalwaterExmax/Emmax=342/442nm;marineshallowtransitionalEx,,,JEmmax=310/423nm;marineshalloweutrophicExmax/Emmax=299/389nm;andmarinedeepEx,,/Em,,,=340/438nm.Differencessuggestthatthehumicmaterialinmarinesurfacewatersischemicallydifferentfromhumicmaterialintheotherenvironmentssampled.TheseresultsexplainpreviousconflictingreportsregardingfluorescencepropertiesofDOMfromnaturalwatersandalsoprovideameansofdistinguishingbetweenwatermasssourcesintheocean.1.IntroductionTheconcentration,composition,andcyclingofdissolvedorganiccarbon(DOC)intheoceanhasbeenthefocusofintensestudyoverthepastfiftyyears.Thechemicalcomplexityandthelowconcen-trationofoneofthemajorcomponents,dissolvedhumicsubstances,makebalancingtheoceaniccar-bonbudgetespeciallydifficult.ThemethodsusedtoremoveDOMfromseawaterinsufficientquantitiesforcompositionalanalysesrequirelargevolumesandlongsampleprocessingtimes,introducingthepossi-bilityforcontaminationandchemicalalterationofthestartingmaterial,severelylimitingthenumberofsampleswhichcanbecollected,andpreventingsys-tematicanalysisofshort-termvariability.Onepropertyofdissolvedorganicmatter(DOM)whichiseasilymeasuredisfluorescence.ManyoftheearlieststudiesoffluorescenceintheoceanconcentratedonusingitasatracerofriverineinputofDOC(Kalle,1949;Duursma,1974),orasaproxyforsalinityandriver-bornesolublecontaminants0304-4203/96/$15.0001996ElsevierScienceB.V.AllrightsreservedSSDI0304-4203(95)00062-3326P.G.Cable/MarineChemistry51(1996)325-346(LaaneandKramer,1991,andreferencestherein).TwotypesofDOMfluorescencesignalshavebeenobservedinseawater,ahumic-like,orgelbstofffluo-rescence(Kalle,1949,1966;ZeppandSchlotzhauer,1981;Ewaldetal.,1983;Cobleetal.,1990,1993;Green,1992;MopperandSchultz,1993;deSouzaSierraetal.,19941,whichoccursat420-450nmfromexcitationat230-260and320-350nm,andaprotein-oraminoacid-likefluorescence(Traganza,1969;Ewaldetal.,1986;Cobleetal.,1990,1993;MopperandSchultz,1993;deSouzaSierraetal.,1994).Theprotein-likefluorescencearisesfromthefluorescenceofaromaticaminoacids,eitherfreeorasproteinconstituents,andisobservedatanemis-sionof300-305nm(tyrosine-like)and340-350nm(tryptophan-like)fromexcitationat220and275nm(Wolfbeis,1985).Attemptstodistinguishbetweenmarineandfresh-waterhumicsubstancesusingfluorescencespec-troscopyhavebeenlargelyunsuccessfulandhaveresultedinamultitudeofcontradictoryspectraldatabecausechoiceofexcitationandemissionwave-lengthsusedtomakeobservationshasbeenarbitraryandbecauseoffailuretocorrectforinstrumentalconfiguration(Ewaldetal.,1983;CobleandBrophy,1994;deSouzaSierraetal.,1994).Recenthigh-sensitivityspectrofluorometricstudieshavedemon-stratedsource-specificdifferencesinfluorescencespectraofseawaterrelativetofreshwatersamples(Donardetal.,1989;Green,1992;deSouzaSierraetal.,19941,betweensurfacewatersanddeepwatersatthesamemarinesite(Cauwetetal.,19901,betweenhumicsubstancesofdifferentorigin,andbetweenfulvicacidsandhumicacidsfromthesamesource(Mianoetal.,1988;Senesietal.,1991;MianoandSenesi,1992).Fluorescencecontouring,inwhichrepeatedemis-sionscansarecollectedatnumerousexcitationwavelengths,resultsinexcitation-emissionmatrices(EEMs)whichprovidehighlydetailedinformationwhichcanbeusedtoidentifyfluorescentcompoundspresentincomplexmixtures(Christianetal.,1981;LochmullerandSaavedra,1986;LeinerandWolf-beis,1988).ThetechniquehasrecentlybeenappliedsuccessfullytothestudyofseawaterDOM,whichisalsoacomplexmixtureoffluorescentcompounds(Cableetal.,1990,1993;Green,1992;Hawes,1992;MopperandSchultz,1993).ItissensitiveenoughtopermitanalysesonunconcentratednaturalDOMinbulkseawaterandfastenoughtobeusefulininvestigationsofsmall-scalevariationinDOMcomposition.EEMspectroscopy(EEMS)offersseveralmajoradvantagesoversingle-scanmethodologies.OncetheEEMhasbeenfullycorrectedforinstrumentalcon-figuration(Cableetal.,19931,datacanbeanalyzedasexcitationspectra,emissionspec