22/03/2016

DissolvedOxygenbytheWinklerMethod

TheWinklerMethodMeasuringDissolvedOxygen
CreatedbyMonicaZ.Bruckner,MontanaStateUniversity

WhatistheWinklerMethod?
TheWinklerMethodisatechniqueusedtomeasuredissolved
oxygeninfreshwatersystems.Dissolvedoxygenisusedasan
indicatorofthehealthofawaterbody,wherehigherdissolved
oxygenconcentrationsarecorrelatedwithhighproductivityand
littlepollution.Thistestisperformedonsite,asdelaysbetween
samplecollectionandtestingmayresultinanalterationinoxygen
content.

HowdoestheWinklerMethodWork?
TheWinklerMethodusestitrationtodeterminedissolvedoxygenin TheWinklermethodisdonebynotingacolorchangewhen
thewatersample.Asamplebottleisfilledcompletelywithwater
titratingafreshwatersample.PhotoprovidedbyMonicaBruckner.
(noairislefttoskewtheresults).Thedissolvedoxygeninthe
sampleisthen"fixed"byaddingaseriesofreagentsthatformanacidcompoundthatisthentitratedwithaneutralizingcompoundthat
resultsinacolorchange.Thepointofcolorchangeiscalledthe"endpoint,"whichcoincideswiththedissolvedoxygenconcentrationinthe
sample.Dissolvedoxygenanalysisisbestdoneinthefield,asthesamplewillbelessalteredbyatmosphericequilibration.

Applications
Dissolvedoxygenanalysiscanbeusedtodetermine:
thehealthorcleanlinessofalakeorstream,
theamountandtypeofbiomassafreshwatersystemcansupport,
theamountofdecompositionoccurringinthelakeorstream.

HowtoSampleCollection,Preparation,AnalyticalProtocols,andConcerns
Dissolvedoxygenshouldbemeasuredasquicklyandcarefullyaspossible.Ideally,samplesshouldbemeasuredinthefieldimmediately
aftercollection.ThefollowingprotocolisadaptedfromWashingtonStateUniversity'sHowtoMeasureDissolvedOxygen(moreinfo)
website.
ReagentList:
2mlManganesesulfate
2mlalkaliiodideazide
2mlconcentratedsulfuricacid
2mlstarchsolution
Sodiumthiosulfate
Thesereagentsareavailableindissolvedoxygenfieldkits,suchasthosemadebytheHachCompany.Pleaseusecautionwhenusingthese
reagents,astheycanbehazardoustoone'shealth.
Procedure:
1.Carefullyfilla300mLglassBiologicalOxygenDemand(BOD)stopperedbottlebrimfullwithsamplewater.
2.Immediatelyadd2mLofmanganesesulfatetothecollectionbottlebyinsertingthecalibratedpipettejustbelowthesurfaceofthe
liquid.(Ifthereagentisaddedabovethesamplesurface,youwillintroduceoxygenintothesample.)Squeezethepipetteslowlyso
nobubblesareintroducedviathepipette.
3.Add2mLofalkaliiodideazidereagentinthesamemanner.
4.Stopperthebottlewithcaretobesurenoairisintroduced.Mixthesamplebyinvertingseveraltimes.Checkforairbubbles
discardthesampleandstartoverifanyareseen.Ifoxygenispresent,abrownishorangecloudofprecipitateorflocwillappear.
Whenthisflochassettletothebottom,mixthesamplebyturningitupsidedownseveraltimesandletitsettleagain.
http://serc.carleton.edu/microbelife/research_methods/environ_sampling/oxygen.html

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DissolvedOxygenbytheWinklerMethod

5.Add2mLofconcentratedsulfuric
acidviaapipetteheldjustabovethe
surfaceofthesample.Carefully
stopperandinvertseveraltimesto
dissolvethefloc.Atthispoint,the
sampleis"fixed"andcanbestored
forupto8hoursifkeptinacool,
darkplace.Asanaddedprecaution,
squirtdistilledwateralongthe
stopper,andcapthebottlewith
aluminumfoilandarubberband
duringthestorageperiod.
6.Inaglassflask,titrate201mLofthe
samplewithsodiumthiosulfatetoa
palestrawcolor.Titratebyslowly
droppingtitrantsolutionfroma
calibratedpipetteintotheflaskand
continuallystirringorswirlingthe
samplewater.
7.Add2mLofstarchsolutionsoablue
colorforms.
8.Continueslowlytitratinguntilthe
sampleturnsclear.Asthisexperiment
reachestheendpoint,itwilltakeonly
onedropofthetitranttoeliminatethe
bluecolor.Beespeciallycarefulthat
eachdropisfullymixedintothe
samplebeforeaddingthenext.Itis
sometimeshelpfultoholdtheflaskup
toawhitesheetofpapertocheckfor
absenceofthebluecolor.
9.Theconcentrationofdissolved
oxygeninthesampleisequivalentto
thenumberofmillilitersoftitrant
used.EachmLofsodiumthiosulfate
addedinsteps6and8equals1mg/L
dissolvedoxygen.

ResultsAnalysis

TheWinklermethodformeasuringdissolvedoxygeninvolvestitratingasamplewitha
seriesofreagents.SchematicdiagramcourtesyoftheWashingtonStateUniversity
DepartmentofEcology,whichcanbeaccessedviathislink:HowtoMeasureDissolved
Oxygen(moreinfo).

Thetotalnumberofmillilitersoftitrantusedinsteps68equalsthetotaldissolvedoxygeninthesampleinmg/L.Oxygensaturationis
temperaturedependentgasismoresolubleincoldwaters,hencecoldwatersgenerallyhavehigherdissolvedoxygenconcentrations.
Dissolvedoxygenalsodependsonsalinityandelevation,orpartialpressure.Clickheretoaccessanonlineoxygensaturationcalculator
thattakestheseparametersintoaccount.Otherwise,usethechartbelowtofindsaturationatagiventemperature.

Oxygensolubilityatdifferentwatertemperatures.ModifiedfromWashingtonStateUniversity
EcologyCitizen'sGuidetoWaterQuality.

http://serc.carleton.edu/microbelife/research_methods/environ_sampling/oxygen.html

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