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Doestheoptimumtemperaturefortherateofreactionoftheenzymecatalase,asmeasuredbyoxygen

production,varybetweenplantandanimalcells,whencomparing:leek,potato,chickenliver,porkliverandfishliver;anddoesitmakeadifferenceiftheanimalis

homeothermicorpoikilothermic?

BiologyExtendedEssayCandidateSessionNumber:

CandidateName:Wordcount:3967

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TableofContents

Acknowledgements.....................................................................................................3

Abstract......................................................................................................................4

Introduction................................................................................................................5

Investigation...............................................................................................................7Variables........................................................................................................................................................................7Materials........................................................................................................................................................................9Method............................................................................................................................................................................9Uncertainties............................................................................................................................................................11Results.........................................................................................................................................................................11Observations.............................................................................................................................................................11Processeddata.........................................................................................................................................................12DiscussionandReasonedArgument..........................................................................17

Conclusion.................................................................................................................20

Evaluation.................................................................................................................21

WorksCited...............................................................................................................22

Appendices...............................................................................................................24Appendix1.................................................................................................................................................................24Appendix2.................................................................................................................................................................26

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Acknowledgements Iwouldfirstlyliketothankmysupervisor_____________forprovidingmewiththebestsupportandguidance.Iamalsoveryappreciativeofthehelpthatourlabtechnician,______________,gavemeduringmyexperiment.Mymotheralsocontributedbypurchasingthesourcesofcatalasethatwasrequiredformyexperiment.

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Abstract Theresearchquestionforthisessayis:Doestheoptimumtemperaturefortherateofreactionoftheenzymecatalase,asmeasuredbyoxygenproduction,varybetweenplantandanimalcells,whencomparing:leek,potato,chickenliver,porkliverandfishliver;anddoesitmakeadifferenceiftheanimalishomeothermicorpoikilothermic?Thisinvestigationiscarriedoutthroughacomparisonofoptimumtemperaturesbyanexperimentwheretheenzymecatalasebreaksdownhydrogenperoxide.Catalaseisanenzymethatbreaksdownhydrogenperoxide,atoxicsubstanceincells,intowaterandoxygen.Therateofreactionofeachsourceofcatalaseatarangeoftemperaturesiscalculatedusingthevolumeofoxygenproducedinoneminute.Thisrateofreactionshowsthepointatwhichtheenzymereachesitsoptimumtemperaturebeforedenaturing. Sourcesofcatalasebeingcomparedare;potato,leek,chickenliver,porkliver,andfishliver.Porkandchickenarehomeothermicanimalsthatdiffergreatlyfromfishliver,apoikilothermicanimal. Theexperimentillustratedafairlyclearrelationshipbetweenthecatalasesourcebodytemperatureoratmospheretemperatureandtheoptimumtemperaturescalculated.Porkandchickenresultedinthehighestoptimumtemperaturesandfishlivercatalasehadthelowestoptimumtemperature.Theoptimumtemperatureforcatalasedoesvarybetweenplantandanimalcellswhentakingintoconsiderationhomeothermicandpoikilothermicanimals.Homeothermicanimalshaveamoredefiniteoptimumtemperatureforcatalasebecausetheirbodytemperatureisconstant.Poikilothermicanimalshaveanindefiniteoptimumtemperatureforcatalasebecauseoftheirvaryingbodytemperatures.However,theplantcatalasewasinconclusive.Thegraphsofmyresultsshowtheexactproductionofoxygenanddepicttheoptimumtemperatureofeachsource.Althoughthisexperimentdidnotresultinanexactoptimumtemperatureforeachsourceofcatalasetested.Wordcount:299

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IntroductionTheresearchquestionforthisessayis: Doestheoptimumtemperaturefortherateofreactionoftheenzymecatalase,asmeasuredbyoxygenproduction,varybetweenplantandanimalcells,whencomparing:leek,potato,chickenliver,porkliverandfishliver;anddoesitmakeadifferenceiftheanimalishomeothermicorpoikilothermic? Myinvestigationwillincludedeterminingtheoptimumtemperatureofcatalasefromexperimentalresultsandresearchusingthesourcesofcatalaselistedabove. Mostchemicalreactionshaveanenergybarrierthatmustbeovercomebeforethereactioncantakeplace.Thisenergyistheenergyofactivation.Catalystsareusedtolowertheactivationenergyneededsothatreactionscanoccurmoreeasily.Enzymesarecatalyststhatarespecifictoonesubstrate.Asubstrateandanenzymeformacomplexthatrequireslessenergyforthereaction.Thiscompoundquicklybreaksdowntoformreactionproducts.(“Enzyme”) Uptoapointtherateofenzymereactionincreasesasthetemperatureincreasesduetothecollisiontheory.Itincreasesuptoacertainpointandthentheenzymeisdenatured,thispointistheoptimumtemperature. Iwillbetestingtheenzymecatalase,asitispresentinalleukaryoticcellsintheperoxisomes.Catalaseisanenzymethatcatalyzesthereactionbywhichhydrogenperoxideisbrokendowntoproducewaterandoxygen.(“Catalase”)Peroxisomesareorganellesboundbyasinglemembranepresentineukaryoticcells.Theyareresponsibleforvariousmetabolicreactionsandthesereactionsproducehydrogenperoxide(H2O2)asabyproduct.Peroxisomesintheliverbreakdowntoxinssuchasalcoholandotherharmfulcompoundsbytransferringhydrogenfromthetoxinstooxygen.“TheH2O2formedbyperoxisomemetabolismisitselftoxic”,(CampbellandReece111)butperoxisomescontaincatalasetobreakdownthistoxicsubstance. Thedecompositionofhydrogenperoxidebycatalasetoproducewaterandoxygenisshownbytheequationbelow.

2H2O2à2H2O+O2("Howdoescatalasebreak")

Ascatalysts,allenzymeshaveaninfluenceontherateofreactions.“Asetofreactantsinthepresenceofanenzymewillformproduct(s)atafasterratethanwithouttheenzyme.”(Damon67)InmyinvestigationIwillbefocusingontheoptimumtemperatureofcatalase.Thismustbetestedbyusingtherateofreactionatdifferenttemperatures.Astemperatureincreases,therateofreactionalsoincreases.Thisoptimumtemperaturecanvarydependingontherolloftheenzymeandthesourceoftheenzyme. Plantsandanimalsallcontaincatalasebutareadaptedtovarioustemperatures.Plantsgrowintheground,whichhasamuchloweraveragetemperaturethanthebodytemperatureofanimals.Leekandpotatoaretwospecies,whicharegrownincoldweatherclimates.

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Iamtestingtwomammalsourcesofcatalase,homeotherms,andonefishsource,poikilothermic.Themaindifferencebetweenpoikilothermsandhomeothermsisthathomeothermshavewarmbloodthatmaintainsasettemperatureandpoikilothermschangedependingonenvironment.Themetabolismsofpoikilothermsareeverchangingandthemetabolismsofhomeothermsareusuallythesame.(Youngbergco2011wp) Theliverofachicken,pigandfishwillbeused,asthisistheorganinanimalsthatcontainsthehighestconcentrationofcatalase.Leekandpotatowillbeusedastheplantspecies. Therateofreactioncanbemeasuredtodetermineatwhichtemperaturethecatalasecarriesoutthereactionmostefficiently.Ratecanbecalculatedby:

(“RatesofReaction”)

Inthedecompositionofhydrogenperoxidebycatalasetheproductsformedareoxygengasandwater.Theproductformedintherateequationwillbethevolumeofoxygenproducedinthereactionwithin60seconds. Myhypothesisisthattheoptimumtemperatureforcatalaserateofreactionwillbehigherinthechickenandpigliverthaninthepotato,leekandfishsourcesofcatalase.Thisisbecausetheenvironmentbywhichthecatalaseisadaptedto,determinestheoptimumtemperatureofthatcatalase.Sincefishisapoikilothermicanimalthemetabolismvariesdependingontheenvironmentandthereforethecatalasewillfunctionatvarioustemperatures.Fishliverwillbelowerthanthemammalcatalaseoptimumtemperaturebecausewatertemperaturesareneverashighasbodytemperature.Chickenandpigarehomeothermicanimalsthatregulatetheirbodytemperaturenaturallyandwillthereforehaveaconstanttemperatureofmetabolicactivity. Ichosethistopicoutofcuriosityforthesubject.AfterconductingseveralenzymelabsintheclassroomIwantedtotakethesefurtherandcreatealargerexperimentandfromthereitgrewintoanExtendedEssaytopic.

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Investigation

VariablesIndependent:Sourceofcatalase–willincludechickenliver,porkliver,fishliver,potatoandleek.Thesesourcesofcatalasemustbepreparedbyblendingandstraining.Dependent:Optimumtemperatureforcatalaseactivity,thiswillbemeasuredbyfindingtherateofreactionforeachtemperature.Thehighestrateofenzymeactivitywillsignifytheoptimumtemperatureforthatsourceofcatalase,measuredbytheproductionofO2.Controlledvariables:Variable Whyitmustbe

controlledHowitwillbecontrolled

VolumeofH2O2andvolumeofcatalasesolution

Ifvolumesfluctuatetheconcentrationsofenzymetosubstrateratiowillbealtered.Thiswillcausetherateofreactiontofluctuate.

ForeachtrialIwillreact2cm3ofhydrogenperoxidewith1cm3ofthecatalasesolution.

Measuringcylinders Theuncertaintiesonmeasuringcylinderscanvary.

Imustusethesamemeasuringcylinderforeachtrialtoensurethattheuncertaintyforthevolumestaysconstant.

Blendingandfilteringmethod

Differentsizedpiecesofthesourcealloweithermoreorlessenzymetoworkonthesubstrate,andaffecttherate.

ForeachofthefourcatalasesolutionsIwillblendandfilterthemwiththesameapparatus.Eachofthesolutionswillhavethesamesurfacearea.

Timethatoxygeniscollected

Thetimeintervalmustbecontrolledforeachtrialbecausethevolumeproducedwillbechangingandthetimewillbekeptconstanttocalculatetherate.

Eachtrialwillbemeasuredafter60secondsandthevolumeofgasproducedwillberecorded.

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Variable Whyitmustbe

controlledHowitwillbecontrolled

Amountofcatalasesourceused

Theamountofwatermixedintothesolutionwilleffecttheconcentrationofcatalaseusedineachtrialanddirectlyaffecttherateofreaction.

Iwillweighoutacontrolledamountoftheliverandplantsourcesbeforeblendingandmixingwithwater.Theamountofwaterwillbecontrolledtoensurethattheconcentrationofcatalaseisequal.

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Materials Belowisalistofapparatusandmaterialsneededfor5trialswithonesourceofcatalase.Allamountsofmaterialwerethesameforeachtest.Material Quantity/VolumeGassyringeanddeliverytube 15cm3measuringcylinders 2100cm3glassbeakers 2Boilingtubeswithdeliverytube 3020volhydrogenperoxide 60mlWaterbaths 4Disposablepipettes 2500cm3glassbeaker 1Boilingtuberacks 2Catalasesolution 30mlCatalasesolutionspreparedandinseparatesealedcontainers 5

MethodMethodPart1 – Preparingthecatalasesolutions

1. Witheachofthelivertypes,add80gofliverwith20mlofdistilledwater.2. Witheachoftheplanttypes,add80goftheplantwith20mlofdistilledwater.3. Blendthewaterandcatalasesourcetogetherwithahandblenderuntilthereare

nolumps4. Strainthefluidwithcheesecloth.5. Putthecatalasesolutionsinplasticsealedcontainersandlabelthem.6. Storethecatalasesolutionsinthefreezer.Theverylowtemperaturesdonot

affectcatalasesolutions.

MethodPart2

1. Pourtheentiremixtureofthefirstsourceofcatalaseintoa100cm3glassbeaker.

2. Placethisbeakerinthefirstwaterbathof30˚Candplaceathermometerandapipetteinthebeaker.

3. Withthesecond100cm3beakerpourapproximately60mlof20volH2O2(hydrogenperoxide)

4. Placethisbeakerinthesamewaterbathasthecatalasesolution(30˚C),andplaceathermometerandapipetteinthisbeaker

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5. Monitorthebeakersinthewaterbathuntilboththehydrogenperoxideandthecatalasesolutionareatthesamecorrecttemperature.

6. Whilethesolutionsaregettingtotemperaturesetuponeboilingtubewithadeliverytubeandapluginaboilingtuberack.

7. Connectthedeliverytubetothegassyringeandensureitisatightseal.8. Presstheendfittingallthewayintotheglassbarreltoemptythegassyringe

readyforthetrial.9. Placeaboilingtubestopperandthestopclockdirectlynexttotheboiling

tube.10. Whenthesolutionsareatthecorrecttemperaturestirthemandthen

measure,usingthepipettesandthetwo5cm3measuringcylinders,1mlofthecatalasesolutionintoonemeasuringcylinderand2mlofH2O2intothesecondmeasuringcylinder.

11. Afterthesolutionshavebeenremovedfromthewaterbaths,transferthemquicklytominimizetemperaturechange.

12. Firstpourthe1cm3ofcatalasesolutionintotheboilingtubeattachedtothegassyringe.

13. Thenaddthe2cm3ofH2O2tothesameboilingtubeandimmediatelyafter,plugtheboilingtubeusingthestopperwithonehandandstartthestopclockwiththeotherhand.

14. Repeatthestepsaboveusingthewaterbathsat10,20,30,40,50,and60˚C,usingafreshsetofequipment.Useiceandcoldwaterinalargeplastictubforthe10˚Cwaterbath.

15. Repeatsteps1-13usingthecatalasesolutionspreparedinmethod1,leek,potato,chickenliver,fishliverandporkliver.Witheachcatalasesourcerepeattheexperiment5timesateachtemperature.Useafreshsetofglasswareforeachtrial.

Eachcatalasesourceateachtemperatureisrepeatedtoallowthecollectionofsufficientandrelevantdata.

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UncertaintiesThereareseveralapparatusuncertaintiesthatmustbetakenintoaccountforthemethodoftheexperiment.Variable Uncertainty(±)Measuringcylinder 0.5mlGassyringe 0.5mlThermometer 1˚CStopclock 2seconds(humanerror)

Results Inthissection,theresultsofmyinvestigationareshowninbothrawdataandprocesseddataforms.Therawdatavalues,volumeofoxygenproducedineachtrial,canbeseeninAppendix:1

ObservationsLeek: Forthissourceofcatalasetherewasagradualincreaseinoxygenproducedthroughoutthe60seconds.Someamountofeffervescencewasformed,verybrightgreencolor.Potato: Afterthesolutionwasfreshlyblendedandfilteredtherewassedimentthatwouldsettleatthebottomofthecontainer.Theliquidpartofthesolutionwasalightbrowncolor.ThepotatoproducedtheleastamountofeffervescenceChickenliver: Thiswasavigorousreaction.Thegasproducedincreasedrapidlyinthefirst10secondsandthenstayedconstantfortheremainderofthetime.Attemperatures,suchas40˚C–60˚Cthegaswouldbegintoretractbyabout1ml.Thiscouldhavebeenduetothechangeintemperatureasthegascooledoff.Atsimilarhightemperaturestheliversolutionwouldbegintosolidify.Porkliver: Thishadamoregradualincreaseofgasproducedthroughoutthe60seconds.Thiscouldbeduetolesserconcentrationofcatalaseinthesolution.Otherobservationswerethesameaschickenliverathighertemperatures.Mackerelliver: Thishadaveryrapidincreaseintheoxygenproduced.Themaximumamountwasreachedinthefirsttensecondsofcombiningthehydrogenperoxideandthecatalasesolution.Effervescencewasshownanditwasayellow/browncolor.Thefishliverwasmuchsmootherandhadfewerclumpsthanthemammalliversourcesofcatalase.

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Processeddata(seeexamplecalculationsinAppendix2.)Tableshowingtheaveragevolumeofoxygenproducedinoneminuteateachtemperatureforeachofthecatalasesources.

Table2.1 AverageVolumeofoxygenproduced(mlmin-1)(±0.5ml)Temperature/˚C(±0.5˚C) PorkLiver

ChickenLiver FishLiver Leek Potato

10 26.8 21.4 29.6 24.0 4.220 28.8 29.6 25.4 38.8 4.230 29.8 30.0 26.2 39.4 8.240 33.0 30.4 25.6 34.8 9.450 32.2 32.8 26.2 13.8 3.060 16.4 27.8 0.40 2.0 1.2

Thetablebelowshowstherateofreactionateachtemperatureforeachsource.Theredhighlightedcellsindicatethehighestratethatwasreached.Thehighestrateiswhatdeterminestheoptimumtemperature.

Table2.2 Rateofreactionofcatalase(cm3sec-1)Temperature/˚C(±1˚C) PorkLiver

ChickenLiver FishLiver Leek Potato

10 0.447 0.357 0.493 0.400 0.07020 0.480 0.493 0.423 0.647 0.07030 0.497 0.500 0.437 0.657 0.13740 0.550 0.507 0.427 0.580 0.15750 0.537 0.547 0.437 0.230 0.05060 0.273 0.463 0.007 0.033 0.020

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Belowisagraphcomparingtheratesofthefourdifferentcatalasesources.Theoptimumtemperaturecanbeobservedwherethehighestpointofthelineis.

0.000

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RateofEnzym

eReaction(cm

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Temperature˚C(±1˚C)

ComparisonoftherateofCatalaseActivityfromDifferentSourcesatDifferentTemperatures

PorkLiver

ChickenLiver

Leek

Potato

FishLiver

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Graphsshowingtheaveragevolumeofoxygenproducedateachtemperatureforeachsourceofcatalase.

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(allgraphsgeneratedbyexcel)

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DiscussionandReasonedArgumentAsstatedbefore,theresearchquestionforthisessayis:Doestheoptimumtemperaturefortherateofreactionoftheenzymecatalase,asmeasuredbyoxygenproduction,varybetweenplantandanimalcells,whencomparing:leek,potato,chickenliver,porkliverandfishliver;anddoesitmakeadifferenceiftheanimalishomeothermicorpoikilothermic?“Uptoapoint,therateofanenzymaticreactionincreaseswithincreasingtemperature,partlybecausesubstratescollidewithactivesitesmorefrequentlywhenthemoleculesmoverapidly.”(CampbellandReece154)Thepointtowhichtherateofanenzymaticreactionincreasesistheoptimumtemperaturefortheenzyme. Thereisapointoneachgraphwheretherateofreactionreachesamaximumpointandthenbeginstodecreasewiththedenaturationoftheenzyme.Denaturationiscausedbytheactivesitechangingshapeduetothehightemperaturesandnotallowingthesubstratetobindwiththeenzymethereforedecreasingtherateofreaction.Thereforethesourcesofcatalasewithaloweroptimumtemperature,theircatalaseactivesitechangesshapemoreeasily.

Therelationshipbetweenrateofreactionofenzymesandtemperatureisshowninthegraphtotheleft.“Chemistshavearuleofthumbthata10°Cincreaseintemperaturegivesadoublingofthereactionrate.ThisruleislooselyderivedfromtheArrheniusequation.”(Oswald)Thisisdepictedinmyresultswhencomparingthegraphsabove.Thepotatoandleekshowthemostnoticeableratecurvesandtheoptimaltemperaturecanbeseenwhentheratedropssignificantlyatacertaintemperature.Thechickenliverand

porklivershowthegeneraltrendoftheincreaseinrate,followingtheArrheniusequation,butthecompletedenaturationoftheenzymecannotbeaseasilyobservedbecauseofthetemperaturesthatwereused.Fishliverhadacompleteoppositecurve,astherateofreactiondecreasesfrom10to20degreesandthenstaysconstantuntilthedenaturationoftheenzymeat50degreesisshown. Fishlivershowedthemostuniquecurvecomparedtotheothersourcesofcatalase.Therehasbeenresearchtoprovethatlivingorganisms“sustainrelativelysimilarratesofmetabolicactivityatwidelydifferenttemperatures”(Somero[abstract]).Thisisduetotheadaptationoracclimationofenzymes,whichhasallowedthemtooperatenormallyatdifferenttemperatureswheretheywould

(Oswald)

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normallybedenaturedifnotadapted.Thisevidencesupportstheideaofpoikilothermic–“theinternalbodytemperaturevariesaccordingtotheambienttemperature”("LearnAboutAnimalAdaptations") andthereforeaffectsthemetabolicrateofthefish.SincetheMackerelfish,whichIusedinmyexperiment,arefoundinawiderangeofdepthsandwatertemperaturesrangingfrom6˚C-20˚C(BigelowandSchroeder)theirpoikilothermiccharacteristicscausetheirmetabolicreactionstostayconstantastheenvironmentaltemperaturefluctuates.Thiscanbeseen,astherateofreactiondoesnotfluctuatebetween20and50degrees.Sincethehighestrateofenzymaticreactionwasat10˚Citwouldbeusefultoconductafurtherexperimentatlowertemperatures.Thefishlivercatalasedenaturedat60˚Candtherewasalmostnooxygenproduced.Thiswastheonlysourcewheretheoxygenproducedbecamealmostzero. Allothersourcesofcatalaseincreasedinrateofreactionastemperatureincreased.Thisisbecauseofthefactoroftemperatureincreasingthekineticenergyofthemolecules,causingmorecollisionsandthereforemoresubstratestobindwiththeenzyme. Chickenhasanaveragebodytemperatureof40.6-41.7˚C.“Birdsarehomeothermic–theyproduceanddissipateheattomaintainarelativelyconstantbodytemperature.Theinternalbodytemperatureofbirdsshowsmorevariabilitythanmammals,andthereforethereisnoabsolutebodytemperature” ("PoultryProductionManual").Thisfluctuationoftemperaturesinbirdsiswhytherateofcatalasereactiondidnotincreaseordecreasebygreatamountswhentemperaturesvaried.Whencomparingthechickenliverandtheporkliver,thechickenliverhadoneincreasefrom10to20˚Candthenstayedmoderatelystableuntildenaturation,withonlyslightincreaseat50˚C;whereastheporkliverhadagradualincreaseinrateofreactionanddidnotstayconstantatanypoint.Thechickenliverdepictsastraighterlineinthevolumeofoxygenproducedandtheporkliverdepictsacurvedlinebeforetheenzymeisdenatured. Pigshaveanaveragebodytemperatureof38.7-40˚C("Temperatureandhumidityindex").Boththeliteraturevaluesforbodytemperatureandmyfindingsofoptimumtemperatureforcatalaseshowthatthechickenhasahighertemperaturethanthepig.Thistrendwasalsoshowninmydatawhencomparingtheoptimumtemperaturesofboththesesourcesofcatalase.Althoughmyinvestigationdoesnotallowforthepreciseoptimumtemperaturetobefoundthegeneraltrendscanbeshownincomparingthedifferentsources. Theleeksourceofcatalasehasthelargestdifferenceinrateafterthedenaturingoftheenzyme.Thelivertypesandpotatoshowedamoregradualchangeinrate.Thistrendonthegraphismostlikelyduetotheamountsofcatalasethatwerepresentinthesolution.TablesinAppendix:1showthatthevolumeofoxygenproducedforanimalspeciesandleekwerethehighestcomparedtotheverylowvolumesofoxygenproducedbythepotato.TodetermineifthisistrueIwouldneedtodeterminetheamountofactiveenzymeinthesample.Thisoxygendifferencecouldbeduetothepotatobeingastorageorgancontainingahighlevelofstarchandleekbeingaleafsourceofcatalase.Theplantsourcesgenerallyhavealoweroptimumtemperaturebecauseplantsdonotproduceabodytemperaturelike

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mammalsdo.Althoughpotatohadthesameoptimumtemperatureasporkliversoplantcatalaseisinconclusive.

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Conclusion FromthegraphsaboveandthecalculationsofaveragesIhavededucedthefollowingoptimumtemperaturesforeachsourceofcatalase. Porklivercatalasehasanoptimumtemperatureof40˚C Chickenlivercatalasehasanoptimumtemperatureof50˚C Fishlivercatalasehasanoptimumtemperatureof10˚C Leekcatalasehasanoptimumtemperatureof30˚C Potatocatalasehasanoptimumtemperatureof40˚CThedatashowsthatanincreaseintemperaturecausesanincreaseinrateofreactioninalltestedsourcesofcatalaseexceptforfishlivercellsbecauseofthesignificantlylowerbodytemperatureandpoikilothermiccharacteristics. Myhypothesiswassupportedtoanextent,bymyfindings.Ipredictedthatthechickenandporklivercatalasewouldhaveahigheroptimumtemperaturethantheplantandfishlivercatalasesandthattheoptimumtemperatureswouldmatchthegeneraltemperatureoftheenvironmentthatthecatalaseisadaptedto.Itisshownbythetrendsonthegraphthattheoptimumtemperatureofchickenliverwassignificantlyhigherthanallothersourcesofcatalaseandtheleekcatalasewasthelowestoptimumtemperature.Theoptimumtemperatureof40˚CforthepotatodidnotsupportmyhypothesisasIexpectedthisvaluetobemuchlower.However,thepotatoproducedthesmallestvolumeofoxygen,whichsuggestsaverylowconcentrationofcatalase. InconclusionIhavefoundthatthesourceofcatalasedoesaffecttheoptimumtemperatureofthecatalaserateofreaction.Previousstudiesshowthat“catalaseactivityoftoadliver(thecoldbloodedanimal)hasitsoptimumat15˚C,butthatofcowliver(thewarmbloodedanimal)at40˚C.”(MitsudaandYasumatsu201) Thisstudyalsotestediftheseoptimumtemperaturesweredependentonenzymepuritybuttheyfoundthattheseoptimumtemperaturesofcatalaseswereindependentofenzymepurity.(MitsudaandYasumatsu200)Both,myfindingsandfindingsinotherresearchshowthattheoptimumtemperaturesaresimilartotheanimal’sbodytemperatureorinthecaseoffish,apoikilothermicanimal,similartoitsatmosphere.Ihavedeterminedthatthereisadifferenceinoptimumtemperaturedependingifthecatalaseanimalsourceishomeothermicorpoikilothermic. Thisinvestigationcouldbetakenfurthercomparingmorespeciesofplantsandanimalsandspecificallydeterminingthereasonforsuchahighoptimumtemperatureforpotatothatwasskewedbytheverylowamountofoxygenproduced.Thecomparisonofmorecold-bloodedanimalscouldimprovethisinvestigation.Theoptimumtemperaturescouldalsobecomemorepreciseifmoretemperatureswithsmallerintervalsweretested.Andtheunknownamountofenzymeinthesamplewasabigproblemindeterminingtheoptimumtemperature.

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Evaluation Therearemanylimitationsanderrorsthathavehadanimpactonmyresultsandconclusions.Thisincludesbothrandomandsystematicerrors. Thepurityandconcentrationofcatalaseisonemajoreffectingfactorbecauseoftheverycleardifferenceintheamountofoxygenproducedwitheachcatalasesource.Inthemammallivertherewasgenerallymoreoxygenproducedthanthepotatoandfishliver.Theleekcellshadaveryhighvolumeofoxygenandthereforecontainedmorecatalasepergram.Thisdoesn’tdramaticallyaffecttheoptimumtemperaturefoundbecauseofastudybyMitsuda,thatenzymepurityisindependentofitsoptimumtemperature.Itismorelikelythatnotallofthecatalasewillbedenaturedathightemperaturesifthesolutionislargerandnotheatedthoroughly.Thiscannotbeimprovedeasily,butcatalasecanbeextractedfromthesourceswithgreatprecision.MyexperimentwouldhaveconcludedmorepreciselyofIhadusedsourcesofcatalasethatwouldreacttoproducesimilarvolumesofoxygen. Theoriginalsourcesandstoragetimeoftheplantsandliverwasunknowninmyexperimentandthiscouldhavecauseddenaturationoftheenzymeassomeoftheliverwasfrozenthawedandrefrozenseveraltimes,whileothersourcesofliverandplantswerefresh.Thiswasnotkeptconstantforeachofthesourcesanditisanunknownfactoraffectingtheprecisionofmyresults.Thiscouldbeimprovedbykeepingallsourcesconsistentandbuyingthemfreshandlocal. Icouldhaveimprovedtheaccuracyofmyresultsandaveragesbyincreasingthenumberoftrials.Icouldhavealsotestedseveraldifferentliversandplantsofthesamespeciestoensurethatitwasnotjustafaultinonesource. Ionlytestedarangeof6temperatures,whichlimitedmyaccuracyindeterminingtheoptimumtemperature.Icouldhaveimprovedmyconclusionbytestingseveralmoretemperaturesbymakingtheintervalssmaller,forexampleintervalsof5˚Cratherthen10˚C. Oneofthegreatesterrorsinmyexperimentwastheunknownconcentrationofenzymebecausemycatalasesolutionswerenotpurifiedorcrystalized.TheconcentrationofcatalaseIwasusingfromeachsourcevariedbecauseoftheothersubstancespresentinthetissues.Itwasclearfromtheresultsthattheliverhadthehighestconcentrationsofcatalase.Thisisdifficulttoimprovewithoutsophisticatedequipment,althoughitispossibletocrystalizecatalaseinapureformandwouldhaveledmetomoreexactoptimumtemperatures. Thisevaluationhashelpedmetodetermineifmyconclusionisreliableornotwhileinvestigatingtheoptimumtemperaturefortherateofreactionoftheenzymecatalasebetweenplantandanimalcells,whencomparing:leek,potato,chickenliver,porkliverandfishliver;andthedifferencebetweenhomeothermicorpoikilothermicanimals.

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<http://www2.ca.uky.edu/poultryprofitability/Production_manual/Chapter7_Venti

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homeotherms/>.

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Appendices

Appendix1

VolumeofoxygenproducedinreactionbetweencatalasefromdifferentcellsandH2O2

LeekTable1.1 Volumemin-1(±0.5ml)Temperature(±1˚C)

Trial1 Trial2 Trial3 Trial4 Trial5

10 26.0 24.0 25.0 22.0 23.020 40.0 37.0 38.0 41.0 38.030 35.0 42.0 38.0 42.0 40.040 33.0 36.0 35.0 36.0 34.050 13.0 8.0 20.0 16.0 12.060 3.0 2.0 2.0 1.0 2.0

PotatoTable1.2 Volumemin-1(±0.5ml)Temperature(±1˚C)

Trial1 Trial2 Trial3 Trial4 Trial5

10 4.0 4.0 5.0 3.0 5.020 4.0 3.0 5.0 4.0 5.030 9.0 8.0 7.0 10.0 7.040 10.0 10.0 8.0 9.0 10.050 3.0 3.0 4.0 2.0 3.060 1.0 1.0 2.0 1.0 1.0

ChickenLiverTable1.3 Volumemin-1(±0.5ml)Temperature(±1˚C)

Trial1 Trial2 Trial3 Trial4 Trial5

10 20.0 24.0 22.0 21.0 20.020 31.0 25.0 29.0 32.0 31.030 30.0 30.0 25.0 33.0 32.040 28.0 29.0 32.0 32.0 31.050 35.0 39.0 36.0 29.0 25.060 30.0 28.0 25.0 30.0 26.0

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PorkLiverTable1.4 Volumemin-1(±0.5ml)Temperature(±1˚C)

Trial1 Trial2 Trial3 Trial4 Trial5

10 27.0 29.0 26.0 24.0 28.020 30.0 28.0 29.0 32.0 25.030 35.0 30.0 25.0 29.0 30.040 31.0 37.0 35.0 32.0 30.050 30.0 34.0 31.0 30.0 36.060 15.0 17.0 20.0 15.0 15.0

MackerelLiverTable1.5 Volumemin-1(±0.5ml)Temperature(±1˚C)

Trial1 Trial2 Trial3 Trial4 Trial5

10 33.0 27.0 27.0 27.0 34.020 28.0 24.0 27.0 24.0 24.030 26.0 27.0 26.0 25.0 27.040 25.0 26.0 28.0 25.0 24.050 29.0 28.0 26.0 25.0 23.060 1.0 0.0 1.0 0.0 0.0

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Appendix2

Calculationforaveragevolumeofoxygenproducedateachtemperature

𝐴𝑣𝑒𝑟𝑎𝑔𝑒 =𝑠𝑢𝑚𝑜𝑓5𝑡𝑟𝑖𝑎𝑙𝑠(𝑚𝑙)

5 Example:

𝐴𝑣𝑒𝑟𝑎𝑔𝑒 =26 + 24 + 25 + 22 + 23𝑚𝑙

5 𝐴𝑣𝑒𝑟𝑎𝑔𝑒 = 24𝑚𝑙ThevaluesfortheaveragegasproducedateachtemperaturewitheachsourceofcatalaseareshowninTable2.1onpage8.

Calculationfortherateofreactionateachtemperatureforeachsourceofcatalase

𝑅𝑎𝑡𝑒 =𝑣𝑜𝑙𝑢𝑚𝑒𝑜𝑓𝑜𝑥𝑦𝑔𝑒𝑛𝑝𝑟𝑜𝑑𝑢𝑐𝑒𝑑

60𝑠 Example:

𝑅𝑎𝑡𝑒 =26.8ml60𝑠

𝑅𝑎𝑡𝑒 = 0.447(mlofoxygen/minute)