Lab # 4: Respiration
Lab4:Respiration
Conceptscovered
9CellularEnergy
9Respiration
9AnaerobicRespiration(doesnotrequireoxygen)
9AerobicRespiration(requiresoxygen)
ATPistheenergycurrencyofthecell.Itisproducedthroughaprocesscalledrespiration.
Theenergymolecules(ATP)generatedthroughrespiration,areavailabletofueltheprocessesofthe
cellasneeded.WhenATPlevelsbecometoolowaspecialproteinsignalsthecelltobeginrespiration.
Aslongasallthecriticalcomponentsforthereactionareavailable,thiscycleprovidesaconstant
sourceofenergyforthecell.
Respirationharvestsbiologicalenergyfromfuelmolecules,suchascarbohydrates,andstoresitasATP.
Togetherwithoxygen,thecellconvertscarbohydratestocarbondioxide,waterandenergy.Asshown
intheequationbelow,respirationisacontrolled,multistepprocesswhichslowlyreleasestheenergy
storedinglucoseandconvertsittoATP.Ifallofthisenergyfromglucosewerereleasedatonce,most
wouldbelostasheatandlight.
Carbohydratescontainhighenergybondsthat,whenbroken,releaseelectrons.Thefirststageofres
piration,glycolysis,breakscarbohydrates(glucose)intopyruvatemolecules.Thoughthebondshold
ingpyruvatetogethercontainagreatdealofpotentialenergy,thisstepyieldslittleenergy.
Glycolysisoccurswithorwithoutoxygenandtakesplaceinthecytoplasmoutsidethemitochondria.
Interestingly,itisapathwayfoundinalllivingthings.
C6H12O6+6O26CO2+6H2O+energy
glucoseoxygencarbondioxidewater
Yeasthasbeenusedtomakeleavenedbread
forcenturies.Whenyeastundergoesfermen
tation,CO2istrappedandcausesthebreadto
rise.Ethanol,anotherbyproductofyeast
fermentation,generatesthealcoholcontent
inbeer,andtheCO2provideseffervescence.
Whatingredientsmustbepresentinorderfor
thisprocesstooccur?
Lab # 4: Respiration
Thesecondstage,calledanoxidationreactionortheKrebsCycle,isalwayscoupledwithareduction
reaction.Bothinvolvethemanipulationofelectrons.Theprocessusedatthispointinrespiration
dependsonthepresenceofoxygen(aerobicrespiration)orwithoutoxygen(anaerobicrespiration).
Membrane
Anaerobic
Aerobic
2 ATP
in mitochondria
6 Carbon Compound
Glucose
4 ADP
Oxidation
4 ATP
3 Carbon Compound
2 ATP
Alcohol +
CO2
Cytosol
+O2
Pyruvate
Yeast fermentation
CO2 + H2O
Homolactic
fermentation
Lactic
Acid
Aerobicrespirationtakesplaceinthemitochondria(a
specializedorganelle)ofthecellandusesoxygenas
thefinalelectrontransporter.Inthisprocess,pyruvate
isoxidizedtogenerateenergy.Thekeytoefficiencyin
thisstepistheuseofoxygentoshuttleelectrons.
34 ADP
34 ATP
Mitochondrion
However,iftheelectronsreleasedweretransferred
directlytooxygen,itwouldcombust.Toavoidthis,
specialmoleculesshuttleelectronstotheATP
productionsite.Sinceoxygenhasaveryhighaffinity
forelectrons,aerobicrespirationisthemostefficient
meansofproducingATP(36perreaction).
Figure4.1:AerobicRespiration
Anaerobicrespirationtakesplaceinthecytoplasmofthecellandusesother,lessefficient,molecules
totransportelectrons.Ifthefinaltransfermoleculeisorganic(containsacarbonatom),theprocessis
calledfermentation.Fermentationisananaerobicprocessthatusesenzymestoreducepyruvateinto
energyrichmolecules.Becauseitcannotfullybreakdowntheglucosemolecule,fermentationisfar
lessefficientthanaerobicrespiration,generatingonlytwoATPmolecules.
Duringphysicalactivity,cellsrequiremore
energy.Aslongasenoughoxygencanbe
deliveredtocells,aerobicrespiration
dominates.
Whenenergyconsumptionexceedsthe
oxygensupply,anaerobicrespirationstarts.
Lacticacidisabyproduct,andiswhatcauses
musclesorenessafterahardworkout!
Lab # 4: Respiration
Experiment4.1:FermentationinYeast
YeastcellsproduceethanolandCO2duringfermentation.WewillmeasuretheproductionofCO2to
determinetherateofanaerobicrespirationinthepresenceofdifferentcarbohydrates.
Wewillbeusingtwodifferenttypesofsugarsinthislab:
x Sucrose(adisaccharide)ismadeupofglucoseandfructose
x Glucoseisamonosaccharide
Defineforthisexperiment:
Hypothesis:
Control(s):
IndependentVariable(s):
DependentVariable(s):
Materials
5Respirometers:(twotesttubesthatfitintoeachother)–smallplasticandlargeglass
1%solutionsofglucose,sucrose
Equal,SplendaandSugarPackets
Yeast
(4)250mlbeaker
*Warmwater
Pipettes
*Watchortimer
Permanentmarker
Ruler
MeasuringSpoon
*Youmustprovide
Lab # 4: Respiration
Procedure
1. Completelyfillthesmallesttubewithwaterandinvertthelargertubeoverit.Pushthesmall
tubeup(intothelargertube)untilthetopconnectswiththebottomoftheinvertedtube
(Figure4.2).Inverttherespirometersothatthelargertubeisupright(thereshouldbeasmall
bubbleatthetopoftheinternaltube).Repeatthisseveraltimesaspractice–striveforthe
smallestbubblepossible.Whenyoufeelcomfortablewiththistechnique,emptythetesttube
andcontinuewiththisexperiment.
Figure4.2:Respirometer
2. Mix1/4teaspoonofyeastinto175mLofwarm(4043°C)waterina250mLbeaker.Stiruntil
dissolved.
Note:Makesuretheyeastsolutionisstirredbeforeeachtesttubeisfilled.
3. Labelboththebigandsmalltesttubes15withamarker.
4. Ina250mlbeaker,mixthepacketofEqualwith100mlofwater.Inanother250mlbeaker,mix
thepacketofSplendawith100mlofwater.Inanother250mlbeaker,mixthepacketofsugar
with200mlofwater.Thesearenow1%solutions.
5. Fillthesmallertubeswith15mLsolutionasfollows:
Tube1:1%glucosesolution
Tube2:1%sucrosesolution
Tube3:1%Equalsolution
Tube4:1%Splendasolution
Tube5:1%sugarsolution
Note:Makesureyourinsethegraduatedcylinderbetweeneachuse.
6. Then,filleachtubetothetopwiththeyeastsolution.
7. Slidethecorrespondinglargertubeoverthesmalltubeandinvertitaspracticed.Thiswillmix
theyeastandsugarsolutions.
Lab # 4: Respiration
8. Placerespirometersinthetesttuberackandmeasuretheinitialairspaceintherounded
bottomoftheinternaltube.RecordthesevaluesintheTable4.1.
9. Allowthetesttubestositinawarmplace(~37C)foronehour.Thismaybeasunny
windowsill,atopawarmoven,underaverybright(warm)light,etc.
10. Attheendoftherespirationperiod,measuretheairspaceintheinternaltubes,andrecordit
inTable4.1.
Table4.1:RecordingRespirometerValues
Tube
Initialgasheight(mm)
Finalgasheight(mm)
NetChange
1
2
3
4
5
Questions
1. Didyounoticeadifferenceintherateofrespirationbetweenthevarioussugars?Didthe
“fake”sugarprovideagoodstartingmaterialforfermentation?
2. Howdoyouknowthatanaerobicfermentationwasoccurring?
Experiment4.2:AerobicRespirationinSoybeans
Wewillevaluaterespirationinbeansbycomparingcarbondioxideproductionbetweengerminated
andnongerminatedbeans.Asshowninthebalancedequationforcellularrespiration,oneoftheby
productsofrespirationisCO2(carbondioxide):
C6H12O6+6H2O+6O2energy+6CO2+12H2O
Lab # 4: Respiration
Wewilluseacarbondioxideindicator(bromothymolblue)toshowoxygenisbeingconsumedandcar
bondioxideisbeingreleasedbythebeans.
Bromothymolblueisanindicatorthatturnsyellowinacidicconditions,greenwhenitisneutraland
bluewhenitisinbasicconditions.
Defineforthisexperiment:
Hypothesis:
Control(s):
IndependentVariable(s):
DependentVariable(s):
Materials
100beans
(3)250mlbeakers
*Papertowels
3measuringcups
Pipette
Bromothymolbluesolution
Parafilm
3Rubberbands
*Water
*Youmustprovide
Figure4.3:BeakerSetup
Procedure
1. Soak50beansfor24hoursin200mlwater.
2. Labelthreebeakers:Beaker1,Beaker2,andBeaker3.
3. Placeseverallayersofmoistpapertowelsatthebottomofthe250mlbeakers.
4. Place50presoakedbeansintoBeaker1,50control(dry)beansinBeaker2,andzerobeansin
Beaker3.
5. Dispense4mlofbromothymolbluesolutionintothebottomofeachmeasuringcup,andplace
onemeasuringcupinsideeachbeakerontopofthebeans.
6. StretchtheParafilmacrossthetopofeachbeaker.Securewitharubberbandtocreateanair
tightseal(Figure4.3).
Lab # 4: Respiration
7. Placethebeakersonashelfortable,andletsitundisturbedatroomtemperature.
8. Observethejarsat10minuteintervalsforthreehours,andrecordanycolorchangeofthe
bromothymolblueinTable4.2.
Table4.2:Colorchangesofthebromothymolblueovertime
Time
Jarwithpresoakedbeans
Jarwithunsoakedbeans
Jarwithnobeans
0min
30min
60min
90min
120min
150min
180min
Questions
1. Howdidthecolorofthebromothymolbluesolutionineachbeakerchangeovertime?
2. Whatcanbeinferredfromthecolorchangeofthebromothymolbluesolution?
3. Whatisthemechanismdrivingthebromothymolbluesolutioncolorchange?
4. Whatarethecontrolsinthisexperiment,andwhatvariablesdotheyeliminate?Whyisit
importanttohaveacontrolforthisexperiment?