grade: grade 9 subject: science unit: chemistry (gr. 9)
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Last Update: April, 2020
Grade:Grade9 Subject:Science Unit:Chemistry(Gr.9)
Title:SeparationExploration–TheScienceoftheSeparationofMixtures
RationaleTherearemanyareasinbiology,biochemistryandchemistrywheremixturesneedtobeseparated.Thescientistmightneedtoseparateonetypeofantibodyfrommanyothers,separatepiecesofDNAfromanotherbysize&charge,orneedtopurifyaproteinforlateruse.Theselaboratorytechniquesrelyontheseparationofmixturesbasedontheirphysicaland/orchemicalproperties,aconcepttaughtinGrade9Science.Later,inGrade11and/orGrade12Biology,theconceptcomesupagainingelelectrophoresis.Theseconceptsarere-visitedandexpandeduponinuniversitychemistrycourses,suchasanalyticalchemistry.Theseanalyticaltechniquesarewidelyusedinindustry.
BackgroundInformationTheWakarchuklabatRyersonUniversity(undertheumbrellaofGlycoNet)isacarbohydrateresearchlabthatusesenzymesfortherapeuticpurposes.Thesepurposesinclude:
• Theuseofenzymestoaddsugarstoproteindrugstomakethesedrugslastlonger(persist)inthebloodstream,therebyreducingdosage,sideeffectsandcost
• Addingsugarmoleculestoculturedneuronalcellsurfacesusingenzymes,sothatthesemodifiedneuronswillfindtheirwaytowheretheyareneededwhengiventoParkinson’spatients
• Researchinghowtomaketheseproteinsusingbacteria(ratherthanmaintainingexpensivemammaliancelllines)
Inthislaboratory,therearemanytechniquesthatareusedtoseparatemixturesand/oranalyzethem.Someoftheseare:
• FPLC(fastproteinliquidchromatography)/HPLC(highperformanceliquidchromatography)
• GelelectrophoresisandSDS-Page• TLC(ThinLayerChromatography)
ForfurtherinformationaboutGlycoNet’sotherresources,visithttps://canadianglycomics.ca/training/high-school-resources/ForfurtherinformationabouttheresearchdoneattheWakarchuklab,visit:https://www.wakarchuklab.com/
Last Update: April, 2020
CurriculumConnectionsGrade9Science:ChemistryUnit:Atoms,ElementsandCompounds
• C1.1assesstheusefulnessofand/orthehazardsassociatedwithcommonelementsorcompoundsintermsoftheirphysicalandchemicalproperties[AI,C]
• C2.2conductaninquirytoidentifythephysicalandchemicalpropertiesofcommonelementsandcompounds(e.g.,magnesiumsulfate,water,carbon,copperII)[PR]
• C2.3planandconductaninquiryintothepropertiesofcommonsubstancesfoundinthelaboratoryorusedineverydaylife(e.g.,starch,tablesalt,wax,toothpaste),anddistinguishthesubstancesbytheirphysicalandchemicalproperties(e.g.,physicalproperties:hardness,conductivity,colour,meltingpoint,solubility,density;chemicalproperties:combustibility,reactionwithwater)[IP,PR,AI]
• C3.4describethecharacteristicphysicalandchemicalpropertiesofcommonelementsandcompounds(e.g.,aluminumisagoodconductorofheat;copperreactstomoistairbydevelopingagreenishsurfaceofcoppercarbonate;sodiumcarbonateisawhite,odourlesspowderthatdissolvesinwater;waterhasuniquephysicalpropertiesthatallowittosupportlife)
LessonObjectives/Concepts
• StudentswilllearnaboutanalyticaltechniquesusedinindustryinaCanadianresearchlaboratorylocatedinToronto,ON(RyersonUniversity)bylisteningtovideoclipsfromCanadianresearchersinthecontextofcarbohydrateresearch
• Studentswilllearnaboutworldissuesandhowsciencecanhelp(waterfiltrationarticle)
• Grade9studentswillbeabletodoapaperchromatographyinquiry,usingatechniquesimilartotheoneusedintheRyersonGlycoNetresearchlab(TLC–thinlayerchromatography)
Time:2periods• 30minutesinthecomputerlab,20-30minforliteracypiece• Inkchromatographyinquiry:1fullperiod
Materials
• Studentsshouldbringtheirownearphonesforuseinthecomputerlab(orlistentothevideosathome)
• Computerfiles:o SeparationExploration-Gr.9StudentResourceo InkChromatographylabreport&rubric
Last Update: April, 2020
Inkchromatography• Permanentmarkers(black,purple,orange,etc.mixedcolours)• Watercolourmarkers(mixedcoloursonly,noblue/yellow/red)• Avarietyofsolvents(water,rubbingalcohol,acetone,othersthatstudentsmaywant
totry)• Avarietyoffilterpaperswithdifferentporesizes• Coffeefilters• 100mlgraduatedcylindersor600mlbeakerwithaglassplateontop• Rulertomeasurethevaluestocalculatetheretentionfactor(Rf)
OverviewPeriod1PartA:ListentotheScientists(20-30minutes)Studentsusedocument,“SeparationExploration–Gr.9StudentResource”toguidethemthroughthevideoclipsontheWeeblywebsitehttp://glyconetchromatography.weebly.com.PartB:LiteracyConnection(20-30minutes)Studentsusesamedocument,“Gr.9SeparationExploration–StudentResource”toguidethemthroughtheliteracypiecehttps://www.scientificamerican.com/article/water-filtration-system/Period2:FullperiodPartC:InkChromatographyInquiryPre-teaching,doneinadvanceofthislesson:
Ø ClassificationofmatterØ Physicalandchemicalpropertiesofmatter
Last Update: April, 2020
Photocredit:http://tinyurl.com/y8xgb86u
Differentsubstancescanbeusedforthechromatographysuchasfilterpaper,papertowelandchalk.Activities/Procedure:Directions:
1. Showstudentstheset-upinthediagrambelow.Askstudentstoselectacolourtoseparateanddottheinkonthefilterpaper,notingthatthesolvent(waterforwatersolublemarkers,rubbingalcoholforpermanentmarkers)muststartoffbelowtheleveloftheink.
Last Update: April, 2020
Photocredit:http://tinyurl.com/yagg9d69
InitiatingandPlanning
• Askstudentswhattheycouldchange(independentvariable)toinfluencetheoutcome(dependentvalue).Someanswerscouldinclude:
Independentvariablestochange
• Filter(usedifferentporesizefilterpapers,whitechalk,coffeefilter,papertowel,artpaper,binderpaper,etc.),
• Solvent(ethanol,water,acetone,vinegar,etc.), • Substancetobeseparated(groundupcolouredfruitsorleaveswouldbeinteresting;
alsospinach,autumnleaves,flowers,beets,etc.).Dependentvariable:measuretheRfvaluesIfthefilterpaperiscutintorectangularstrips,theamountofmovementalongthepapercouldbemeasured(dependentvariable).ItiscalledtheRfvalue(RetentionFactor),which
Last Update: April, 2020
representshowmuchasubstanceissloweddownasitmovesalongthepaper.Itisacomparisonofhowmuchtheinkparticlesmoveinrelationtothesolventfront(howfarthesolventmoved).
𝐑𝐟 =𝐌𝐢𝐠𝐫𝐚𝐭𝐢𝐨𝐧𝐝𝐢𝐬𝐭𝐚𝐧𝐜𝐞𝐢𝐧𝐚𝐬𝐨𝐥𝐯𝐞𝐧𝐭
𝐌𝐢𝐠𝐫𝐚𝐭𝐢𝐨𝐧𝐝𝐢𝐬𝐭𝐚𝐧𝐜𝐞𝐨𝐟𝐭𝐡𝐞𝐬𝐨𝐥𝐯𝐞𝐧𝐭𝐟𝐫𝐨𝐧𝐭
Sometroubleshooting:
1. Makesurethatyouchoosecoloursofmarkersthataremixturesofothercolours(purple=red+blue,orange=yellow+red).Primarycolours(red,blue,yellow)willnotseparate,astheyarenotmixtures.
2. Makesurethatthechromatographypaperisnotinthesolventverydeeplyoreverythingwillsmear.
PerformingandRecordingHavestudentstryitout,keepingonetreatmentthesame(thecontrol)andchangesomethingelse(changeonevariableatatime).Laboratoryreport:canbeintheformofanarrativelabreport;seethedocument“InkChromatographyLaboratoryReportandRubric”.
SummaryAsaclass,studentslistentoavideo“TheGlycoNetStory”togetthecontextbehindcarbohydrateresearchandwhyitissoimportantforhumanhealth(therapeuticproteins,biofilms,etc.)andenergyneeds(biofuels).Individually,theylistentoandanswerquestionsaboutdifferentresearcherswhotalkaboutthebiochemicalanalysisthattheydo.InGrade9,thereisaconceptabouttheseparationofmixturesthatcanbeseeninitsappliedcontextinanalyticalchemistryandbiologyintheworkplace.Techniquessuchasgelelectrophoresis,TLC(thinlayerchromatography)andSPE(solidphaseextraction)arehighlighted.Afterthisintroduction,studentsreadaboutanamazinginventionthatfilterswaterfromastraw.Thenextperiod,studentscontinuewiththisthemebydoingapaperchromatographyinvestigationwithdifferentcolouredinks.TeacherscouldchoosetofollowthisconceptintotheEcosystemsUnitanddoaWaterFiltrationInquiry(ifstudentshavenotdoneoneinearliergrades).
Assessment:Usetherubricinthefile,“Inkchromatographylaboratoryreportandrubric”
Extensions
1. Thestudentscouldperformacandychromatographyexperimentinwhichcandies,suchasgobstoppers,arerestinginthemiddleofafilterpaperandwaterisaddedby
Last Update: April, 2020
dropper(itactsasthesolvent).Thecolouredparticlesdissolveandmoveapartfromeachother,dependingonsize.Thiscouldbedoneasaquickinquiry–canchangethetypeoffilterpaper(coffeefiltervs.filterpaperordifferentporesize),changethesolvent(differenttypesofsolution–acid,base,neutral,saltsolution,etc.).2.DNADaywithCurioCity–DNAandFoodSecurity:http://curiocity.archive.letstalkscience.ca/Explore/ArticleId/5321/dna-day-2017.aspx3.UsethesamepartA(videoclips)andPartB(literacyconnection)butdoawaterpurificationinquiryandconnectto11UChemistry(SolutionsandSolubility)withconnectionstothemanyexpectationsaroundwaterpurification,treatmentanddistributionaroundtheworld.4.TeachersmaychoosetocontinuewiththisconceptanddoaWaterFiltrationInquiryduringtheGrade9EcosystemsUnit.ThismayconnecttosomeGrade9Geographyexpectationsaswell.5.AnotherextensionincludesaSTEM(Science,Technology,EngineeringandMathematics)quantitativeactivityinwhichwatercontaminatedwithchlorineispassedthroughafilterandthewateristestedwithchlorineteststripsafterwards.https://www.teachengineering.org/activities/view/wst_environmental_lesson02_activity2
References
• NakitaBeunbrazo,July2017.PresentationforTDSB(Powerpointpresentation).Wakarachuklaboratory,RyersonUniversity.
• Dr.WarrenWakarchuk,July2017.PresentationforTDSB(Powerpointpresentation).Wakarachuklaboratory,RyersonUniversity.
Last Update: April, 2020
NarrativeLaboratoryReport1Rationale:Theideabehindanarrativelaboratoryreportistomovestudentsbeyondaquestionandanswerstyleofanalysisorevenaformallaboratoryreport.Theyhavetoaskthemselvesthecorequestions:WhatdidIlookfor?HowdidIlookforit?WhatdidIfind?Whatdiditmean?Itishopedthatbyaskingstudentstostepbackandtakeawiderviewoftheirwork,theywillmovetowardsopeninquirywheretheybegintowonderandgeneratetheirownquestionstoanswer(andbeingcomfortablewiththat).Aftertheinitialuncertaintyaboutmarksandsteppingoutoftheircomfortzone,studentsgenerallylikethefreedomtoworkinthisway,ifthetopiccaptivatesthem.Itistheresponsibilityoftheteachertoselectopportunitiesforthestudentsthathaveaninterestingcontextorsomeaspectthatisfascinating.WhatwasIlookingfor?(Describeyourresearchquestionherealongwithyourhypothesis)
• Shouldbe1-2sentences,includesatestablequestion&hypothesisHowdidIlookforit?(Describeyourmethod)
• Shouldbereproducibleandstep-by-stepWhatdidIfind?(Showanyobservationsthatyouhad)
• Shouldincludetables(succinct&organizedwithheadings)andgraphs(labelaxes,includetitle,legendifnecessary)
Whatdoesthismean?(AnalysisandConclusion)• Shouldincludeanalysisofscientificobservationsanddata;shoulddiscuss
resultsandwhytheyoccurred• Writteninparagraphform
1ModifiedfromLenapeDistrictHighSchool’sNarrativeLabReport:https://www.lrhsd.org/cms/lib/NJ01000316/Centricity/Domain/26/templatepercent20narrativepercent20lab.pdf
Last Update: April, 2020
RubricforNarrativeReport Level1 Level2 Level3 Level4WhatwasIlookingfor?
Researchquestiondidnotincludeaproperlytestablequestion
Researchquestionwassomewhatfocusedwithatestablequestion
Researchquestionwaswellfocusedwithatestablequestion
Researchproblem/questionwashighlyfocusedwithatestablequestion
HowdidIlookforit?
Methodwasincomplete
Methodwasmostlythere
Methodwaswritteninagenerallyreproducibleway
Methodwaswritteninadetailedandhighlyreproducibleway
WhatdidIfind?
Observationswerepresentbutwerenotorganized/succinct;graphslackingaxes,labels&title
Observationsweresomewhatorganized&succinctwithtableheadings,goodgraphs
Observationswerewellorganized&succinctwithtableheadings,excellentgraphs
Observationswerehighlyorganized&succinctwithtableheadings,excellentgraphs
Whatdoesthismean?
Analysisdidnotfocusonthekeyquestionanddidnotaddressscientificerror
Analysissomewhatcapturedthekeyquestionsbutdidnotaddressthescientificerrorwell
Analysisgenerallycapturedthekeyquestionsandaddressedmostofthescientificerror
Analysiscapturedthekeyquestionsandworkedinscientificerrorinahighlevelway
Last Update: April, 2020
AnswerKeyPartA:ListentotheScientistsTheGlycoNetStoryvideo:https://www.youtube.com/watch?v=CQGEGIoqTpQ
Letthevideoplay.Stopatthe1minutemark.1. Whatistheimportanceofunderstandingglycomics(thestudyofsugars)?
Communicationbycellsorinvasionintocellsisoftenmediatedbysugars.Ifweunderstandglycomics,wecaninterferewithinfectionprocesses.
2. WhatisGlycoNet?
GlycoNetisapan-Canadiannetworkofresearchersthatformedin2015todevelopsolutionstounmethealthneeds.
3. Whybringtheseresearcherstogether?Thispoolingofeffortsshouldresultinastreamlinedprocesstogetproductstomarketaswellascompetemoreeffectivelyontheinternationalstage.
Playvideofrom1min–2:474. WhatkindofgoodcanresultfromtheseGlycoNetpartnerships?
Anythingrangingfromnewvaccinestonewmedicinesfordiseasesthatarecurrentlyuntreatable.
5. Whatareashavetheymadeprogressin?Influenzaanddiabetes
MeettheGlycoNetScientistsfromRyersonUniversity,WakarchukLaboratory.Gotowww.glyconetchromatography.weebly.comVideoclip#1:Tasnim1–GraduateStudent
1. WhatisTasnim’sareaofresearch?Herareaofresearchisenzymaticsynthesis.Sheattachescertaincomponentsontoproteins,carbohydratesorotherbiomoleculesthatthelabneeds.
2. WhywouldTasnimfluorescentlylabel/tagmolecules?Researchthis.Itisnotmentionedinthevideobutwhenyouaretryingtopurifyamolecule,itisbeneficialtobeabletoseethedesiredproduct.Inthiscase,sheisvisualizingthemoleculeswithafluorescentdye.
Last Update: April, 2020
Videoclip#2:Tasnim2–AtypicaldayforTasnimAbukar
1. TasnimmentionsthatsheusesTLC(ThinLayerChromatography)onatypicalday.WhatisTLC?Whatisitusedfor?TLCisusedtoseparatemixturesintoindividualcomponents.Withpaperchromatography,aninkcanbeseparatedintotheparticlesthatcomposeit.Forexample,ifpurpleinkweredottedontoapieceoffilterpaperanddippedintowateroranothersolventsuchasalcoholoracetone,thesolventwouldmoveupthepaperthroughcapillaryactionandcarrytheinkparticleswithit.Sincetheinkparticlesdifferinsomeway(size,charge,attractiontothepaper,etc.),theymovedifferentamountsandcanbeseenseparately.Formoreinformation,refertohttp://chemguide.co.uk/analysis/chromatography/thinlayer.html.
2. WhatdidTasnimsaywasthebasisoftheseparationintheTLC?
Shesaidthatthemoleculeswouldseparatebasedonpolarity.3. TasnimmentionedthatanothertechniquethatsheusesalotisFPLC(fastproteinliquid
chromatography).Whatisthis?Chromatographyinvolvestheseparationofmixturesintotheirindividualcomponentsbasedondifferentpropertiesofthecomponents.ItisusedalotinbiotechnologyandissimilartoHPLC(highperformanceliquidchromatography)butatahighpressure.Theseparationcouldbeduetosize,chargeorhydrophobicinteraction.Reference:http://fplc.weebly.com/background.html
4. Whatiselution?Inproteincolumnchromatography,asolutioncontainingthedesiredprotein(plusimpurities)ispouredthroughthecolumn.Theproteinwillsticktothematerialinsideofthecolumnandtheimpuritieswillflowthrough.Later,anothersolutionwillbepouredthroughtoreleasethedesiredprotein,whichisthencollectedinpureformforfurtheruse.Thisprocessof“stripping”thecolumniscalledelutionandiscommonlyusedinbiotechnologyandbiochemistry.Reference:https://en.wikipedia.org/wiki/Elution
Videoclip#3:RayMartinez-Rodriguez-Summerstudent(1:10minutes)
1. WhatdoestheenzymethatJoseworkswithdo?
Itmovessugarsfromadonormoleculetoanacceptormolecule.2. Whataretwoproblemswiththisprotein(enzyme)?
Oneproblemisthatitishardtopurifyandtheotheristhatitisnotveryactive.
Last Update: April, 2020
3. JosedescribeshowaHistagcanhelpfixoneofthetwoproblemsmentionedabove.
Whatdoesithelpwith?DoanInternetsearchforHis-tag(histidinetag)purificationandIMAC(immobilizedmetalaffinitycolumn).Hementionsusinganickelcolumn.DrawapictureofanIMACcolumnandexplain.Scientistsaddhistidinetoproteinstohelptotagthemforapurificationprocess.Whentryingtoscaleupasubstanceforlargescaleproduction,itisimportanttobeabletoidentifyandpurifythefinalproduct.Thishelpswiththepurificationprocess.Seequestion#4fortherestoftheanswer.
4. Immobilizedlactaseinacalciumalginategelandpouringmilkthroughittobreakdown
thelactaseisasimilarideatotheIMACmentionedabove.Howarethesetwoproceduressimilarandhowaretheydifferent?TherepeatingHis(histidine)taghasacertainaffinityformetalssuchasnickelwhichareembeddedinthecolumn.Asamplethatishistidine-taggedcanbepouredthroughtheNicolumn,stickingtothecolumnwhileimpuritiesareflushedout.Thehis-taggedsamplecanbelaterrecoveredbyelutingwithanothersubstancesuchasimidazole,whichcompeteswithhistidineforthenickel.Inthisway,arecombinantproteincanberecovered.Thisisdifferentfromusingimmobilizedlactasesincelactosepassesthroughandtheenzymebreaksitdown.Noelutingisnecessarysincethegoalistogetridoftheoffendinglactosebybreakingitdown.Thebreakdownproductsarestillpresentinthelactose-freemilk.Thetwoprocessesaredifferentbecausethegoalsaredifferent.IMACusesaffinitytoremoveacertainmoleculefromothermoleculesinthelysate.Immobilizedlactasecarriesoutahydrolysisreactionononecomponentofthemixtureandthenreleasestheproductsbackintosolution.Thetwoprocessesaresimilarbecauseinbothcases,amixtureofsubstancesispouredthroughthecolumn.WithIMAC,onecomponentisremoved.Withimmobilizedlactase,lactoseisremovedbybreakingitdownintoothermolecules.Reference:“HowdoesHis-tagpurificationwork?”http://www.bio-rad.com/featured/en/his-tag-purification.html
Videoclip#4:Dr.TingDu-PostDoctoralFellow(26seconds)
1. Dr.Dutalksaboutpurifyinghertargetprotein.Howdoesshedothat?ShedoesthiswithFPLC(fastproteinliquidchromatograph)orAKTA(aproteinpurificationsystem).
Last Update: April, 2020
2. Shealsotalksaboutconstructinganoperon.Whatisanoperon?
Anoperonisaclusterofgenesthatcodesforaparticularproteinthatisregulatedbyasinglepromoter.
3. Whatisthefunctionoftheproteinsintheoperonthatsheistalkingabout?
TheseenzymesputanO-glycan(chainofcarbs)ontohertargetprotein.References:1.http://fplc.weebly.com/uses.html.2.https://www.britannica.com/science/operon
Videoclip#5:LauraKell,ResearchTechnician(58s)AsLauratalksaboutherjob,shementionstwomainresponsibilities.Oneofthemistrainingandmentoringotherteammembersandassistingthemtopushtheprojectsalong.
1. Isitsurprisingtoyouthataresearchtechnicianhasthisresponsibilityaspartoftheirjob?Answerswillvary.
2. Whatistheotherpartofherjob?
HerresearchincludesusinglabtechniquessuchasproteinpurificationusingFPLC(aformofliquidchromatographytoanalyzeandpurifyproteins).Shealsousesenzymeassayswithfluorescentlylabeledsubstrate(amoleculethatisacteduponbyanenzymetoproduceaproduct)whicharereactedwithenzymeswhicharethendevelopedonaTLC(thinlayerchromatography)plate.
3. Lauradescribeshertypicaldayandsays,“Well,thereisnotypicalday.”Doesthisappeal
toyou?Wouldyoulikeajobwheretherearedifferentthingshappeningday-to-dayorsomethingmoreconsistent?Answerswillvary.
4. Whatisanenzymeandsubstrate?Researchthis.
Anenzymeisaproteinthathelpstobreakdownorbuildupothermolecules.Asubstrateisamoleculethatanenzymeactsupontocreateaproduct.Anexampleistheenzymelactase,whichhelpstobreakdownthedoublesugarlactose(asugarfoundinmilk)intosinglesugars.
Clickonthe“LabTechniques”Tabatthetopofthewebsitepage.YouwillseesomeoftheRyersonUniversityscientistsatwork,demonstratinglabtechniquessuchas:gelelectrophoresis,TLCandSPE(solidphaseextraction).
ViewtheTLCdemonstrationvideo.Summarizetheconceptin1-2sentences.
Last Update: April, 2020
Summary:TLCworksbyseparatingamixtureintoitsindividualcomponentsbytheirproperties.
PartB:LiteracyConnectionReadthefollowingbriefScientificAmericanarticleprovidedbyyourteacher:https://www.scientificamerican.com/article/water-filtration-system/OntheLines:MainIdea,Who?Whathappened?Include2definitionsofkeywordsAnswerswillvary.BetweentheLines:Why?Benefits?Consequences?Answerswillvary.
BeyondtheLines:Doyouagree?Howdoesthisaffectyourcommunity/city/country?Answerswillvary.
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