grade: grade 9 subject: science unit: chemistry (gr. 9)

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/


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


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


Photocredit:http://tinyurl.com/y8xgb86u

Differentsubstancescanbeusedforthechromatographysuchasfilterpaper,papertowelandchalk.Activities/Procedure:Directions:

1. Showstudentstheset-upinthediagrambelow.Askstudentstoselectacolourtoseparateanddottheinkonthefilterpaper,notingthatthesolvent(waterforwatersolublemarkers,rubbingalcoholforpermanentmarkers)muststartoffbelowtheleveloftheink.


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


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


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.


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


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


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.


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.


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).


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.


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.

grade: grade 9 subject: science unit: chemistry (gr. 9)

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