GCSE Science - Chromatography
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This page has been archived and is no longer updated. Find out more about page archiving. GCSE Bitesize BBC Radio 1 BBC 1Xtra Science Chromatography Chromatography is an important analytical technique because it allows chemists to separate substances in complex mixtures. There are a variety of types of chromatography, which can be used in different contexts. Analytical procedures Qualitative analysis means any method of identifying the chemicals in a sample. Quantitative analysis means working out how much of a particular chemical there is in a sample. Any analysis must be carried out on a sample of a substance that is representative of the bulk of the substance. This means that the results from the analysis can be used to draw conclusions about the rest of the substance. It is important to mix solutions thoroughly before testing, and to take many samples at random from the solution when taking repeat readings. It is important to follow standard procedures when analysing chemicals. This ensures that the results are reliable and valid. Standard procedures cover the following: collecting samples storing samples preparing samples for analysis Paper chromatography In chromatography, substances are separated as they travel in a mobile phase which passes through a stationary phase. Different substances travel at different speeds, so some move further than others in a given time. In paper chromatography, the stationary phase is paper. The mobile phase may either be an aqueous (waterbased) liquid or a nonaqueous organic (carbon based) solvent [solvent: A solvent is the liquid in which the solute dissolves to form a solution.] . An example of an organic solvent is propanone which is the main chemical in nail varnish remover. For each chemical in the sample, there is a dynamic equilibrium [dynamic equilibrium: When the rate of a forward reaction is the same as the rate of the reverse reaction, so the concentration of product remains constant over time.] between the stationary phase and the mobile phase. The overall separation depends upon how strongly attracted the chemicals are to the mobile and the stationary phases. Watch this video clip to see how paper chromatography can be used to solve a crime. In order to see this content you need to have both Javascript enabled and Flash installed. Thin layer chromatography (TLC) Thin layer chromatography (TLC) is similar to paper chromatography but instead of paper, the stationary phase is a thin layer of an inert [inert: Unreactive.] substance (eg silica) supported on a flat, unreactive surface (eg a glass plate). TLC has some advantages over paper chromatography. For example: the mobile phase moves more quickly through the stationary phase the mobile phase moves more evenly through the stationary phase there is a range of absorbencies for the stationary phase TLC tends to produce more useful chromatograms [chromatogram: To separate different substances dissolved in a liquid.] than paper chromatography, which show greater separation of the components in the mixture and are therefore easier to analyse. Analysing chromatograms A chromatogram [chromatogram: To separate different substances dissolved in a liquid.] can be compared with a chromatogram of a standard reference material to identify which chemicals the mixture contained. The mixture on the left separated into three substances which match the pure standard reference materials used on the right Locating agents Sometimes the substances being separated are colourless. In this case, locating agents can be used to show where the spots are. Locating agents bind to the chemicals in the spots. Sometimes, another chemical is then added, which reacts with the locating agent to produce a coloured spot, or the chromatogram is put under ultraviolet [ultraviolet (UV): Electromagnetic waves with frequencies higher than those of visible light, beyond the violet end of the visible spectrum. UV light causes damage to DNA leading to sunburn and possible future skin cancer.] light and the locating agent glows to show where the spots are. R f value The movement of a substance during chromatography, relative to the movement of the solvent [solvent: A solvent is the liquid in which the solute dissolves to form a solution.] , is measured by calculating its retardation factor (R f ).
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GCSEBitesizeBBCRadio1BBC1XtraScienceChromatographyChromatographyisanimportantanalyticaltechniquebecauseitallowschemiststoseparatesubstancesincomplexmixtures.Thereareavarietyoftypesofchromatography,whichcanbeusedindifferentcontexts.AnalyticalproceduresQualitativeanalysismeansanymethodofidentifyingthechemicalsinasample.Quantitativeanalysismeansworkingouthowmuchofaparticularchemicalthereisinasample.Anyanalysismustbecarriedoutonasampleofasubstancethatisrepresentativeofthebulkofthesubstance.Thismeansthattheresultsfromtheanalysiscanbeusedtodrawconclusionsabouttherestofthesubstance.Itisimportanttomixsolutionsthoroughlybeforetesting,andtotakemanysamplesatrandomfromthesolutionwhentakingrepeatreadings.Itisimportanttofollowstandardprocedureswhenanalysingchemicals.Thisensuresthattheresultsarereliableandvalid.Standardprocedurescoverthefollowing:collectingsamplesstoringsamplespreparingsamplesforanalysisPaperchromatographyInchromatography,substancesareseparatedastheytravelinamobilephasewhichpassesthroughastationaryphase.Differentsubstancestravelatdifferentspeeds,sosomemovefurtherthanothersinagiventime.Inpaperchromatography,thestationaryphaseispaper.Themobilephasemayeitherbeanaqueous(waterbased)liquidoranonaqueousorganic(carbonbased)solvent[solvent:Asolventistheliquidinwhichthesolutedissolvestoformasolution.].Anexampleofanorganicsolventispropanonewhichisthemainchemicalinnailvarnishremover.Foreachchemicalinthesample,thereisadynamicequilibrium[dynamicequilibrium:Whentherateofaforwardreactionisthesameastherateofthereversereaction,sotheconcentrationofproductremainsconstantovertime.]betweenthestationaryphaseandthemobilephase.Theoverallseparationdependsuponhowstronglyattractedthechemicalsaretothemobileandthestationaryphases.Watchthisvideocliptoseehowpaperchromatographycanbeusedtosolveacrime.InordertoseethiscontentyouneedtohavebothJavascriptenabledandFlashinstalled.Thinlayerchromatography(TLC)Thinlayerchromatography(TLC)issimilartopaperchromatographybutinsteadofpaper,thestationaryphaseisathinlayerofaninert[inert:Unreactive.]substance(egsilica)supportedonaflat,unreactivesurface(egaglassplate).TLChassomeadvantagesoverpaperchromatography.Forexample:themobilephasemovesmorequicklythroughthestationaryphasethemobilephasemovesmoreevenlythroughthestationaryphasethereisarangeofabsorbenciesforthestationaryphaseTLCtendstoproducemoreusefulchromatograms[chromatogram:Toseparatedifferentsubstancesdissolvedinaliquid.]thanpaperchromatography,whichshowgreaterseparationofthecomponentsinthemixtureandarethereforeeasiertoanalyse.AnalysingchromatogramsAchromatogram[chromatogram:Toseparatedifferentsubstancesdissolvedinaliquid.]canbecomparedwithachromatogramofastandardreferencematerialtoidentifywhichchemicalsthemixturecontained.
ThemixtureontheleftseparatedintothreesubstanceswhichmatchthepurestandardreferencematerialsusedontherightLocatingagentsSometimesthesubstancesbeingseparatedarecolourless.Inthiscase,locatingagentscanbeusedtoshowwherethespotsare.Locatingagentsbindtothechemicalsinthespots.Sometimes,anotherchemicalisthenadded,whichreactswiththelocatingagenttoproduceacolouredspot,orthechromatogramisputunderultraviolet[ultraviolet(UV):Electromagneticwaveswithfrequencieshigherthanthoseofvisiblelight,beyondthevioletendofthevisiblespectrum.UVlightcausesdamagetoDNAleadingtosunburnandpossiblefutureskincancer.]lightandthelocatingagentglowstoshowwherethespotsare.RfvalueThemovementofasubstanceduringchromatography,relativetothemovementofthesolvent[solvent:Asolventistheliquidinwhichthesolutedissolvestoformasolution.],ismeasuredbycalculatingitsretardationfactor(Rf).
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ChromatographymeasurementTheRfvalueisworkedoutusingthisformula:
Rfvalue=distancetravelledbysubstance/distancetravelledbysolventCalculatingtheRfvalueallowschemiststoidentifyunknownsubstancesbecauseitcanbecomparedwithRfvaluesofknownsubstancesunderthesameconditions.GaschromatographyIngaschromatography(GC),themobilephaseisaninert[inert:Unreactive.]gas(eghelium).Thestationaryphaseisaverythinlayerofaninertliquidonaninertsolidsupportsuchasbeadsofsilicapackedintoalongthintube(thisflexibletubeiscoiledmanytimesinsideathermostaticallycontrolledoventokeepitataconstanttemperature).GCisusedtoseparatecomplexmixtures.Itismuchbetteratthisthanthinlayerorpaperchromatography.Thisisbecauseitismoresensitiveallowingthedeterminationnotonlyofwhatchemicalsareinthemixture,butalsohowmuchofeachchemicalthereis.
GaschromatographyThemixturetobeanalysedisinjectedintothestreamofcarriergas[carriergas:Theinertgas(oftenhelium)usedingaschromatographytocarrythemixtureofsubstancesthroughthecolumn.].Asitpassesalongthecolumn(longthintube)itseparatesintothedifferentsubstances.Substanceswithagreateraffinity(attraction)forthemobilephasereachthedetectorattheendofthecolumnmorequickly.Substanceswithagreateraffinityforthestationaryphasemovemoreslowlythroughthecolumn.Gaschromatographyisusedtodetectbannedsubstancesinurinesamplesfromathletes.AnalysinggaschromatogramsAgaschromatogram[chromatogram:Toseparatedifferentsubstancesdissolvedinaliquid.]mightshowthetimealongthexaxisandthestrengthofresponsealongtheyaxis.Theamountoftimethatasubstancetakestopassthroughthecolumniscalleditretentiontime.Theretentiontimeofanunknownsubstancecanbecomparedwithstandardreferencedatatohelptoidentifyit.Threemainpiecesofinformationcanbegatheredfromagaschromatogram:thenumberofcompounds[compound:Asubstanceformedbythechemicalunion(involvingbondformation)oftwoormoreelements.]inthemixturerepresentedbythenumberofpeakshowmuchofeachcompoundispresentrepresentedbytheheightofthepeak(higher=more)theretentiontimeindicatedbythepositionofthepeak
Gaschromatographygraph
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Thisgaschromatogramshowsthat:substanceAwaspresentinthesmallestquantity(ithasthesmallestpeak)substanceAhadtheshortestretentiontimesubstancesBandCwerepresentinequalamountssubstanceFhadthelongestretentiontimesubstanceFwaspresentinthegreatestquantity(ithasthelargestpeak)substanceFhadthegreatestaffinityforthestationaryphaseMorefromChromatography
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