Chemistry Reference
In-Depth Information
immediatelyafteriltration.Themostcommonlyusedmethodsarebasicallyreversed-phaseHPLC
separationscoupledwithUVlightdetector.
NMRspectroscopyandmassspectrometry(MS)areusefultechniquesforstructuralelucida-
tionofunknowncompounds,buttheresultsobtainedaredificulttoquantify.Liquidchromatog-
raphy(LC)methodshavebeenusedextensivelyforthedeterminationofhighlyintensesweeteners
because,inmanyinstances,thesamplematricesfromwhichtheyaretobedeterminedmaybecom-
plex.Inaddition,asweetenermaybeusedincombinationwithothersweetener(s).Nevertheless,in
recentyears,capillaryelectrophoresis(CE)methodshavebeendevelopedthatcompetesuccessfully
withLCmethodologies.TherearenomethodologiesproposedforthedeterminationofNTMand
onlyafewmethodsforalitameandthaumatin.Mostmethodsforthedeterminationofthaumatin
involveimmunochemicalassays(IC)andmeasurementinanenzyme-linkedimmunosorbentassay
reader.Ontheotherhand,thelargestnumberofmethodologieshasbeenproposedforthedetermi-
nationofsaccharinbecauseitistheoldestknownandusedhigh-intensitysweetener.Automated
continuousdeterminationbyFIAhasbeendevelopedforACS-K,ASP,cyclamate,andsaccharin;
generally,theseproceduresinvolvespectrophotometricandelectroanalyticaldetections.
4.15.1 Determination of aCS-K
Inthelastfewyears,therehasbeenaninterestinthedevelopmentofanalyticaldevicesforthe
detectionandmonitoringofvariousbiologicalandchemicalanalytes.Forseveraldecades,analytical
chemistswereinspiredfrombiologicalsciences,andnowadays,thedetectionofanalytesusingbio-
sensorsisverycommon.Biosensorsofferthecapabilitytodevelopamethodfortherapidscreeningof
thesesweetenerswitharespectivelowcost(NikolelisandPantoulias2000).Lipidilmscanbeused
fortherapiddetectionorcontinuousmonitoringofawiderangeofcompoundsinfoodsandinthe
environment.Suchelectrochemicaldetectorsaresimpletofabricateandcanprovideafastresponse
andhighsensitivity.Investigationshavetakenplacefortheinteractionsoftheartiicialsweeteners
ACS-Kwithfreelysuspendedbilayerlipidmembranes(BLMs)thatcanbeusedforthedirectsensing
ofthesesweeteners.TheinteractionsofthesweetenerswiththeseBLMsproducetransientelectro-
chemicalcurrentsignals,themagnitudeofwhichcouldbeusedtoquantifytheconcentrationofthe
sweetener.Determinationofthemechanismofsignalgenerationinvolvesanalysisofthestructural
effectscausedbyinteractionsofthesweetenerswithmodellipidmembranes.Differentialscanning
calorimetrystudieshaveshownthattheinteractionsofsweetenerswithlipidvesiclesstabilizethe
gelphaseoflipidilms.Monolayercompressiontechniquesatanair-waterinterfacehaverevealed
anincreaseinthemolecularareaoflipidswhensweetenersareaddedtotheaqueoussubphase.Such
structuralchangesareknowntocauseelectrostaticandpermeabilitychangeswithlipidmembranes.
AconductometricmethodbasedontheuseofsurfacestabilizedBLMs(s-BLMs)composedfrom
eggphosphatidylcholineisdevelopedformonitoringACS-Kandothersweeteners.Theinteractions
ofsweetenerswiths-BLMsproduceareproducibleelectrochemicalioncurrentsignalincreasethat
appearswithinafewsecondsafterexposureofthemembranestothesweetener.Thecurrentsignal
increasesrelativelytotheconcentrationofthesweetenerinbulksolutioninthemicromolarrange.
Thealterationsinthecurrentsignalareobservedevenfromlowconcentrationsbetween0.4and
7 μMinelectrolytesolutions.Thes-BLM-basedbiosensorisstableforlongperiodsoftime(over
48h)andcanbeeasilyconstructedatlowcost(andthereforecanbeusedasadisposablesensor)
withfastresponsetimesontheorderofafewseconds.Thelipidlayerisdepositedintoanascent
metallicsurfacewhileimmersingthemetalwireintothelipidsolution.Thewirewiththelipidlayer
isimmersedintoKClaqueoussolution,andioniccurrentisstabilizedoveraperiodof10to15min
dependingonthediameterofthesilverwire.Calibrationsaresubsequentlydonebystepwiseaddi-
tionsof0.1or1.0mMsweetenerstandardsolutionaddedtotheKClelectrolytewhilecontinuously
stirring.Oncethecalibrationplotoritsequationissetup,theunknownsweetenerconcentrationof
asolutioncanbeindependentlydeterminedusingafreshBLMonanascentmetallicsurface,and
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