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1)whatisspectroscopy
2)whatisthebasicprincipleofNMR
3)whatischemicalshift
4)factorsaffectingchemicalshift
5)whatisintegrationofapeakinNMR
6)spinspincoupling
7)firstvssecondorderspectra
8)PopleNotation
10)NMRrelaxation
11)T1Relaxation
12)T2Relaxation
13)whydoweneed2DNMR
14)typesof2DNMR
15)whatiscosy
16)whatisNOESY
Whatisspectroscopy?
Spectroscopyinvolvesthestudyofinteractionofradiationwithmatter
WhatisthebasicprincipleofNMR
TheprinciplebehindNMRisthat
nucleihavespindenotedbyI,nucleusofspinIwillhave2I+1possibleorientations.
Intheabsenceofanexternalmagneticfield,theseorientationsareofequalenergy.Ifa
magneticfieldisapplied,thentheenergylevelssplit.
Ifanexternalmagneticfieldisapplied,anenergytransferispossiblebetweenthe
lowerenergytoahigherenergylevel.
Theenergytransfertakesplaceatawavelengththatcorrespondstoradiofrequencies
andwhenthespinreturnstoitslowerlevel,energyisemittedatthesamefrequency.
Thesignalthatmatchesthistransferismeasuredinmanywaysandprocessedin
ordertoyieldanNMRspectrumforthenucleusconcerned.
Thereare4levelsofinformationtobegainedfromprotonNMRoranyrelatedtodifferent
aspectsofthespectrum:
1.HowmanytypesofH?
Givenbyhowmanybasicgroupsofsignalsarethere
2.HowmanyHofeachtype?
givenbytheintegration(relativearea)ofeachgroup.
3.Whichisthegrouptowhichhydrogenisattached?
givenbythechemicalshiftofeachgroup
4.informationaboutneighbouringgroup
givenbycouplingpatterns.
Whatischemicalshift?
chemicalshiftistheresonantfrequencyofanucleusrelativetoastandardina
magneticfield.
Chemicalshiftisusuallyexpressedinpartspermillion(ppm)byfrequencygivenby
factorsthatcausedifferentchemicalshifts
1)Localdiamagneticshielding
2)Localparamagneticshielding
3)Anisotropy
localdiamagneticshielding
theexternalfieldcauseselectronstocirculateinaplaneperpendiculartothefield
direction.thecurrentsoinducedgeneratesasmalllocalfieldopposedtoBotherefore
shieldthenucleus
localparamagneticshielding:
whenanexternalmagneticfieldappliedalongthezaxisdistortsthewavefunctionof
thegroundstatebymixingintoitsmallfractionoftheexcitedstatewavefunction.inthis
waythefieldpartiallyhelpovercometheenergygapbetweenthehigherexcitedorbital
andcreatesapathfortheelectrontocirculateinxyplane.thisinducedcurrent
generatesamagneticfieldwhichdeshieldthenucleus
Anisotropy:
Thisjustmeans"anonuniform"magneticfield.
Electronsinpisystems(e.g.benzene,alkynes)interactwiththeappliedfieldandare
causedto"circulate"withinthepisystem.Thismotioninducesamagneticfieldthat
causestheanisotropy.
Asaresult,thenearbyprotonswillexperience3fields:theappliedfield,theshielding
fieldofthevalenceelectronsandthefieldduetothepisystem.
Dependingonthepositionoftheprotoninthisthirdfield,itcanbeeithershieldedor
deshielded,
WhatisintegrationofpeakinNMRmean?
AmeasureoftheareaofthepeakswhichisproportionaltothenumberofHthatthe
peakrepresents.
SPINSPINCOUPLING
Itistheinteractionbetweenthespinmagneticmomentsofdifferentnuclei.InNMR
spectroscopyitgivesrisetomultipletpatterns
Splittingpatternofpeaks,ingeneral,followthetreediagrampattern
Incaseisthegivennucleusiscoupledtonequivalentnucleicertainpealsoverlap
whichresultsindoubleortripletheareaunderthepeakdependingonthenumberof
peaks
The(n+1)Ruleanempiricalruleusedtopredictsplittingpatternofpeaksin1Hand
13CNMRspectra
Itstatesthatifagivennucleusiscoupledtonnumberofnucleithatareequivalent,the
multiplicityofthepeakisn+1.
Incasethegivennucleusiscoupledtononequivalentnucleiitleadsto2^ndifferent
peaks
Mechanismofspinspincoupling
Onenuclearspin
Intheabsenceofnuclearspin,electronspinsarepairedaccordingtothePauli
principle.Theupdown'anddownup'stateshavethesameenergy
Ifonenuclearspinispresent,thetwospinstatesnolongerhaveexactlythe
sameenergy.
Theinteractionbetweennuclearandelectronspinsisnamedthehyperfine
interactionandactstolowertheenergyofthespinthatispairedwiththe
nuclearspin
Twonuclearspin
Intheaboveimage,twodifferenttransitiontakesplaceforBnucleusinABbond
Fromfavouredgroundstate.Inthisarrangement,Bnucleusisinitslowestenergy
stateandallofthenuclearandelectronicarepaired,resultinginlowestenergyforthe
system.ThesecondoptionisnucleusofatomAandBareparallelresultinginhigher
energysystem
NowlettheBnucleusundergotransitionandinvertitsspin.theexcitedstatethat
resultsfromthehigherenergygroundstatehasalowerenergybecauseallitsnuclear
andelectronicspinsarepaired,whiletheexcitedstatethatresultsfrommorestable
groundstateisofhigherstatebecauseitsspinsarenotpaired
ThusweseetwodifferenttransitionsforBnucleusdependingonthespinofA,asa
result,weseeadoubletinthespectrum
Relationbetweencouplingconstatntandchemicalshift
Firstorderspectraisobservedwhenthereisweakcouplingthatisdifferencein
chemicalshiftisfargreaterthanthecouplingconstant
Secondorderspectraarespectrawhichrequiremathematicalanalysisfor
interpretation.Generallyobservedwhensystemisweaklycoupledthatisthedifference
inchemicalshiftbetweentwogroupissimilarinmagnitudetothecouplingconstant,j
WhatisSpinsystemnotationorPopleNotation
Itisconvenientshorthandnotationtodescribethetypeofspinsystem.
Eachchemicallydifferenttypeofprotonisgivenaletter:A,B,Candsoforth
Ifagrouphastwoormoreprotonofonetype,theyaredenotedbysubscriptsasinA2
orB3
Protonsofsimilarchemicalshiftvaluesareassignedlettersthatareclosertoone
anotherinthealphabet,suchasA,B,C.
Protonsofwidelydifferentchemicalshiftareassignedlettersfarapartinthealphabet,
suchX,Y,XvsA,B,C.
InatwoprotonsystemthatexhibitsfirstorderiscalledanAXsystem
InatwoprotonsystemthatexhibitssecondorderiscalledanABsystem
Inatwoprotonsystemthatexhibitsidenticalchemicalshiftandaremagnetically
equivalentisdenotedbyA2
Inatwoprotonsystemthatexhibitsidenticalchemicalshiftandarenotmagnetically
equivalentisdenotedbyAA
ifthreeprotonsareinvolvedandtheyhaveaverydifferentchemicalshift,aletterfrom
themiddleofthealphabetisused,usuallyMegAMX.
ifthreeprotonsareinvolvedandtheyhaveverysimilarchemicalshiftitisdenotedby
ABC
ExaminingthesystemoftwoprotonHAandHBonadjacentcarbonatoms
weexpecttoseetwodoubletswitheachofequalintensityusingN+1rule.
Butweseetwodoubletonlywhenchemicalshiftislargecomparedtothecoupling
constant
IncaseofHAandHBidenticalchemicalshiftandaremagneticallyequivalent
(A2)weseeonlyonepeak
Forallothercasesasadifferenceinachemicalshiftbycouplingconstantbecomes
smallerandsmallerthesplittingpatterncontinuouslychangesi.etheintensityofthe
innerpeakscontinuouslyincreasesinintensitywhiletheouterpeakofthedoublet
decreasesinintensity.
whatisNMRrelaxation?
NMRrelaxationistheprocessesbywhichanexcitedmagneticstatereturnstoits
equilibriumdistribution.
Ithastwocomponentlongitudinalandtransverserelaxation
T1Relaxation(longitudinalrelaxation)
T1isthetimeconstantforregrowthoflongitudinalmagnetisation(Mz)
Requiresenergytransferfromspinstoenvironment("lattice")
Energytransferisstimulatedbyfieldfluctuatingcausedbymolecularmotionofa
nearbyprotonorelectronneartheLarmorfrequency
Graphindicatingprocessbywhichthenetmagnetisationreturnstoitsinitialmaximumvalue
(Mo)
T2RelaxationorTransverserelaxation
T2isthetimeconstantfordecay/dephasingoftransversemagnetisation
Mayoccurwithorwithoutenergytransfer
AnythingcausingT1relaxationalsocausesT2relaxation,butT2relaxationcanoccur
withoutT1relaxation.
MajorcausesofT2onlyrelaxationincludedephasingbystaticlocalfield
Graphindicatingthedephasingoftransversemagnetisation
Whydowerequire2DNMR
1)Itgivesusawaytoknowwhichproton/carbonarecorrelatedwitheachother
2)Ithelpsresolveoverlapofcomplexstructurebycorrelatingchemicalshift
2DNMRBasics
Thepreparationtime
Timerequiredtoallowtheexcitednucleitogetbacktheirequilibriumstate
Theevolutiontime
Duringtheevolutiontime,thespinsystemisevolvingundertheeffectofdifferent
factors,eachcoherenceevolvesatitsowncharacteristicfrequencyasafunctionofthe
chemicalshiftandofthescalarcouplingofthecorrespondingnucleus.
TheMixingtime
Itismadeofapulsesequencewhichachievestransfersofmagnetisationdifferent
frequenciesarecorrelated.
Thedetectiontime
Theacquisitionofthemodulatedsignaltakesplaceduringthedetectionperiod.The
sequencewejustdescribeddoesnotconstitutebyitselfa2DNMRexperiment.
typesof2DNMR
1)Homonuclear
Throughbond:COSY,TOCSY,2DINADEQUATE,2DADEQUATE
Throughspace:NOESY,ROESY
2)Heteronuclearcorrelation
OnebondcorrelationHSQC,HMQC
LongrangecorrelationHMBC
(ihavecoveredonlyCOSYANDNOESY.IalsohaveabasicideaofTOCSYANDROESY.
RestIamyettocover)
WhatisCOSY
COSY(CorrelatedSpectroscopY)isausefulmethodfordeterminingwhichsignals
arisefromneighbouringprotons.
Correlationsappearwhenthereisspinspincouplingbetweenprotonsandwhere
thereisnocoupling,nocorrelationisexpectedtoappear.
Thismethodisveryusefulwhenthemultipletsoverlap
WhatisNuclearOverhausereffect?
Whenaspecificnucleusismagneticallyexcitedanditsneighbourisatequilibrium,
relaxationoccursbetweenthetwonucleiBecauseofwhichtheintensityofthenucleus
thatwasatequilibriumchanges.
WhatisNOESY
InNOESY,thenuclearOverhausercrossrelaxationbetweennuclearspinsthat
happensduringthemixingperiodisusedtoestablishthecorrelationsnucleithatare
spatiallycloseratherthanthosethatarethroughbondcoupledtoeachother.
NOESYisusefulfordeterminingwhichsignalsarisefromprotonsthatarecloseto
eachotherinspaceeveniftheyarenotbonded.
OneapplicationofNOESYisinthestudyoflargebiomolecules