NUCLEARMAGNETICRESONANCE

(NMR)

NMR Spectroscopy:
Introduction

History of NMR
NMR Hardware and Software
Superconducting Cryo-Magnet
Probe
RF Console
Computer + NMR Software
+ Printer/Plotter
Solution NMR
Sample Preparation
Presentation of Data

History- NMR Nobel Prize Winners

1943 Otto Stern
(Physicist)

"for his contribution to the development of the molecular ray

method and his discovery of the magnetic moment of the proton".

1944 Isidor I Rabi

(Physicist)

"for his resonance method for recording the magnetic properties

of atomic nuclei".

1952 Felix Bloch

& Edwin M
Purcell
(Physicist)
1991 Richard Ernst
(Chemist)

"for their development of new methods for nuclear magnetic

precision measurements and discoveries in connection
therewith"

2002 Kurt Wthrich

(Chemist)

"for the development of methods for identification and structure

analyses of biological macromolecules"

2003 Paul Lauterbur

& Sir Peter
Mansfield
(Chemist-Physicist)

"for their discoveries concerning magnetic resonance imaging"

"for his contributions to the development of the methodology of

high resolution nuclear magnetic resonance (NMR)
spectroscopy".

nobelprize.org

History of NMR
1938 - NMR in molecular beams
Rabi (Columbia University)
1946 - NMR of Liquids and Solids
Purcell, Torrey, Pound (Harvard)
Bloch, Hansen, Packard (CalTech)
1952 - First commercial NMR spectrometer (30 MHz)
1962 - First Superconducting Magnet for NMR
1968 - First Pulse Fourier Transform NMR
1969 - First Concept of MRI Scanners
1971 - First 2D NMR Experiment COSY (Jean Jeener)
1985 - Protein Structures
2009 - First Gigahertz NMR Spectrometer (1.0 GHz)

MRI

NMR Manufacturers
Bruker corp.
Varian Inc.
JEOL Ltd
Agilent Tech.

HIGH RESOLUTION

Oxford Instruments plc.

LOW RESOLUTION
Qualion NMR
OST Engineering
picoSpin LLC

Composition analyses:
- Oil and petrochemical industries
- Edible oils
- Soluble fat and water contents
- Pharmaceutical products
- FAME and LNG

NMR Magnet Safety

NMR magnets are always charged!

NMR magnets contain Cryogens (liquid Helium and

Nitrogen)

11

NMR Safety
NMR magnets are always charged!
Interfere with medical devices (i.e. pacemakers, metallic implants)
Interfere with electronic and mechanical devices and may damage
them (cell phones, pagers, watches, etc.)
Erase credit cards, ID cards, floppy disks, hard disks (I-pods and
some other mp3 players).
Attract ferromagnetic objects of any size (i.e. paper clips, coins, keys,
pens, scissors, screw drivers, wrenches, metallic chairs, gas cylinders,
etc.).
NMR magnets contain Cryogens (liquid Helium and Nitrogen)
Cryogens can cause severe burns if handled improperly (use eye
protection and gloves).
Cryogens evaporate and may cause asphyxiation if a lab is not
properly ventilated.
During a refill the refill tubing may shatter. Frozen rubber cuts like
glass!!

NMR INSIDE

Liquid Helium:
- Inert
- Light and abundant (after H2)
- Protect the samples from
heating of superconductor.
Liquid Nitrogen:
- Stable
- Abundant
- Thermal buffer between He and
environment
Magnet:
- Paramagnetic
- Source of external magnetic
field

Niobium-Tin
Niobium-Titanium

NMR Probes

bruker.com

Dual: 1H/13C
Dual: 15N/19F
Triple: 1H/13C/15N
Quadruple: 1H/13C/15N/31P

Nuclear Spins
NMR is concerned with change in the direction of
spin orientation as the result of the absorption of
radiofrequency radiation.

http://www.xearththeory.com

Spins and Chemical Shift

The distribution of nuclear
spins is random in the
ABSENCE of an external
magnetic field.

+
+

An external magnetic field causes

nuclear magnetic moments to align
parallel and antiparallel to applied field.

+
+

H0

+
+

There is a slight excess of nuclear

magnetic moments aligned parallel to
the applied field (thermodynamic
stability)

H0

+
+

13

C NMR Probes (conventional vs cryroprobe)

bruker.com

SCHEME OF NMR

Magnet (Applied Field)

Radiofrequency energy is transmitted and

absorption is detected

Preparasi sampel NMR

5 mm

Use clean + dry NMR tubes and caps

(tubes can be re-used, caps should not!)
0.5 ml deuterated solvent
(i.e. CDCl3, C6D6, acetone-d6,etc.)
substrate requirements for routine spectra:
10 mg for proton NMR
100 mg for carbon-13 NMR
min. filling height of tube: 2 inches (5 cm)
Cleaning of tubes:
1. rinse with solvent you were using
2. rinse with acetone
3. dry in (vacuum-)oven at low temperature

Preparasi sampel NMR

Clean
clear
solution

GOOD!

Suspension or
opaque
solution

Precipitate

Concentration
gradient

Two
phases

Bad Samples!

Not
enough
solvent

Shigemi

State of the Art of NMR

What Next??

Nuclear Magnetic Resonance (NMR) adalah salah satu teknik spektroskopi yang dapat memberikan
informasi tentang lingkungan magnetik suatu inti.
Dalam keadaan resonansi, inti akan menyerap radiasi elektromagnetik di dalam daerah gelombang radio
dengan frekuensi yang tergantung pada lingkungan kimianya. Lingkungan kimia tersebut dipengaruhi
oleh ikatan kimia, konformasi molekul dan proses dinamik.
NMR umumnya digunakan dalam kimia organik untuk menentukan struktur molekul dan konformasi
dengan mempelajari inti 1H dan 13C.
Supaya terjadi transisi proton dari keadaan spin yang satu ke keadaan spin yang lainnya, sampel
ditempatkan di daerah medan magnet Bo dan frekuensi radiasi
elektromagnetik u diubah-ubah sampai tercapai resonansi.
Senyawa yang digunakan sebagai standar adalah tetrametilsilan (CH 3)4Si (TMS), karena protonprotonnya beresonansi pada medan magnet yang lebih besar daripada medan magnet untuk resonansi
proton senyawa organik lainnya. TMS memiliki 12 proton yng setara lingkungan kimianya dan
memberikan 1 puncak, TMS bersifat inert.

Geseran Kimia () = Perbedaan letak resonansi suatu proton tertentu dengan letak resonansi proton TMS. Inti
dapat berada dalam keadaan shielded atau deshielded saat inti dipengaruhi oleh medan magnet disekitar
elektron. Elektronegativitas adalah salah satu faktor penting terhadap distribusi elektron sekitar ikatan.

ChemicalShiftData
Aspreviouslymentioned,differentkindsofprotonstypicallycomeatdifferentchemicalshifts.Shown
belowisachartofwheresomecommonkindsofprotonsappearinthescale.Notethatmostprotons
appearbetween0and10ppm.Thereference,tetramethylsilane(TMS)appearsat0ppm,andaldehydes
appearnear10ppm.Thereisapageinyourlabhandoutwithmoreprecisevaluesforthischart.
Notethatthesearetypicalvaluesandthattherearelotsofexceptions!

R
R

NH

OH
Ph

OH

O
H

1
D
o

(
R
HO

9
re

0
o

fth

7
g

w
e

5
n

fi

NR2 O
CH3

C H3

Me

Si M
Me
Me

R
CH3

CH3
TM S

3
p

Me
T

Ph CH 3

Cl

4
n

C H3

OC H3

6
io

2
e

1
U d
l
o
c

0
re

p
ft
h

tru

H spektrum dari Etanol

TheHardPartInterpretingSpectra
LearninghowanNMRmachineworksisstraightforward.Whatislessstraightforwardislearninghowto
usethedatawegetfromanNMRmachine(thespectrumofethylacetateisshownbelow).Thatsbecause
eachNMRspectrumisapuzzle,andtheresnosinglefactthatyousimplyhavetomemorizetosolvethese
spectra.Youhavetoconsiderlotsofpiecesofdataandcomeupwithastructurethatfitsallthedata.What
kindsofdatadowegetfromNMRspectra?For 1HNMR,therearethreekindseachofwhichwewill
considereachoftheseseparately:

1) Chemicalshiftdatatellsuswhatkindsofprotonswehave.
2) Integralstellsustheratioofeachkindofprotoninoursample.
3) 1H1Hcouplingtellsusaboutprotonsthatarenearotherprotons.

ChemicalShiftData

Integrasi
Integralstellustheratioofeachkindofproton.Theyarelines,theheightsofwhichareproportionaltothe
intensityofthesignal.Considerethylacetate.Therearethreekindsofprotonsinthismolecule,theCH 3
nexttothecarbonyl,theCH2nexttotheOandtheCH3nexttotheCH2.Theratioofthesignalsarising
fromeachofthesekindsofprotonsshouldbe3to2to3,respectively.So,ifwelookattheheightofthe
integralstheyshouldbe3to2to3.Withthisinformation,wecanknowwhichistheCH 2signal(itsthe
smallestone),buttodistinguishtheothertwo,wehavetobeabletopredicttheirchemicalshifts.Thechart
onthepreviouspageallowsustomakethatassignment(theCH 3nexttotheC=Oshouldappearat~2PPM,
whiletheotherCH3shouldbeat~1PPM).

3H'S

O
O

O
O

CH3

O
H 3C

3H'S

2 H'S

Integrasi

Chemicalshiftandintegrasi

Jason,C.Kwan,etal.,JournalofNaturalProduct,February18,2010

Jek-ek = 2-4 Hz

Jak-ek = 2-4 Hz

H
H

Jak-ak = 11-13 Hz

H
O

H
3

J = 9-12 Hz

J = 14-16 Hz

H1HCoupling

H
H

Orto = 6-8 Hz
Meta = 1-3 Hz
Para = < 1 Hz

More1H1HCoupling
Whathappenswhenthereismorethanoneprotonsplittinganeighboringproton?Wegetmorelines.
Considerthemoleculebelowwherewehavetwoprotonsononecarbonandoneprotononanother.

N
o
b HA + yA is
o
fth

HAa
c
in
T
(c
fe

HA +
A an
h
id
h
a
e

HA'
' pd
e
e
y
e
le
lth

m
a
d

pH
n

e.
B

ic
r
e
a

te
'

t
w
a

th
B

i
c
tp

HB
ras

e
tic
m

e
ro

th

t
h
d

HA '

e
u

s
c

HA

HB

ig

Ha
thp
elitn
sto
ath
Bis
ba
e ls
ch
a iftb
ue
s c
afi
lee
HAnldn
od
irofH
n
Aa
' v
a
l
s
o
s
p
l
itn
d
o
u
b
le
t
)b
a
g
n
e
t
ic
fi

i
z
e

ea
to
e

mre
m
e

e
d

itfe u

tw
c

H1HCoupling

WhyareThereThreeLinesforHB?
HBfeelsthesplittingofbothHAandHA.So,letsimaginestartingwithHBasasingleline,thenletsturn
onthecouplingfromHAandHAoneatatime:
H

Ifu
s
N

Bw

h
t
th
N
s
b
e
v
o

n
in

HB-H Aco

c
g

e
e

H B-H Aco
p
u
rla

o w
,le

o
le

c
s
' oHw
,le
litsA '
tn
p

ut's
HA

HB

Becausethetwolinesinthemiddleoverlap,thatlineistwiceasbigasthelinesontheoutside.More
neighboringprotonsleadstomorelinesasshownonthenextslide.

h
in
ut's
e

o
.
O

SplittingPatternswithMultipleNeighboringProtons
Ifaprotonhasnneighboringprotonsthatareequivalent,thatprotonwillbesplitinton+1lines.So,ifwe
havefourequivalentneighbors,wewillhavefivelines,sixequivalentneighborswell,youcandothe
math.Thelineswillnotbeofequalintensity,rathertheirintensitywillbegivenbyPascalstriangleas
shownbelow.
e in te n s ite s p a te rn
n o .o n
f e g
i h b reo lars tiv
0
1
s in g le t(s )
1
2
3

1 1
1 2 1
1 3 3 1

x a m p le
e

d o u b le t(d )

trip le t()
q u a e
tr t(q )

4
5
6

1 4 6 4 1
1 5 1 0 1 0 5 1

p e n te t
s e
x e
t t

1 6 1 5 2 0 1 5 6 1 s e p te t

H
H

H
H

Wekeepemphasizingthatthispatternonlyholdsforwhentheneighboringprotonsareequivalent.Whyis
that?Theansweristwoslidesaway.

MoreAboutCoupling
Earlierwesaidthatprotonscoupletoeachotherbecausetheyfeelthemagneticfieldoftheneighboring
protons.Whilethisistrue,themechanismbywhichtheyfeelthisfieldiscomplicatedandisbeyondthe
scopeofthisclass(theydontjustfeelitthroughspace,itstransmittedthroughtheelectronsinthe
bonds).Itturnsoutthatwhentwoprotonsappearatthesamechemicalshift,theydonotspliteachother.
So,inEtBr,wehaveaCH3nexttoaCH2,andeachprotonoftheCH3groupisonlycoupledtothe
protonsoftheCH2group,nottheotherCH3protonsbecausealltheCH3protonscomeatthesame
chemicalshift.

T b
a

HH
HCCBr
HH

T re p
a
a
H

Br

h
nth
Br

lu

Metodologi Penentuan Struktur

Struktur Molekul = Kerangka + Gugus Samping
Contoh:

O
H3C

O
CH3

H3C
H3C
N
CH3

CH3

kerangka

3 X CH3

gugus
samping

kerangka

N(CH3)2 + OCH3

gugus
samping

Manakah yang lebih dahulu ditentukan:

Kerangka atau gugus samping
Metodologi1
Senyawaalifatik(siklik):Gugussamping
Tahapankerjanya:

Kerangka

Identifikasigugussamping
Tentukankerangka
Tempatkangugussampingpadakerangka

Metodologi2
Senyawaaromatik:Kerangka
Tahapankerjanya:

GugusSamping

Tentukankerangka
Tentukanpolasubstitusipadakerangka
Identifikasigugussamping

Metodologi 1
C10H16O

DBE=(2216)/2=3

Note:DBE(DoubleBondEquivalents)
Halogen=hidrogen
Nitrogen=gugusCH
OksigendanSulfur=tidakdiganti
Formula:(2n+2x)/2;n=karbon
x=hidrogen

Gugussamping:3xCH3(C3H9)
O
H3C
H3C

Kerangka:C7H7Odengan3DBE
CH3

Metodologi 2
C9H13NO

DBE=(2214)/2=4

Kerangka:benzena(4H)

Gugussamping:C3H9NO(0DBE)

C6H4(4DBE)

PrediksikanStrukturSenyawaBerikut
C4H5O2Br
3

C5H4O2

C3H6O2

C4H7ON

C3H9N3

1
2

C6H7N2Br

2
1

C6H9O2N

2
1

C7H8O4

C7H7N

2
1

C8H10

6
4

C8H10O2

C15H14O

PENGANTAR NMR 2 DIMENSI

HSQC : one bond

H-13C correlation

HMBC : long range H- C correlation

COSY : proton-proton coupling
1

13

NOESY/ROESY : spatial relationships between protons

TOCSY :

long range proton-proton connectivities

Maitotoxin

COSY

HSQC (edit)

NOESY

H1HCoupling

Youllnoticeinthespectrathatweveseenthatthesignalsdontappearassinglelines,sometimesthey
appearasmultiplelines.Thisisdueto1H1Hcoupling(alsocalledspinspinsplittingorJcoupling).
Hereshowitworks:Imaginewehaveamoleculewhichcontainsaproton(letscallitH A)attachedtoa
carbon,andthatthiscarbonisattachedtoanothercarbonwhichalsocontainsaproton(letscallitH B).It
turnsoutthatHAfeelsthepresenceofHB.Recallthattheseprotonsaretinylittlemagnets,thatcanbe
orientedeitherwithoragainstthemagneticfieldoftheNMRmachine.WhenthefieldcreatedbyH B
reinforcesthemagneticfieldoftheNMRmachine(B 0)HAfeelsaslightlystrongerfield,butwhenthefield
createdbyHBopposesB0,HAfeelsaslightlyweakerfield.So,weseetwosignalsforH Adependingonthe
alignmentofHB.ThesameistrueforHB,itcanfeeleitheraslightlystrongerorweakerfieldduetoH As
presence.So,ratherthanseeasinglelineforeachoftheseprotons,weseetwolinesforeach.

o w th
F
( a
m
c

is
B

A is
H

HA

o a
F
(s
m
c

th

is
B

B is
H

B
H

rth ith e
d
a H H
o
AB
C C

lin

NotallCouplingsareEqual
Whenprotonscoupletoeachother,theydosowithacertainintensity.Thisiscalledthecoupling
constant.Couplingconstantscanvaryfrom0Hz(whichmeansthattheprotonsarenotcoupled,even
thoughtheyareneighbors)to16Hz.Typically,theyarearound7Hz,butmanymoleculescontain
couplingconstantsthatvarysignificantlyfromthat.So,whathappenswhenamoleculecontainsaproton
whichiscoupledtotwodifferentprotonswithdifferentcouplingconstants?Wegetadifferentpatternas
describedinthediagrambelow.

So,iftheprotonsarenotequivalent,theycanhavedifferentcouplingconstantsandtheresultingpattern
willnotbeatriplet,butadoubletofdoublets.Sometimes,nonequivalentprotonscanbeonthesame
carbonasdescribedonthenextslide.

CouplingConstantsinAlkenes
Couplingconstantsinalkenescanalsodifferdependingonwhethertheprotonsarecisortranstoeach
other.Notethatinaterminalalkene(i.e.,analkeneattheendofacarbonchain),thecisandtransprotons
areNOTequivalent.Oneisonthesamesideasthesubstituent,theotherisontheoppositeside.The
couplingoftransprotonstoeachotheristypicallyverylarge,around16Hz,whilethecouplingofcis
protons,whilestilllarge,isalittlesmaller,around12Hz.Thisleadstothepatternshownbelow,andan
exampleofamoleculewiththissplittingpatternisshownonthenextslide.

HA

1 H

M
H

HX

I
fu
s

2
th

HA
w

HA
- HX
c

1 H

2
H A -H M c

N
th

6 z

Thereareothertimeswhenprotonsonthesamecarbonarenonequivalent,whichwellseelater.

H
e

HO

HO

CH 3

H
H

HO

HO
H

H HO

CH 3

H
H HO

HO

H
H

m
Me

OH
Me

N
s

Me
OH

Me
H

H
Me

i
n
e
th

Me
OH

OH

H
Me

y
e

Me
OH

DiastereotopicProtonsinanNMRSpectrum

Me OH

H
H

H Me OH

OH

H H

OH

H H
H

Me OH

OH

Ha Hb

HO

HO Me

H H

Thereareseveralimportantfeaturestothisspectrum.NoticethattheprotonsmarkedH aandHbare
diastereotopicandthattheyappearatdifferent.WecantassignwhichisHaandwhichisHb,butwe
knowtheyaredifferent.Also,noticethatthelinesforH aandHbarenotofequalheights;theoutsidelines
aresmallerthantheinsidelines.SuchleaningofNMRlinesiscommonandisdescribedonthenext
slide.

LeaningofNMRlines
Recallthatifwehavetwoprotons,HaandHb,andtheyarecoupledtoeachother,Hawillbesplitintotwo
linesbyHb,andHbwillbesplitintotwolinesbyHa.So,weshouldseeaspectrumwheretherearefour
lines,twoforeachproton.But,itturnsoutthatthepatternsleantowardseachotherinthattheoutside
linesofthepatternaresmallerthantheinside:

o
p

th

lin

o
p
th

u
a

Ha Hb
C

Howmuchleaningwehavedependsonhowclosetoeachotherthesignalsareinthespectrum,andhow
stronglytheyarecoupledtoeachother.Theclosertheyare,thegreatertheleaning,thestrongerthe
coupling,thegreatertheleaningasshownonthenextslide:

BacktothePreviousNMRSpectrum

Me OH

H
H

H Me OH

OH

H H

OH

H H
H

Me OH

OH

Ha Hb

HO

HO Me

H H

NoticetheappearanceoftheOHpeakinthisspectrum;itsbroadandnotcoupledtoitsneighbors.Thisis
commonwithOHpeaks.OHpeaksareunusualinthattheirappearanceisvariable;forthesamemolecule,
sometimestheyarecoupledsometimestheyarenot,sometimestheyaresharp,sometimestheyarebroad.
Infact,sometimestheyareverybroad,sobroadthatyoucantevenseethem!ThisisduetoOHgroups
undergoingexchangewitheachotherandwithtracesofwaterinthesolvent.Thisprocessiscatalyzedby
tracesofacidorbaseinthesolventwhicheffectstherateoftheexchangeandtheappearanceoftheOH.

MoreonthisNMRSpectrum

Me OH

H
H

H Me OH

OH

H H

OH

H H
H

Me OH

OH

Ha Hb

HO

HO Me

H H

NowletsconsidertheappearanceofthePhgroup.Itsamess!Notethattherearemorecouplingsthanwe
wouldexpectbasedonanearestneighboranalysis.Thatsbecausealltheprotonscomeatsimilar,and
overlapwitheachother.Ingeneral,whenevertherearealotofsignalscoupledtoeachotherthatare
bunchedupinaportionofthespectrum,wegetcomplexpatternswithextralines.Thereasonforthisis
calledvirtualcoupling,andanexplanationofthisisbeyondthescopeofthisclass,butyoushouldbeaware
whenyouaresolvingspectrathatsometimes,thiscausesthepatternstobemorecomplexthanyouwould
initiallyexpect.