SEMMELWEIS UNIVERSITY, DIALOG CAMPUS PUBLISHER

Development of Complex Curricula for Molecular Bionics and Infobionics Programs within a consortial* framework**

Consortium leader

PETER PAZMANY CATHOLIC UNIVERSITY

Consortium members

SEMMELWEIS UNIVERSITY, DIALOG CAMPUS PUBLISHER

The Project has been realised with the support of the European Union and has been co-financed by the European Social Fund ***

**Molekuláris bionika és Infobionika Szakok tananyagának komplex fejlesztése konzorciumi keretben

***A projekt az Európai Unió támogatásával, az Európai Szociális Alap társfinanszírozásával valósul meg.

CATHOLIC UNIVERSITY

UNIVERSITY

World of Molecules

The structure of the molecules how we see (with spectroscopy)

(Molekulák Világa )

(A molekulák szerkezete: ahogyan azt (spektroszkópiával) látjuk)

Compiled by dr. Péter Mátyus

with contribution by dr. Gábor Krajsovszky

Formatted by dr. Balázs Balogh

Table of Contents

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1. Ultraviolet Spectroscopy 4 – 8

2. Infrared Spectroscopy 9 – 14

3.

H-NMR Spectroscopy 15 – 35

4.

C-NMR Spectroscopy 36 – 44

5. Mass Spectroscopy 45 – 50

6. X-ray Crystallography 51 – 58

7. Application of

H-NMR Spectroscopy (Examples) 58 – 80

Ultraviolet Spectroscopy

Planck’s law:

ΔE = hν = hc / λ

Lambert-Beer’s law:

A = logI

/I = εcλ

Absorption of ultraviolet (UV) or visible (VIS) light results in electronic transitions, promotion of electrons from low-energy ground-state orbitals to higher energy excited-state orbitals (p-p* transition is the most probably).

The UV spectrum spans from 100 nm to 400 nm, while the VIS spectrum ranges from 400 nm (violet) to 750 nm (red).

Molecules that require more energy for electron promotion absorb at shorter wavelenghts. Molecules that require less energy for electron promotion

absorb at longer wavelenghts.

log

5

4

3 2

1

h = E

-E

LUMO HOMO

200 300 400 nm

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Axes of UV-VIS spectra

Bathochromic shift: increasing of λ

Hypsochromic shift: decreasing of λ

Hyperchromic shift: increasing of absorbance (A, loge) Hypochromic shift: decreasing of absorbance (A, loge)

The wavelenght of absorption is usually reported as λ

, the wavelenght at the highest point of the curve. The absorption of energy is recorded as

absorbance (A). Molar absorptivity, or molar extinction coefficient is the

value for e.

bathochromic shift

increasing conjugation

Comparison of UV spectra of compounds with increasing/decreasing conjugation

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hypsochromic shift

decreasing conjugation

bathochromic shift

Comparison of UV spectra of the same compound in different solvents

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hypsochromic shift

Infrared Spectroscopy

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Planck’s law: E = h ν = h c ν *

ν* : wavenumber [cm

]

Transmission: T = I / I

(percent transmission %T)

Nuclei of atoms bonded by covalent bonds undergo vibrations, or

oscillations (Hooke law). When molecules absorb infrared radiation, the absorbed energy causes an increase in the amplitude of the vibrations of the bonded atoms. The molecule is then in an excited vibrational state. The absorbed energy is subsequently dissipated as heat when the molecule returns to the ground state.

Fundamental modes of vibration are the streching vibrations (bond

lenghts change) and the bending vibrations (bond angles change).

Types and shapes of bands

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transmittance %:

wavenumber: the reciprocal of wavelength, in cm-1 or in Kaysers strong

medium weak

sharp

wide

I 100 T I

0

λ

ν

1

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1-BUTANOL

dr. P é ter T é t é nyi

Butanol liquid film, standardized Data operations: Blank, Flat, Abex, Smooth Resolution: 4 cm ^{- 1} Time: 11.51.38

Date: 1996.09.12 butanol6.sp

4400 4000 3500 3000 2500 2000 1800 1600 1400 1200 1000 800 600 450

CM - 1 0

10 20 30 40 50 60 70 80 90 100

aliph C - H aliph C - H aliph C - H aliph C - H

aliph C - C assoc OH

assoc OH

C - O C - O

C H ₃

CH ₂

CH ₂

CH ₂

OH

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Name of the Department: DOC dr. P é ter T é t é nyi

Data operations: Blank, Flat, Abex, Smooth Resolution: 4 cm ^{- 1}

Time: 11.36.36

Date: 1996.09.12 Version ID: Report Builder, Rev. 1.10

brombut6.sp

4400 4000 3500 3000 2500 2000 1800 1600 1400 1200 1000 800 600 450

CM - 1 0

10 20 30 40 50 60 70 80 90 100

%T

aliph C - H aliph C - H

aliph C - C aliph C - Br

1-BROMBUTANE

C H ₃

CH ₂

CH ₂

CH ₂

Br

dr. P é ter T é t é nyi

Allylamine liquid film, standardized - 1

Time: 13.17.16

Date: 1997.01.08 allami6.sp

4400 4000 3500 3000 2500 2000 1800 1600 1400 1200 1000 800 600 450

CM - 1 0

10 20 30 40 50 60 70 80 90 100

%T

assoc N - H as assoc N - H as assoc N - H as

assoc N - H s assoc N - H s assoc N - H s

aliph C - H aliph C - H aliph C - H aliph C - H olefine C - H

olefine C - H _olefine C=C aliph C - N( - H)

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ALLYLAMINE

C H ₂

CH

CH ₂

NH ₂

Name of the Department: DOC dr. P é ter T é t é nyi

2,4 - Pentanedione liquid film, standardized Data operations: Blank, Flat, Abex, Smooth Instrument model: Perkin - Elmer FTIR - 1600

Resolution: 4 cm ^{- 1} Time: 11.07.12

Date: 1996.11.25 Version ID: Report Builder, Rev. 1.10

pendio6.sp

4400 4000 3500 3000 2500 2000 1800 1600 1400 1200 1000 800 600 450

CM - 1 0

10 20 30 40 50 60 70 80 90 100

%T

assoc OH enol assoc OH enol

olefine C - H enol olefine C - H enol

aliph C - H aliph C - H aliph C - H aliph C - H

ketone C=O diketone

ketone C=O

diketone ^ketone C=O C=C enol keto - enol

aliph C - C aliph C - C

diketo - form keto - enol - form

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2,4-PENTANEDIONE

C H₃

C

CH₂ C

CH₃

O O

C H₃

C CH

C

CH₃

O O

H

1 H-NMR spectroscopy

Nuclear magnetic resonance (NMR) spectroscopy provides information about the carbon (

C NMR) and hydrogen (

H NMR) atoms in the

molecule. NMR spectroscopy is based upon the absorption of radio waves by certain nuclei in organic molecules when they are in a strong magnetic field.

The amount of energy required to flip the magnetic moment of a proton from parallel to antiparallel depends, in part, upon the strenght of external magnetic field. If the magnitude of magnetic field is increased, the energy difference between the parallel and antiparallel states increases. If this value is increased, the nucleus is more resistant to being flipped and higher-energy, higher frequenty radiation is required.

For example, a 200 MHz instrument uses a field strenght of about 46 gauss,

B ₀

z

x

y

Superconducting with cryogenic liquids:

generate B

constant magnetic field

Sample tube:

5-20 mg sample soluted in 0,5-1,0 ml of appropriate NMR solvent

Radiofrequency generator:

generate B

radiofrequency field

Detector

reciver coil detects the signal B

Schematic diagram of an NMR instrument

spinmoment

magnetic moment:

: magnetogyric ratio h : Planck constant

I : spin quantum number

I is dependent on number of

protons and neutrons ^I

NMR inactive even even 0

C,

O

NMR active odd odd 1

N

P m

Magnetic properties of nuclei

γP μ

1) 2π I(I

γ h μ

1) 2π I(I

P h

p z

Magnetic behaviour of nuclei in magnetic field

z

B ₀

m : magnetic quantum number

I - ...

1, - I I, m

1 2I

1 2 I

) 2 (

m 1 ( )

1 2

és

p p p

2π γ m h μ _z

Larmor precession

Space quantization of the magnetic moment

B ₀

) 2 ( m 1

) 2 ( m 1

E

0 0

z γB

π m h μ B

E 2

γB 0

π E h

2

γB hν π h 2 0

kT E -

N e

Condition of resonance:

Population of the energy levels:

Frequency of the precession:

γB 0

(Larmor frequency)

4,7 T 9,4 T E

B

200 MHz 400 MHz

γB hν π h 2 0

γB 0

π E h

2

B ₀

Effect of electromagnetic field and origin of spectrum

• it puts as much magnetization as possible in the

<xy> plane Pulse /2:

• it has the effect of inverting the populations Pulse :

z’

y’

x’

B ₀

B

time frequency

y’ y x’ x z’ z

FID (Free Induction Decay) NMR spectrum

FT

Fourier transformation

B ₀ ^{>> B}

Axes of ¹ H-NMR spectra

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absorbance, A

scale

ppm 0

10 (TMS)

6 instrument

TMS X X

ν 10

ν

δ ν

3

J

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Chemical shift

The local magnetic field B „sensed” by a proton is different from the external magnetic field B

provided by the spectrometer. The reason is that electrons circulating in the vicinity of a proton extert their own magnetic fields that oppose the external field. Because electrons are the source of this shielding, it follows that the more electron density there is near a proton, the greater will be the shielding. Electronegative groups near a proton pull electons away and decrease the shielding of a proton. Electropositive groups near a proton, in contrast, increase the surrounding electron density and increase the shielding of the proton.

γB hν π h

0 2

0 σB B

10 B 5 0 (ppm)

Decreasing electrondensity Less shielded nucleus Greater chemical shift (paramagnetic shift)

Deshielding effect

Higher frequenty (low-/downfield)

Increasing electrondensity More shielded nucleus Smaller chemical shift

(diamagnetic shift) Shielding effect

Lower frequenty (high-/upfield)

Chemical shift

The signals are not singlets, but multiplets having fine structures.

The phenomenon is the consequence of spin-spin coupling and characterized by the spin-spin coupling constant, J [Hz]. Each interacting nucleus splits the signal into two peaks. If the number of interacting nuclei is n, the signal is split into 2n peaks.

signal of X hydrogen in the absence of coupling partner signal of X hydrogen in the presence of A coupling partner

3

J

Splitting / Coupling

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Signal of X hydrogen

Signal of A hydrogen AX spin system

A

X

Spin system means an arrangement of nuclei being in spin-spin interaction with each other (several spin systems can be present within one molecule).

Splitting / Coupling

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First-order spectrum: distance (in Hz) of the signals of nuclei coupling with each other is considerably larger than the distance of peaks formed by splitting (Δν/J > 10), as well as the chemically equivalent groups of nuclei are magnetically equivalent, too: A

X

spin system

• when two protons have the same chemical shift, no splitting is observed

• in case of first order spin system, splitting caused by equivalent nuclei is the same (coupling constants are the same), that’s why the splitting pattern of an X nucleus (that consists of 2n lines, principally) is simplified into n+1 peaks (2nI+1 rule)

• intensity ratio of the peaks within the signal is given by the corresponding (n+1) row of the Pascal triangle

• within the signal, the distance (in Hz) of either two adjacent peaks gives the value of the coupling constant

•

3 3 1 1

A

X

Splitting / Coupling

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Spin-spin splitting

Number of hydrogen multiplicity attached to the neighboring carbons

0 singlet

1 doublet

2 triplet

3 quartet

4 quintet

5 sextet

more than 5 multiplet

Higher-order spectrum: there is a spin-spin coupling between nuclei the chemical shift values of which are close to each other (Δν/J < 10), and/or the molecule contains nuclei that are magnetically non- equivalent: AB, AA’BB’ spin systems

The coupling constant cannot be determined directly; the spectrum can be analysed with difficulties.

Splitting / Coupling

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F

F

H

H

Chemically equivalent nuclei can be transformed into each other by means of any symmetry operation (rotation and/or reflection) within the molecule (their situation to the symmetry elements is the same), so the transformed molecule is indistinguishable from the original one.

F

F

H

H

Magnetically equivalent nuclei are chemically equivalent. Each member of their group has the same spin-spin interaction with each member of another group of chemically equivalent nuclei.

• No symmetry: diastereotopic

• Axis of symmetry: homotopic

• Plane of symmetry: enantiotopic Cl

H

Cl H

Br H

Cl

H

J

= J

J

≠ J

Splitting / Coupling

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AB

AX A

2

AB

A

2

AX

A

X

3

J

J ≠ f(B

) Δ = f(B

)

X A

Signal of X hydrogen without spin-spin coupling

Signal of A hydrogen without spin-spin coupling

Splitting / Coupling

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8 7 6 5 4 3 2 1 0 ppm Cl₂HC

O

OCH₃

Integrated intensity

Magnitude of the area under the resonance signal is proportional to the number of nuclei participating in the resonance. Relative number of nuclei in different chemical environment can be determined.

8 7 6 5 4 3 2 1 0 ppm

H₂C O

OCH₃

O

OCH₃

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7 6 5 4 3 2 1 0

3H

2H

3H

3H

3H

2H H

CH

C C

OCH

O

H

C C

OCH

CH

O

ppm

t q s

s q t

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1.0436 2.0552 4.0601 3.0012

Integral 3.8395 3.4748 1.4576 1.4177 1.3838 1.3519 1.3081 1.2702 1.2303 1.1944 0.8436 0.8077 0.7719

( p p m )

0 . 5 1 . 0

1 . 5 2 . 0

2 . 5 3 . 0

3 . 5 4 . 0

AM 200; BUTANO L; H- 1 CDCL3; 99. 02. 08

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2.0001 2.0231 2.0664 3.0845

Integral 3.4369 3.4030 3.3691 1.8662 1.8323 1.7944 1.7605 1.5114 1.4715 1.4356 1.3997 0.9592 0.9214 0.8855

0 . 5 1 . 0

1 . 5 2 . 0

2 . 5 3 . 0

3 . 5 4 . 0

AM 200; NT/ 1- BRO M - BUTAN; H- 1 CDCL3; 2000. 07. 10

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13 C-NMR spectroscopy

H₃C CH2

H b a

H₃C CH₂

CH3

b a

b

H3C CH2

C O OH b a

H₃C CH₂ Cl b a

H₃C CH2

NH2

b a

H₃C CH₂ F b a

H3C CH2

OH b a

b ppm 8.4 15.4 9.0

19.0

18.7

17.9 14.0

a ppm 8.4 15.9 27.8

36.9

38.9

57.3 79.3

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Axes of ¹³ C-NMR spectra

200

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absorbance, A

scale

ppm 0

10 (TMS)

TMS 6 X X

ν 10

ν

δ ν

Advantages of ¹³ C-NMR

• Provides structural information on the carbon skeletone directly.

• Functional groups having no H-s (e.g., C=O, C≡N or similar groups) result in chemical shifts.

• There are no overlapping chemical shifts usually.

• Couplings do not modify the spectra.

• More information could be gained on stereostructure of the compound ( space effect)

Disadvantages of ¹³ C-NMR

• It has considerably less sensitivity, compared to

H-NMR.

• Heteronuclear coupling decreases further the signal/noise ratio.

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γ Space effect

a b

a b

b

b a

a

a ppm b ppm 23.1-6.8=16.3 18.8-2.8=16.0

18.8-2.8=16.0 18.8-2.8=16.0

18.8+2.0=20.8 18.8+2.0=20.8

23.1+0.0=23.1 18.8+0.0=18.8

c ppm d ppm

c d

c d

c d

c d

27.1+1.4+9.0- -3.4=34.1

27.1+1.4+5.4=

=33.9

27.1+1.4-6.4=

=22.1

27.1+1.4-0.2=

=28.3

27.1+6.0+

+5.4-2.9=

=35.6

27.1+1.4+5.4=

=33.9

27.1+1.4-6.4=

=22.1

27.1+6.0-0.1=

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1-BUTANOL

1-BUTANOL ^OH

a b

c d

a: 61.6, t b: 34.6, t c: 18.8, t

d: 13.6, q a b

c d

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77.6760 77.0400 76.4040 34.7526 33.5249 21.2780 13.1134

1 0 2 0

3 0 4 0

5 0 6 0

7 0 8 0

AM 200; NT/ 1- BRO M - BUTAN; C- 13 CDCL3; 2000. 07. 10

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BUTYL ACETATE BUTYL ACETATE

a b

c d

e H ₃ C C f

O O

b: 170.3, s c: 64.2, t d: 30.6, t a: 20.7, q e: 18.0, t f: 13.6, q b

c

d

a e

f

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13 C spectrum recorded by proton decoupling technique

• Hydrogen nuclei are continuously irradiated by another electromagnetic source during the

C recording, for saturation of the hydrogen nuclei.

• The spectrum consists of only singlets, because

H-

C spin- spin interactions have been disappeared.

• Intensity of the chemical shift of carbons without hydrogens is increased due to the nuclear Overhauser effect (nOe).

• Overlapping of chemical shifts seldom happens.

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DEPT spectrum

13 C NMR spectrum with ¹ H decoupling

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Mass spectrometry

Mass spectrometry (MS) is a technique for measuring the mass, and therefore the molecula weight, of a molecule. It is also possible to gain structural information about a molecule by measuring the masses of fragments produced when molecules are broken.

A small amount of sample is vaporized into the mass spectrometer, where it is bombarded by a stream of high-energy electrons (about 70 electron volts).

When it strikes an organic molecule, it dislodges a valence electron from the molecule, producing a cation radical-cation because the molecule has lost an electron and has an odd number of electrons.

Separation of ionic particle (fragments) by electric and/or magnetic field.

The continuous measuring of the intensity of the ion beam results in mass

spectrum. Mass spectra of different organic compounds are always

• EI: Electron-impact Ionization

(hard electron beam, ionisation in gas phase)

• CI: Chemical Ionization

(soft ionisation in gas phase)

• FAB: Fast Atom Bombardment (soft ionisation in liquid phase)

• ESI: Electrospray Ionization

(thermal ionisation for macromolecules)

• MALDI: Matrix Assisted Laser Desorption Ionisation (photolytic ionisation for macromolecules)

• APCI: Atmospheric Pressure Chemical Ionisation

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Ionisation methods

Double-focusing mass spectrometer

E

B Magnetic induction Electric

gradient

2 zU mv ²

r zvB mv ² 2U

B r z

m ^{2 2}

Ion source

Analysator

Detector

2-Methylbutan-1-al

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1-Butanol

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n-Butyl acetate

X-ray Crystallography

1895. Wilhelm Konrad Röntgen: discovering the X-ray (1901. Nobel Prize) 1912. Max von Laue: diffraction of X-ray

Electromagnetic radiation, arising from the nucleus

l: 0,1-100 Å = 0,01-10 nm (crystall-lattice distances: 1-20 Å = 0,1-2 nm) E

: 100 eV – 100 keV

Applications of X-ray:

X-ray radiology (medicinal relation) X-ray crystallography

single crystall technique (Laue methodology)

powder diffractometry (Debye-Scherrer methodology)

Origin of X-ray:

by X-ray tube or by synchrotron radiation Types of X-ray:

X-ray photon-generating effect, i.e.

„Bremsstrahlung” – this radiation is produced by high energy electrons progressively decelerated by the material of the anode.

Characteristic radiation – each element has electronic orbitals of characteristic energy.

Removal of an inner electron by an energetic

photon provided by a primary radiation source, an

electron from an outer shell drops into its place.

Simplified structure of an X-ray tube

C

W

W

X

A K

U

U

- +

Interaction between X-ray and to be analysed crystall

X-ray crystallograpy is based on the fact that the spacing between atoms (100 pm – 300 pm) is similar in magnitude to the wavelenght of X-rays.

The electron clouds of the atoms scatter the X-rays, with the degree of

scattering proportional to the atomic number. The scattering pattern from a single molecule is too weak to be detected, therefore it must be amplified and able to be detected.

incoming ray

optical grid

diffracted rays

diffraction

Diffraction pattern for a crystall

d can be calculated, size of elementar cell can be determined

Bragg diffraction occours when the waves undergo interference in accordance to Bragg’s law:

for a crystalline solid the waves are scattered from the lattice planes separated by the interplanar distance d. The waves remain in phase since the path lenght of each wave is equal to an integer multiple of the wavelenght.

Θ Θ

λ

d

Lattice Planes Bragg’s Law

d

θ θ

·

· x

x/d=sin θ x=d.sin θ 2x=2d.sin θ 2x=n.λ

n.λ=2d.sin θ

Bragg’s law

E=z.U E=h.c/λ

λ=h.c/z.U

Duane-Hunt’s law

d=n.h.c/(z.2.U.sin θ)

N N

O H₃C

NO₂ N O

1

N N

NO₂

O H₃C N

O

2

Structural Formula of 1 and 2 and their ORTEP (Oak Ridge Thermal

Ellipsoid Plot) figure

Application of ¹ H-NMR spectroscopy (Examples)

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8 7 6 5 4 3 2 1 0 ppm

t

qi sx

t

2H

2H 2H

3H CH

CH

CH

CH

Br

•

H-NMR spectrum of 1-bromobutane

Molecular symmetry and rotation

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t

sx

d

m (qi)

1H

2H

3H

3H

CH₃CHCH₂CH₃ Br

•

H-NMR spectrum of 2-bromobutane

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12 11 10 9 8 7 6 5 4 3 2 1 0 ppm

m

d

s d

6H

5H

2H 2H 1H

•

H-NMR spectrum of 1-phenyl-4-methylpentan-2-one

C CH₂ O

CH CH₃

CH₃ CH₂

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• A flask contains a liquid with the label:

‘...ropyl acetate’

3H

2H

2H

3H C

O

H

C O CH CH

CH

C

O

H

C O CH

CH

CH

t

sx

• Possible structures:

• The recorded

H-NMR spectrum:

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•

H-NMR spectrum of normal and isopropyl acetate

5 4 3 2 1 0

ppm C

O

H₃C O CH₂CH₂CH₃

3H 2H

2H

3H

C O

H₃C O CH CH₃

CH₃

5 4 3 2 1 0

ppm

3H

6H

sp

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• Benzene was alkylated with 1-chloropropane under Friedel-Crafts conditions,

H-NMR spectrum of the recorded main product:

6H 1H

CH CH₃

CH₃

sp

d

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CH

CH

CH

8 7 6 5 4 3 2 1 0

ppm

3H 2H

2H

t sx

t

•

H-NMR spectrum of the side-product:

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1H 2H

2H

3H

t

sx td

t

•

H-NMR spectrum of butyraldehide

C O

H

CH₂CH₂CH₃

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8 7 6 5 4 3 2 1 0

ppm

3H

1H 1H

1H

dd

dd dd

•

H-NMR spectrum of vinyl acetate

C C

H H

H O

C H₃C

O

X A

M

AMX

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3H

dd

dd dd

•

H-NMR spectrum of ethyl vinyl ether

C C

H H

H O

CH₃CH₂

1H 1H 1H

2H

X A

M

AMX

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12 11 10 9 8 7 6 5 4 3 2 1 0 ppm

•

H-NMR spectrum of E-hex-2-en-1-al

3H 2H 2H

1H 1H 1H

C C

H CH₂CH₂CH₃ H

C O

H

dd dt ddt qd

sx

t

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1H

1H 2H

2H

OH

•

H-NMR spectrum of phenol

AA’MM’X A

M

A’

M’

X X MM’

AA’

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12 11 10 9 8 7 6 5 4 3 2 1 0 ppm

1H 2H

NO

OH

O

N

AX

• Phenol was nitrated with nitric acid, two products were isolated.

1

H-NMR spectrum of one of them:

d

t

A X X

A X

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OH

NO₂

NO₂

1H 1H 1H

dd d

d

•

H-NMR spectrum of the other one:

A

M X

AMX A

M X

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12 11 10 9 8 7 6 5 4 3 2 1 0 ppm

1H 1H

1H 1H

1H

AMPX

NO

OH

• Nitration under milder conditions resulted also in two products.

1

H-NMR spectrum of the major product:

dd

dd ddd (td)

ddd (td)

A M

P X

M A P

X

semmelweis-egyetem.hu

•

H-NMR spectrum of the side-product:

2H

2H 1H

OH

NO₂

AA’XX’

A A’

X’

X

XX’ AA’

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12 11 10 9 8 7 6 5 4 3 2 1 0

ppm

1H

1H 1H 2H

•

H-NMR spectrum of the 3-nitrophenol

OH

NO₂

AMPX P

M A

X ddd

t

ddd

t

P A

X

M

semmelweis-egyetem.hu

•

H-NMR spectrum of phenylalanine

•

H-NMR spectrum of alanine

12 11 10 9 8 7 6 5 4 3 2 1 0

1H

ppm

3H

1H

2H

CH H₃C

NH₂ COOH

CH H₂C

NH₂ COOH

A

X

ABX

homotopic hydrogens

diastereotopic hydrogens

semmelweis-egyetem.hu

12 11 10 9 8 7 6 5 4 3 2 1 0 ppm

1H

2H CH

H

C

NH

COOH

AB

stereogenic center

diastereotopic hydrogens: different chemical shift

NH

HOOC

H

Ph H

H NH

HOOC H

H H

Ph NH

HOOC H

H Ph

H

X

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*

A

XY

O H

C O

H CH

CH

H H

H H

A

XY

•

H-NMR spectrum of acetaldehyde diethyl acetal

plane of symmetry

diastereotopic hydrogens:

different chemical shift

A

X σ

X

A

1H

3H

6H

2H 2H

semmelweis-egyetem.hu

8 7 6 5 4 3 2 1 0

• 1H-NMR spectrum of 2,4-dimethylpentanedioic acid

σ

HOOC COOH

H

H

C H CH

H

H

C

two stereogenic center plane of symmetry

chemically non-equivalent hydrogens (diastereotopic)

different chemical shift

two stereogenic center symmetry axis

chemically equivalent hydrogens (homotopic)

same chemical shift

HOOC COOH

CH

H H CH

H

H A

X

R S

S A

M

XY S

A

X(Y) (X)Y

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