Spektroskopi UV-Vis

The Electromagnetic Spectrum

 Atoms produce sharp lines of absorption

Molecules have many excited modes of vibration and rotation at
room temperature. The rotational and vibrational levels are
superimposed on the electronic levels

Electron transitions may occur from any of several vibrational

and rotational states of one electronic level to any of several
vibrational and rotational states of a higher electronic level.
Thus, the UV spectrum of a molecule consists of a broad band of
absorption centered near the wavelength of the major transition

Internal Energy of Molecules

Etotal=Etrans+Eelec+Evib+Erot+Enucl
Eelec: electronic transitions (UV, X-ray)
Evib: vibrational transitions (Infrared)
Erot: rotational transitions (Microwave)
Enucl: nucleus spin (nuclear magnetic
resonance) or (MRI: magnetic resonance
imaging)

Latar Belakang spektroskopi UV/Vis

Dasar penyerapan sinar UV
Spektroskopi Ultraviolet/visible spectroscopy
melibatkan penyerapan sinar ultraviolet/visible
oleh molekul sehingga menyebabkan terjadinya
perpindahan elektron dari keadaan dasar (ground
state) ke keadaan elektron tereksitasi.
Sinar Ultraviolet/Visible

Panjang gelombang (l) antara 190 dan 800 nm

Why should we learn this stuff?

After all, nobody solves structures with
UV any longer!
Many organic molecules have chromophores that absorb UV
UV absorbance is about 1000 x easier to detect per mole than NMR
Still used in following reactions where the chromophore changes. Useful
because timescale is so fast, and sensitivity so high. Kinetics, esp. in
biochemistry, enzymology.
Most quantitative Analytical chemistry in organic chemistry is conducted using
HPLC with UV detectors
One wavelength may not be the best for all compound in a mixture.

Affects quantitative interpretation of HPLC peak heights

Tipe transisi
Ada beberapa tipe transisi elektronik pada molekul organik yaitu:
s to s* (alkana)
s to p* (senyawa karbonil)
p to p* (alkena, senyawa karbonil, alkuna, senyawa azo)
h to s* (oksigen, nitrogen, sulfur, and senyawa halogen )
h to p* (senyawa karbonil)
Gambar 1. Tipe transisi elektronik

Transisi elektronik
UV/Vis

Vacuum UV or Far UV
(<190 nm

 Thus, UV / Visible spectra often called Electronic Spectra

General energy differences of 30 150 kcal/mole
For most molecules, the lowest energy orbitals are the
sigma (s) - orbitals (s - bonds)
The electrons of sigma bonds are too tightly bound to be
promoted by radiation in the 200-700 nm region.
Therefore alkanes, saturated alcohols, simple alkenes show no or
very little UV absorption.

p- orbitals are at somewhat higher energy levels

Non-bonding (n-orbitals), i.e., unshared electron pairs, occur at
even higher energies
Unoccupied or anti-bonding orbitals (p* and s*) have the
highest energy
9

Contoh eksitasi elektron pada spektra

Transisi dari orbital molekul
tertinggi yang terisi (HOMO) ke
orbital molekul terendah yang
terisi (LUMO) memerlukan jumlah
energi yang paling sedikit
sehingga hal inilah yang biasanya
terjadi.

Beberapa istilah pada Spektroskopi UV/Vis

Kromofor

adalah atom atau gugusan atom yang menyerap

sinar UV-Vis baik itu menghasilkan warna atau tidak
Auksokrom adalah gugusan atom yang dapat memperpanjang
konjugasi pada kromofor (meningkatkan intensitas,
kadang sekaligus panjang gelombang)
(substituen dg pasangan elektron bebas OH, NH, SH)
Pergeseran batokromik (pergeseran merah ) adalah
pergeseran ke arah panjang gelombang yang lebih
panjang
Pergeseran hipsokromik (pergeseran biru) adalah pergeseran
ke arah panjang gelombang yang lebih pendek
Efek hiperkromik adalah efek yang dapat menaikan
intensitas serapan
Efek hipokromik adalah efek yang dapat menurunkan
intensitas serapan

Terminologi pergeseran lmax

12

Conjugated Unsaturated Systems

Conjugated unsaturated systems are molecules with two

or more double or triple bonds each alternating with a
single bond.
Conjugated unsaturated systems are species that have
delocalized p bonds, i.e. a p-orbital on an atom adjacent
to a double bond producing p p* transitions.
Single electron as in the allyl radical (CH2=CHCH2)
Vacant p orbital as in allyl cation (CH2=CHCH2+)
P orbital of another double bond (CH2=CH CH=CH2
Compounds whose molecules contain conjugated
multiple bonds have absorption maxima at wavelengths
longer than 200 nm, i.e. in the UV range.
Conjugated systems absorb strongly in the UV / Visible
portion of the electromagnetic spectrum, therefore they
can be investigated with Ultraviolet Spectroscopy.
13

14

Pengaruh konjugasi
E

E = h

l max

162

217

258

296
15

16

 More complicated alkenes (carbon-carbon double bond) and

nonconjugated dienes usually have absorption maxima below
200 nm, i.e. do not absorb in the UV range.
Carbonyl Compounds
Compounds with carbon-oxygen double bonds (carbonyl) also
absorb light in the UV region.
The carbonyl excitation process involves movement of an
electron from one of the unshared (nonbonding) pair to the p*
orbital of the carbon-oxygen double bond.
Transitions - n p*
Non-bonding electrons, such as those in a carbonyl group
(and some alkyl halides), will absorb in the UV region, but at
lower intensity than conjugated systems.
Carbonyl absorption in the UV does not require additional
conjugation in the molecule.
If a molecule does not absorb in the UV, then it does not
contain a conjugated system of alternating double bonds or a
17
carbonyl group

Hukum dasar pada UV/Vis Spektroskopi

Hukum Lambert-Beer
Spektra UV dari suatu senyawa biasanya diperoleh dengan
melewatkan cahaya pada panjang gelombang tertentu (sinar
monokromatik) melalui larutan encer pada senyawa di
dalam pelarut yang tidak dapat memberikan serapan.
Intensitas serapan diukur dengan persentase dari cahaya yang
masuk melalui sampel sehingga melewati sampel
% Transmitansi = (I / I0) * 100%
Dimana :

I = intensitas of cahaya yang diteruskan

I0 = intensitas of cahaya yang diberikan

Karena serapan cahaya merupakan fungsi konsentrasi dari

penyerapan molekul, maka cara yang lebih tepat untuk menyatakan
intensitas serapan adalah dengan menggunakan hukum LambertBeer.
Absorbance = -log(I / I0) = cl
dimana
= absorptivitas molar
c = konsentrasi molar zat terlarut
l = panjang sel sampel (cm)

 Molar Absorptivity () - also called the Molar Extinction

Coefficient - is a measure of the strength or intensity of
the absorption.
It is the proportionality constant relating the observed
absorbance (A) at a particular wavelength to the molar
concentration (C) of the sample and the length (l) of the
path of the light beam through the sample cell.
A =

C l

= A / (C l )

Data Example:
2,5-dimethyl-2,4-hexadiene (in methanol)
Wavelength of Maximum Absorbance (lmax) 242.5 nm
Molar Absorptivity ( ) 13,100 M-1 cm-1 (Log = 4.1)

20

Conjugated systems show large values of

= 1000 100,000 (Log

= 3 5)

Carbonyl compounds show smaller values of

= 10 100

Electronic
Structure

p and n

(Log
Example

= 1 - 2)

Transition

max

Log

257
279

486
15

2.7
1.2

210
315

11,500
14

4.1
1.1

217
258

21,000
35,000

4.3
4.5

ca 200
255

8,000
215

3.9
2.3

lmax
(nm)

n-butyl iodide
acetone

p - p and n Acrolein
(propenal)
(C=C & C=O)

n s*
n p*
p p*
n p*

p-p

1,3-butadiene
p p*
1,3,5-hexadiene p p*

aromatic p

benzene

Ar p p*
Ar p p*

21

Analisis struktur dengan UV-Vis

Tentukan harga lmax

Hitung harga
Bandingkan harga lmax teoritis dengan
hasil empiris

22

Pemilihan pelarut
Tabel 1. Pelarut yang umum digunakan dan panjang gelombang
minimum yang dihasilkan dari pengukuran
menggunakan kuvet setebal 1 cm
Pelarut

l minimum (nm) l cut-off (nm)

asetonitril

190

210

air

191

205

sikloheksana

195

210

hexane

195

210

metanol

201

210

etanol

204

210

Dietil eter

215

210

diklorometana

220

235

kloroform

237

245

karbon tetraklorida

257

265

UV Spectroscopy

III. Chromophores
C. Substituent Effects
General from brief study of these general chromophores, only the weak n
p* transition occurs in the routinely observed UV
The attachment of substituent groups (other than H) can shift the energy
of the transition
Substituents that increase the intensity and often wavelength of an
absorption are called auxochromes
Common auxochromes include alkyl, hydroxyl, alkoxy and amino groups
and the halogens

24

UV Spectroscopy

III. Chromophores
C. Substituent Effects
General Substituents may have any of four effects on a chromophore
i. Bathochromic shift (red shift) a shift to longer l; lower energy
ii.

Hypsochromic shift (blue shift) shift to shorter l; higher energy

iii.

Hyperchromic effect an increase in intensity

Hypsochromic

Bathochromic
Hypochromic

200 nm

Hyperchromic

iv. Hypochromic effect a decrease in intensity

700 nm
25

UV Spectroscopy

III. Chromophores
C. Substituent Effects
1. Conjugation most efficient means of bringing about a bathochromic and
hyperchromic shift of an unsaturated chromophore:
H2C

lmax nm

CH2

175

15,000

217

21,000

258

35,000

465

125,000

-carotene

n p* 280

12
900

n p* 280

27
7,100

p p* 189
O

p p* 213

26

UV Spectroscopy

This quantification is referred to as the Woodward-Fieser Rules which we will apply

to three specific chromophores:
1. Conjugated dienes
2. Conjugated dienones
3. Aromatic systems

lmax = 239 nm

27

UV Spectroscopy

IV. Structure Determination

A. Dienes
1. General Features
For acyclic butadiene, two conformers are possible s-cis and s-trans

s-trans

s-cis

The s-cis conformer is at an overall higher potential energy than the strans; therefore the HOMO electrons of the conjugated system have less
of a jump to the LUMO lower energy, longer wavelength

28

UV Spectroscopy

IV. Structure Determination

A. Dienes
1. General Features
Two possible p p* transitions can occur for butadiene Y2 Y3* and Y2
Y4*
Y4 *
Y3*

s-trans

Y2

175 nm forb.
217 nm

175 nm
253 nm

s-cis

Y1

The Y2 Y4* transition is not typically observed:

The energy of this transition places it outside the region

typically observed 175 nm
For the more favorable s-trans conformation, this transition is
forbidden
The Y2 Y3* transition is observed as an intense absorption

29

UV Spectroscopy

IV. Structure Determination

A. Dienes
1. General Features
The Y2 Y3* transition is observed as an intense absorption ( =
20,000+) based at 217 nm within the observed region of the UV
While this band is insensitive to solvent (as would be expected) it is
subject to the bathochromic and hyperchromic effects of alkyl
substituents as well as further conjugation
Consider:

lmax = 217

253

220

227

227

256

263

nm

30

UV Spectroscopy

IV. Structure Determination

A. Dienes
2. Woodward-Fieser Rules
Woodward and the Fiesers performed extensive studies of terpene and
steroidal alkenes and noted similar substituents and structural features
would predictably lead to an empirical prediction of the wavelength for
the lowest energy p p* electronic transition
This work was distilled by Scott in 1964 into an extensive treatise on the
Woodward-Fieser rules in combination with comprehensive tables and
examples (A.I. Scott, Interpretation of the Ultraviolet Spectra of Natural
Products, Pergamon, NY, 1964)

31

UV Spectroscopy

IV. Structure Determination

A. Dienes
2. Woodward-Fieser Rules - Dienes
The rules begin with a base value for lmax of the chromophore being
observed:
acyclic butadiene = 217 nm
The incremental contribution of substituents is added to this base value
from the group tables:
Group

Increment

Extended conjugation

+30

Each exo-cyclic C=C

+5

Alkyl

+5

-OCOCH3

+0

-OR

+6

-SR

+30

-Cl, -Br

+5

-NR2

+60
32

UV Spectroscopy

IV. Structure Determination

A. Dienes
2. Woodward-Fieser Rules - Dienes
For example:
Isoprene - acyclic butadiene =
one alkyl subs.
Experimental value

Allylidenecyclohexane
- acyclic butadiene =
one exocyclic C=C
2 alkyl subs.
Experimental value

217 nm
+ 5 nm
222 nm
220 nm

217 nm
+ 5 nm
+10 nm
232 nm
237 nm

33

UV Spectroscopy

IV. Structure Determination

A. Dienes
3. Woodward-Fieser Rules Cyclic Dienes
There are two major types of cyclic dienes, with two different base values
Heteroannular (transoid):

= 5,000 15,000
base lmax = 214

Homoannular (cisoid):

= 12,000-28,000
base lmax = 253

The increment table is the same as for acyclic butadienes with a couple
additions:
Group

Additional homoannular

Increment
+39

Where both types of diene

are present, the one with
the longer l becomes the
base
34

UV Spectroscopy

IV. Structure Determination

A. Dienes
3. Woodward-Fieser Rules Cyclic Dienes
In the pre-NMR era of organic spectral determination, the power of the
method for discerning isomers is readily apparent
Consider abietic vs. levopimaric acid:

C OH
O

abietic acid

C OH
O

levopimaric acid

35

UV Spectroscopy

IV. Structure Determination

A. Dienes
3. Woodward-Fieser Rules Cyclic Dienes
For example:
1,2,3,7,8,8a-hexahydro-8a-methylnaphthalene
heteroannular diene =
214 nm
3 alkyl subs. (3 x 5)

+15 nm

1 exo C=C

+ 5 nm
234 nm

Experimental value

235 nm

36

UV Spectroscopy

IV. Structure Determination

A. Dienes
3. Woodward-Fieser Rules Cyclic Dienes

C OH
O

abietic acid

C OH
O

levopimaric acid

heteroannular diene =

214 nm

4 alkyl subs. (4 x 5)
1 exo C=C

+20 nm
+ 5 nm

239 nm

homoannular diene =

253 nm

4 alkyl subs. (4 x 5)

+20 nm

273 nm
37

UV Spectroscopy

IV. Structure Determination

A. Dienes
3. Woodward-Fieser Rules Cyclic Dienes
Be careful with your assignments three common errors:
R

This compound has three exocyclic

double bonds; the indicated bond is
exocyclic to two rings

This is not a heteroannular diene; you would

use the base value for an acyclic diene

Likewise, this is not a homooannular diene;

you would use the base value for an acyclic
diene
38

UV Spectroscopy

IV. Structure Determination

B. Enones
1. General Features
For auxochromic substitution on the carbonyl, pronounced hypsochromic
shifts are observed for the n p* transition (lmax):
O
H

293 nm

O
CH3

279

Cl

235

O
NH2
O
O
O

214
204
204

This is explained by the inductive withdrawal

of electrons by O, N or halogen from the
carbonyl carbon this causes the n-electrons
on the carbonyl oxygen to be held more
firmly
It is important to note this is different from
the auxochromic effect on p p* which
extends conjugation and causes a
bathochromic shift
In most cases, this bathochromic shift is not
enough to bring the p p* transition into
the observed range

OH

39

UV Spectroscopy

IV. Structure Determination

B. Enones
2. Woodward-Fieser Rules - Enones

a
C C C
O

Group

d g a
d C C C C C
O
Increment

6-membered ring or acyclic enone

Base 215 nm

5-membered ring parent enone

Base 202 nm

Acyclic dienone

Base 245 nm

Double bond extending conjugation

30

Alkyl group or ring residue

a, , g and higher

10, 12, 18

-OH

a, , g and higher

35, 30, 18

-OR

a, , g, d

35, 30, 17, 31

a, , d

-Cl

a,

15, 12

-Br

a,

25, 30

95

-O(C=O)R

-NR2
Exocyclic double bond

Homocyclic diene component

39
40

UV Spectroscopy

IV. Structure Determination

B. Enones
2. Woodward-Fieser Rules - Enones
Aldehydes, esters and carboxylic acids have different base values than
ketones
Unsaturated system

Base Value

Aldehyde

208

With a or alkyl groups

220

With a, or , alkyl groups

230

With a,, alkyl groups

242

Acid or ester
With a or alkyl groups

208

With a, or , alkyl groups

217

Group value exocyclic a, double bond

+5

Group value endocyclic a, bond in 5

or 7 membered ring

+5

41

UV Spectroscopy

IV. Structure Determination

B. Enones
2. Woodward-Fieser Rules - Enones
Unlike conjugated alkenes, solvent does have an effect on lmax
These effects are also described by the Woodward-Fieser rules
Solvent correction
Water

Increment
+8

Ethanol, methanol

Chloroform

-1

Dioxane

-5

Ether

-7

Hydrocarbon

-11

42

UV Spectroscopy

IV. Structure Determination

B. Enones
2. Woodward-Fieser Rules - Enones
Some examples keep in mind these are more complex than dienes
cyclic enone =
215 nm
O
2 x - alkyl subs. (2 x 12) +24 nm
239 nm
Experimental value

238 nm

cyclic enone =
extended conj.
-ring residue
d-ring residue
exocyclic double bond

215 nm
+30 nm
+12 nm
+18 nm
+ 5 nm
280 nm

Experimental

280 nm

43

UV Spectroscopy

IV. Structure Determination

C. Aromatic Compounds
2. Substituent Effects
a. Substituents with Unshared Electrons

pH can change the nature of the substituent group

deprotonation of oxygen gives more available n-pairs,

lowering transition energy

protonation of nitrogen eliminates the n-pair,

raising transition energy
Primary

Secondary

Substituent

lmax

lmax

-H

203.5

7,400

254

204

-OH

211

6,200

270

1,450

-O-

235

9,400

287

2,600

-NH2

230

8,600

280

1,430

-NH3+

203

7,500

254

169

-C(O)OH

230

11,600

273

970

-C(O)O-

224

8,700

268

560
44

UV Spectroscopy

IV. Structure Determination

C. Aromatic Compounds
2. Substituent Effects
b. Substituents Capable of p-conjugation

When the substituent is a p-chromophore, it can interact with

the benzene p-system

With benzoic acids, this causes an appreciable shift in the

primary and secondary bands

For the benzoate ion, the effect of extra n-electrons from the
anion reduces the effect slightly
Primary

Secondary

Substituent

lmax

lmax

-C(O)OH

230

11,600

273

970

-C(O)O-

224

8,700

268

560

45

UV Spectroscopy

IV. Structure Determination

C. Aromatic Compounds
2. Substituent Effects
c. Electron-donating and electron-withdrawing effects

No matter what electronic influence a group exerts, the

presence shifts the primary absorption band to longer l

Electron-withdrawing groups exert no influence on the

position of the secondary absorption band

Electron-donating groups increase the l and of the

secondary absorption band

46

UV Spectroscopy

IV. Structure Determination

C. Aromatic Compounds
2. Substituent Effects
c. Electron-donating and electron-withdrawing effects
Primary
lmax

lmax

-H

203.5

7,400

254

204

-CH3

207

7,000

261

225

-Cl

210

7,400

264

190

-Br

210

7,900

261

192

-OH

211

6,200

270

1,450

-OCH3

217

6,400

269

1,480

-NH2

230

8,600

280

1,430

-CN

224

13,000

271

1,000

C(O)OH

230

11,600

273

970

-C(O)H

250

11,400

-C(O)CH3

224

9,800

-NO2

269

7,800

Electron donating

Substituent

Electron withdrawing

Secondary

47

UV Spectroscopy

IV. Structure Determination

C. Aromatic Compounds
2. Substituent Effects
d. Di-substituted and multiple group effects

With di-substituted aromatics, it is necessary to consider both

groups

If both groups are electron donating or withdrawing, the

effect is similar to the effect of the stronger of the two groups
as if it were a mono-substituted ring

If one group is electron withdrawing and one group electron

donating and they are para- to one another, the magnitude of
the shift is greater than the sum of both the group effects

Consider p-nitroaniline:
O
H2N

O
H2N

N
O
48

UV Spectroscopy

IV. Structure Determination

C. Aromatic Compounds
2. Substituent Effects
d. Di-substituted and multiple group effects

If the two electonically dissimilar groups are ortho- or metato one another, the effect is usually the sum of the two
individual effects (meta- no resonance; ortho-steric hind.)

For the case of substituted benzoyl derivatives, an empirical

correlation of structure with observed lmax has been
developed

This is slightly less accurate than the Woodward-Fieser rules,

but can usually predict within an error of 5 nm
O

G
49

UV Spectroscopy

IV. Structure Determination

C. Aromatic Compounds
2. Substituent Effects
d. Di-substituted and multiple group effects

Parent Chromophore

lmax

R = alkyl or ring residue

246

R=H

250

R = OH or O-Alkyl

230
Substituent increment

Alkyl or ring residue

10

-O-Alkyl, -OH, -O-Ring

25

-O-

11

20

78

-Cl

10

-Br

15

-NH2

13

13

58

-NHC(O)CH3

20

20

45

-NHCH3
-N(CH3)2

73
20

20

85
50

UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

The portion of the EM spectrum from 400-800 is observable to

humans- we (and some other mammals) have the adaptation of
seeing color at the expense of greater detail

400

500

600

700

800

l, nm
Violet

400-420

Indigo

420-440

Blue

440-490

Green

490-570

Yellow

570-585

Orange

585-620

Red

620-780
51

UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

When white (continuum of l) light passes through, or is reflected

by a surface, those ls that are absorbed are removed from the
transmitted or reflected light respectively

What is seen is the complimentary colors (those that are not

absorbed)

This is the origin of the color wheel

52

UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

Organic compounds that are colored are typically those with

extensively conjugated systems (typically more than five)

Consider -carotene

-carotene, lmax = 455 nm

lmax is at 455 in the far blue region of the

spectrum this is absorbed
The remaining light has the complementary
color of orange

53

UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

Likewise:

lycopene, lmax = 474 nm

O
H
N
N
H
O

indigo

lmax for lycopene is at 474 in the near blue region of the

spectrum this is absorbed, the compliment is now red
lmax for indigo is at 602 in the orange region of the
spectrum this is absorbed, the compliment is now indigo!

54

UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

One of the most common class of colored organic molecules are

the azo dyes:

N N

EWGs

EDGs

From our discussion of di-subsituted aromatic chromophores, the

effect of opposite groups is greater than the sum of the individual
effects more so on this heavily conjugated system
Coincidentally, it is necessary for these to be opposite for the
original synthetic preparation!

55

UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

These materials are some of the more familiar colors of our

environment
NO2

HO
N
N

H2N
OH

N N

O3S

N N

NH2
SO3

Para Red

Fast Brown

Sunset Yellow (Food Yellow 3)

56

The colors of M&Ms

Bright Blue

Royal Blue

Common Food Uses

Beverages, dairy products, powders, jellies, confections,
condiments, icing.

Common Food Uses

Baked goods, cereals, snack foods, ice-cream, confections,
cherries.

Orange-red

Lemon-yellow

Common Food Uses

Gelatins, puddings, dairy products, confections, beverages,
condiments.

Common Food Uses

Custards, beverages, ice-cream, confections, preserves,
cereals.

Orange
Common Food Uses
Cereals, baked goods, snack foods, ice-cream, beverages,
dessert powders, confections

57

UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

In the biological sciences these compounds are used as dyes to

selectively stain different tissues or cell structures

Biebrich Scarlet - Used with picric acid/aniline blue for staining

collagen, recticulum, muscle, and plasma. Luna's method for
erythrocytes & eosinophil granules. Guard's method for sex
chromatin and nuclear chromatin.
HO
O3S

N N

N N
SO3

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UV Spectroscopy

V.

Visible Spectroscopy
A. Color
1. General

In the chemical sciences these are the acid-base indicators used for
the various pH ranges:

Remember the effects of pH on aromatic substituents

Methyl Orange

O3S

N N

Yellow, pH > 4.4

CH3
N
CH3

O3S

H
N N

CH3
N
CH3

Red, pH < 3.2

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