Spectroscopy

Ultraviolet Visible Molecular

Absorption Spectroscopy
Chapters 13 & 14 Skoog Holler Nieman 5th ed.
 UV/VIS Basics

“Absorption measurements based on UV and VIS

radiation find widespread application for the
identification and determination of myriad organic
and inorganic species. Molecular UV/VIS absorption
methods are perhaps the most widely used of all
quantitative analysis techniques in chemical and
clinical laboratories throughout the world.”
• The range of wavelengths for common UV/VIS is
180 nm < λ < 700 nm. The energy range can be
described: h = Plank' s constant
hc c = speed of light
E= = hv
λ λ = wavelength
v = frequency
• The photon energy is typically described as kJ/mol
for the UV/VIS region.
• The type of quantum transition is Bonding
Electrons. There are four transitions.
• Limited to chromophores.
• Transition Probability and cross sectional area of
absorbing species dictates magnitude of absorption.
• Subject to solvent effects.
 UV/VIS Terms

• E.g. A solution containing a UV absorbing chromophore

(λmax 250 nm) is impinged with UV radiation (180 to 350
nm) of power PO (note: this is not a monochromatic light
source) (UV/VIS-PDA)
• Some of PO is absorbed due to electron excitation within
the chromophore, and is related to [analyte] and transition
probability
• What emerges from the other side of the sample of a
specific path length is the output power, P.
• The ratio of P to PO forms the basis of transmittance (T)
and absorption (A).
• Beer’s Law
 Beer’s Law
SHN pp302-303

• The derivation of this law is well described.

Po
A = ε ⋅ c ⋅ b = log = − log T
P
where :
ε is the molar extinction coefficient (M -1cm -1 )
c is the analyte concentration (moles/L)
b is the cell path length (cm)

PO P

b
• The sample path length, the analyte concentration,
its medium, its cross sectional area, and most
importantly its extinction coefficient, dictate the
resultant P relative to PO
 The extinction coefficient

• The extinction coefficient is the term that is seldom

thoroughly discussed in carrying out UV/VIS.
• Rearranging Beer’s Law to solve for ε indicates
that it is inversely dependent on concentration and
path length, and directly so on absorbance. A
ε=
c ⋅b
• The ε is actually a complex function which, for any
particular peak in a spectrum, is dependent on the
cross sectional area of the molecule (As in cm²) and
its probability of transition (P);
ε = 8.7 × 10 19 ⋅ P ⋅ As
• The transition probability, or oscillator strength, is
actually a dimensionless value, which relates the
integrated absorption coefficient AC for all the
frequencies in the band.
(4 ⋅ me ⋅ c ⋅ ε o )
f = 2
⋅ AC
Av ⋅ e
 More on Extinction Coefficient
(4 ⋅ me ⋅ c ⋅ ε o )
f = ⋅ AC
Av ⋅ e 2
me : mass of electron (9.11 × 10 −31 Kg )
c : speed of light (2.998 × 10 8 m / s )
ε o : vacuum permit. (8.854 × 10 − 12 C 2 m − 2 N − 1 )
Av : Avogadro's Number (6.023 × 10 23 mol − 1 )
e : Charge on electron (1.602 × 10 − 19 C )

• Intense electronic transitions approach transition probabilities of

unity, while significantly less intense transitions have transition
probabilities significantly less than unity (orders of magnitude).
• The electronic transition from ground state to a higher energy
state would yield a transition probability of unity for all possible
transitions if the electron could vibrate in all three dimensions.
• The integrated absorption coefficient is hidden within the
transition probability, but is quite a significant component. As
stated, the AC is the sum of all the intensities of all the
transitions, so the greater it is, the greater is the transition
probability.
• Calculation of the transition probability is possible using the
transition dipole moment, µ. The transition dipole moment
requires information regarding wavefunctions of the initial and
final states of the molecule. It is therefore possible to calculate
transition probability with a theoretical approach, provided the
wavefunctions are known.
 Extinction Coefficient Ranges

ε = 8.7 × 10 19 ⋅ P ⋅ As

• Typical capture cross section areas are (for

organics) ~10–15cm².
• With transition probabilities ranging from: 0< P <1,
we can realize extinction coefficients generally
ranging from 10² to 105. Note that transition
probabilities <0.01 are called “forbidden”
transitions, in other words, not likely to occur
relative to other more favorable routes of electronic
excitation.
• In summary,
– εmax ~104 to 105 yields strong absorption (intensity)
– εmax ~103 yields medium absorption (intensity)
– εmax <103 yields low absorption (intensity)

• Recall, absorption is the process of exciting an

electron from one energy level to another, of
which there are four possible transitions.
 σ, π, and lone pair (n) electrons

• The σ, π, and lone pair (n) electrons can all undergo

excitation from ground state to excited energy states.
Take a look at a depiction of the various electrons
within a simple molecule, say formaldehyde.

• The four transitions are σ to σ*, π to π*, n to σ*,

and n to π*. Recall that the promoted electrons are
usually bonding electrons.
 σ to σ* Transitions

• These are the bonding

electrons. <150 nm
• The energies required to
excite an electron from
the ground state σ to
excited state σ* is large -
recall: hc
E= = hv
λ
• Exciting an electron from σ
to σ* requires considerable
energy, actually two to three
times more energy compared
to other transitions.
• Conventional UV
spectroscopy cannot be used
to detect these transitions.
– Absorption of radiation by
glass and quartz
– The compounds need to be
in a vacuum. (can you
explain why?)
 n to σ* Transitions

• These are the lone pair

electrons or nonbonding
electrons. 150 to 250nm.
• The transitions energies
are lower than those of σ
to σ*.
• The extinction coefficients
are small.
• Note that there is a
hypsochromic shift upon
exposure of these
compounds to polar
solvents. Why?
• The number of molecules
with n to σ* transitions is
small.
 n to π* Transitions

• Unsaturated molecules that

contain atoms such as
oxygen, nitrogen, and sulfur
exhibit a weak band in their
absorption spectra which can
be assigned to n to π*
transitions.
• For aldehydes and ketones,
this band arises from
excitation of a non-bonding
(lone-pair) electron on the
oxygen atom into the
antibonding π* orbital of the • If the carbonyl group is
carbonyl moiety. part of a conjugated
• In saturated aldehydes and system, the band due to n
ketones, the band due to n to to π* transition generally
π* transition generally occurs appears in the range of
in the range between 270-300 300-350 nm, and has an
nm, with reasonably high εmax substantially greater
εmax. than that of the carbonyl
• In unsaturated carbonyl alone (~10x greater).
compounds, the spectrum is
usually a composite of the
ethylenic moieties and those
of the carbonyl function.
 ...n to π* Transitions
• The addition of species such as OH and Cl, act as
electron donors to the π system of the carbonyl group.
This causes an increases in energy of all the π orbitals,
bonding and anti-bonding, but doesn’t affect the energy
of the n orbital. This causes an increases in the energy
difference between the n and π* orbitals and thus we see
an increase in the energy required to effect the transition,
i.e., shift to shorter wavelength.
• The pH of the medium has a large effect on n to π*
transitions. Consider the protonation of pyridine. This
results in what appears to be a disappearance due to lone-
pair bonding with proton. Under basic conditions (at a pH
>> pKa), absorption at 300 nm is observed, which for
interest is not present in benzene.
• Blue shift or Hypsochromic shift: Hydrogen bonding
has significant impacts on n to π* transitions, that results
in a blue shift, or a shift to shorter wavelengths. Consider
a compound with a carbonyl group in hexane. The
addition of a ethanol (through hydrogen bonding and with
energy ~ equal to the hydrogen bond) will lower the
energy level of the unexcited state of the n orbital, thus
increasing the energy required to effect the transition.
Wavelength shifts of 30 nm are not uncommon here.
• Increasing the polarity of the solvent has an effect on the
n to π*, but not as significantly as does a change in
polarity.
• εmax ranges from ~10 to 100 M-1cm-1
 π to π* Transitions
• Characterized by the
promotion of an electron
from a bonding π orbital to
an anti-bonding π* orbital.
• Selection rules, such as those
based on symmetry
concepts, determine whether
a transition to a particular π*
orbital is allowed or
forbidden.
• There are several
possibilities of π to π*
transitions in molecules
containing several π orbitals
constituting the π-electron
system. The extent to which
transitions are allowed or
forbidden varies, and spectra
may exhibit bands with a
range of intensities (e.g.,
ethylene and benzene).
• Ethylene: The allowed
transition, Ag to B1u
transition is intense (174
nm).
 ...π to π* Transitions
• Benzene:
– A1g to E2u is an
allowed transition
(~180 nm)
(εmax~60,000)
– A1g to B1u (E2 band)
(~204 nm)
(εmax~8,000) and A1g
to B2u (B band)
(~256 nm)
(εmax~200) are
forbidden.
– Note that these bands
are strongly affected
by ring substitution.
 ...π to π* Transitions

• The effect of solvents on the position of bands due to π

to π* transitions is usually opposite to that for n to π*
transitions. A hydrolytic solvent as a medium for a
molecule with potential π to π* transitions will cause a
red shift or bathochromic shift in wavelength.
• Generally speaking, this is due to the fact the excited
state in a π to π* transition is more stable in a hydrolytic
solvent compared to a more hydrophobic solvent.
• An increase in the extent of conjugation on aromatic
compounds causes the absorption bands to shift to longer
wavelengths.
• For aromatic species, the effect of substituting proton in
benzene with various species is quite marked.
• Generally, π to π* εmax ranges from ~1,000 to 10,000 M-
1cm-1
 Auxochromes

• Auxochromes:
– Do not absorb in UV, but can shift chromophore
peaks to longer wavelengths (red shift) and can also
increase their intensities.
– Note that both -OH and NH2 have an auxochromic
effect the B band of benzene.
– The auxochromic substituents have at least one pair
of non-bonding electrons (lone pair) that can interact
with the π electrons of the ring (stabilizes the π*
state).
 Cut-off Points of
Some Common Solvents
 Absorption Characteristics of Some
Common Chromophores

• There are a number of variables that influence absorbance:

– nature of the solvent
– pH of the solution
– temperature (refractive index changes)
– electrolyte concentration
– interfering substances
 Some UV Spectra

• Note that these are NOT “fingerprint” spectra.

• There are huge solvent effects.
 Electronic Energy Levels Including
Transitions from
Vibrational and Rotational Levels

• We have to consider that each energy level has

numerous vibrational energy levels, and those
have numerous rotational energy levels.
• The electronic transition can therefore occur
from energy levels above the pure electronic
ground level energy level. This results in the
broad distribution of absorption seen in UV/VIS.
–There are three basic processes by which a molecule can absorb
radiation; all involve raising the molecule to a higher internal energy
level, the increase in energy being equal to the energy of the absorbed
radiation (hv). The three types of internal energy are quantized; that is,
they exist at discrete levels.
–First, the molecule rotates about various axes, the energy of rotation
being at definite energy levels, so the molecule may absorb radiation and
be raised to a higher rotational energy level, in a rotational transition.
–Second, the atoms or groups of atoms within a molecule vibrate
relative to each other, and the energy of this vibration occurs at definite
quantized levels. The molecule may then absorb a discrete amount of
energy and be raised to a higher vibrational energy level, in a vibrational
transition.
–Third, the electrons of a molecule may be raised to a higher level of
electron energy, corresponding to an electronic transition. Since each
of these internal energy transitions is quantized, they will occur only at
definite wavelengths corresponding to an energy (hv) equal to the
quantized jump in the internal energy.
–There are, however, many different possible energy levels for each type
of transition, and several wavelengths may be absorbed. The transitions
can be illustrated by an energy level diagram. The relative energy levels
of the three transition processes are in the order electronic > vibrational
> rotational, each being about an order of magnitude different in its
energy level.
–Rotational transitions thus can take place at very low energies (long
wavelengths, that is, the microwave or far-infrared region)
–Vibrational transitions require higher energies in the near-infrared
region
–Electronic transitions require still higher energies (in the visible and
ultraviolet regions).
–Christain, G.D., Analytical Chemistry 5th Edition, 1994, pp. 402