UV-Visible spectroscopy:

Ultraviolet and visible spectrometers have been in general use for the last 35 years
and over this period have become the most important analytical instrument in the
modern day laboratory. In many applications other techniques could be employed but
none rival UV-Visible spectrometry for its simplicity, versatility, speed, accuracy and
cost-effectiveness.

The chemical sample which may include

atom, ion or molecule. These analyte
especially molecules possesses different
electronic states (energy level).

The figure illustrates the general pattern of electronic energy levels, of a molecule
and the fact that the transitions are brought about by the absorption of different
amounts of energy. Light in the UV-VIS part of the spectrum is used to promote
electrons from the ground state to various excited states.
 Examples of transitions and resulting λmax
Molecule Transition λmax
CH3-CH3  - * 135
CH3-OH  - * 150
n - * 183
CH2=CH2  - * 175
C6H6  - * 245

Measurement of Absorbance:
General terms and their relations :

Transmittance: It is the ratio between Relations between,

power of transmitted radiation to the A, T, and path length:
power of incident raciation. (for the development of Beers Law)

PSolution P
T 
PSolvent P0

Absorbance: Negative logarithm of the

transmittance is called Absorbance.

P0
A  - logT  log
P
A  - log(%T)  2
In the form of Lamberts Beers law:
Path length / cm 0 0.2 0.4 0.6 0.8 1.0

A  bC %T
Absorbance
100
0
50
0.3
25
0.6
12.5
0.9
6.25
1.2
3.125
1.5
 Lamberts Beers Law: Absorptivity and molar absorptivity:

Magnitude of molar absorptivity's

Lambert and Beer studied the relation
between absorbance of a solution with
→ It is the property of absorbing species.
length and concentration of solution → It varies between 0 → ≈ 105
and found that: → If ε ‹ 103 low intensity of peak (abs), is
considered as forbidden transition,
Absorbance  Concentration probability ≈ 0.01%.
Absorbance  Path length
for forbidden transition, ε = 0 – 1000
Finally generate a equation:
Where
A = 𝜀bc …… (i) b = pathlength → ε > 104 high intensity of absorption peak,
c = concentration
𝜀 and a are → Unit of absorptivity is determined by the
proportionality unit of b and c.
constants
A = 𝑎bc …… (ii) • b in cm, c in g/L,
i,e. a has unit of L g-1cm-1
• b in cm, c in M (m/L),
𝜀 = molar absorptivity coefficient
a = absorptivity coefficient
i.e. 𝜀 has unit of L mol-1cm-1
 A = 𝜀bc
𝐴
or 𝜀 =
𝑏𝑐
• "𝜀 is a measure of the amount of light absorbed per unit concentration" when it
passes through 1cm length.
• Molar absorptivity is a constant for a particular substance, so if the concentration
of the solution is halved so is the absorbance, which is exactly what you would
expect.
• Let us take a compound with a very high value of molar absorptivity, say 100,000 L
mol-1 cm-1, which is in a solution in a 1 cm path length cuvette and gives an
absorbance of 1.
𝜀=1/1.c
Therefore, c = 1 / 100,000 = 1 × 10-5 mol L-1
• Now let us take a compound with a very low value of 𝜀, say 20 L mol-1 cm-1 which is
in solution in a 1 cm path length cuvette and gives an absorbance of 1.
𝜀=1/1.c
Therefore, c = 1 / 20 = 0.05 mol L-1
• It concludes that - a compound with a high molar absorptivity is very effective at
absorbing light (of the appropriate wavelength), and hence low concentrations of
a compound can be easily detected. This is reverse for the compounds with low
absorptivity.
 Application of Beer’s law to mixtures:

Total absorbance of mixture can be obtained by taking the

sun of absorbance by individual species.
ATotal = A1 + A2 + A3 +…….+ An
= ε1bC1 + ε2bC2 + ε3bC3 +………..+ εnbCn
If polychromatic radiation is used,
• Absorbance at wavelength λ1, A’= ε’1bC1 + ε’2bC2 + ε’3bC3 +………..+ ε’nbCn
• Absorbance at wavelength λ2, A’’= ε’’1bC1 + ε’’2bC2 + ε’’3bC3 +………..+ ε’’nbCn

Validity of Beer’s Law:

In the Beers Law
• A α b is valid in most of the case when concentration is same.
• A α c is not valid in most of the cases when b is constant.
(a) fundamental/ real limitation and
(b) other. [depends on manner/ change of composition/ (instrumental or
chemical deviation)]
 Validity of Beer’s Law:
Real Limitation Of Beers Law:
Beer’s law is successful in describing the absorption behavior of dilute solutions only;
in this sense it is a limiting law.
o The analyte solution must be dilute, high concentrated solution will not obey
Beer’s law.
o At high concentration > 0.01M the absorbance may be reduced due to the effect
in charge distribution by interaction with neighboring molecules. Such type of
interaction can alter the ability of analyte molecules to absorb the given
wavelength.

o High concentration of some other colorless salts adversely affects the absorption
of light. Similar electronic interaction with analyte may occur.

o Change of refractive index of the medium do not adhere the Beer’s law. If the
concentration change causes significant deviation of refractive index departure
in Beers law will be originated.
A correction can be made by substituting ε with,
 = true x h/( h² + 2)²
 Apparent Chemical Deviations
• Caused by association/dissociation/reaction with solvent…..}.
A common example of this behavior is found with acid/base indicators.
Deviations arising from chemical factors can only be observed when
concentrations are changed.
430 570
HIn H+ + In- HIn 6.30 X 102 7.12 X 103
Color 1 Color 2 (Ka = 1.42 X10-5)
In- 2.06 X 104 9.61 X 102
If we calculate absorbance data for
the total indicator concentration
K a  1.42  10 5
If, [In] total  2  10 - 5 M
Gives, [In-]  1.12  10 -5 M and [HIn]  0.88  10 -5 CHIn, M [HInN] [IN-] A430 A570
0.88x10- 1.12x10-
2x10-5 5 5 0.236 0.073
2.22x10- 1.78x10-
A 430  (2.06  10 4  1  1.12  10 5 )  (6.30  10 2  1  0.88  10 5 )  0.236 4x10-5 5 5 0.318 0.175
5.27x10- 2.73x10-
5 5
A 570  (9.61  10  1  1.12  10 )  (7.12  10  1  0.88  10 )  0.073
2 3 8x10-5 5 5 0.596 0.401
8.52x10- 3.48x10-
12x10-5 5 5 0.771 0.640
11.9x10- 4.11x10-
16x10-5 5 5 0.922 0.887

Beer’s law is not obeyed at both wavelength. The direction of fluctuation of absorbance from Beer’s law is opposite at two wavelength.
 Apparent Instrumental Deviations with Polychromatic Radiation
A good picture of the effect of polychromatic radiation on Beer’s law can be
presented as follows.
If we considered just the two and λ”, and assuming that Beer’s law applies at each of
these individual wavelengths,

The absorbance at λ‘
P0′
A = log ′ = 𝜀′bc P = Po 10-bc
P
The absorbance at λ‘‘
P′′0
A = log ′ = 𝜀′′bc P” = P”o 10-”bc
P′
The Overall Absorbance
(P0'  P0'' )
A m  log ' , m  measured
(P  P ) ''

Total incident power = P'’o + P”'o'

putting the valuespower
Total transmitted of P =&P’P+ P”
(P0'  P0'' )
A m  log '
(P0 x 10 - 'bc  P0'' x 10 - ''bc )
A m  log (P0'  P0'' ) - log(P0' x 10 - 'bc  P0'' x 10 - ''bc )

In the very special case where ’ = ’’, the above equation simplifies to Beer’s law. Am = ’bc
Instrumental Deviations in the Presence of
Stray Radiation

(P0  PS )
A'  log ................ (19)
(P  PS )

Ps is the power of non absorbed stray radiation.

Validity of Beer’s Law:

•The radiation source must be monochromatic
•The analyte solution must be dilute, <0.01M
•High concentration of some other colourless salts adversely affects
•Change of refractive index
•Solution containing stray radiation
•There should be no chemical reactions like hydrolysis, association, dissociation,
polymerization
• The graph of A vs c must be straight line passing through origin.
Instrumentation for UV visible spectroscopy

Components are,
 (1) sources,  (2) wavelengths selectors  (3) sample containers,
 (4) radiation detectors,  (5) signal processors and readout devices.
Light Sources
For the purpose of molecular absorption a continuous source is
required with constant power over a considerable range of
wavelengths.
Deuterium and Hydrogen Lamps
Deuterium or hydrogen between the two electrodes will be
excited at low pressure and gives continuum UV spectrum.
The excited molecule dissociates into atomic species and UV
photons.

D2 + Ee D2* D’ + D’’ + hn , •Produce a useful continuum spectrum in the region

of 160 to 375nm.
Ee = ED2 = ED' + ED'' + hn •At longer wavelengths, the lamps produce emission
Varies between 160 -375 nm lines, which are superimposed on the continuum
KE of two D atoms varies spectrum
lineraly and continuously •Note that the maximum intensity occurs at
Fixed quantized energy ～225nm
Tungsten Filament Lamps ‘incandescent’ lamps
•Are commonly used in the visible and near-infrared region.
•Useful in wavelength range (320 to 2500 nm)
• At the usual operating temperature of about 3000 K, only about 155 of the
total radiant energy falls in the visible region.
•In visible region the energy output of tungsten lamp varies as forth power of
operating voltage.
•For stable radiation voltage should be closely controlled by electronic voltage
regulators or constant voltage transformers.
In Tungsten halogen Lamps
•A quartz envelope houses tungsten filament containing small amount of iodine.
(quartz can resist high temperature ≈3500K)
•This has double life time than ordinary tungsten lamp, because of iodine gas.
W gas + I2 = WI2 (volatile compound), when WI2 strikes the filament, tungsten is
redeposited by deposition.

These lamps provide a continuous source of radiation form 375 nm down to about 160 nm
hence quartz windows must be used with these types of lamps since glass absorbs strongly
at wavelengths less than about 350 nm
•Xenon arc lamp:
•Radiation is produced by passing current through xenon atmosphere.
•It is applicable for 200-1000nm, has intense peak around 500nm.
 •Wavelength selector:
Are the device/s which restricts radiation to a narrow band, that is absorbed or
emitted from analyte.
Determine sensitivity and selectivity of the method
.These are filters, monochromators, spectrograph etc.
Radiation filters: Filter absorb all but a restricted band of radiation from a
continuum source. Interference filters (Febry-Perot filters):\
White radiation

Glass plate
Metal film
Dielectric layer

Narrow band of radiation


1 2 3 4
A

 4'
1' 2' 3'
B

nl = 2th
 Monochromators generally have a diffraction grating disperse the radiation into its
component wavelengths. contains: entrance slit, collimating lens, mirrors, focusinglens,
a dispersion medium (prism and grating) and an exit slit.
Mechanism of diffraction from an
echellette-type grating

(a) grating monochromator

nλ = d(sin i + sin r)
Use geometry to derive this relation

(b) prism monochromator

http://www.monzir-
•A refraction grating disperse radiation into its component
pal.net/Instrumental%20Analysis/Content
wavelength.
•Different wavelength are focused to exit slit by rotating the _Vis1.htm
grating.
•Can provide radiation of effective bandwidth (spectral band) of
less than 1 nm (expensive) to about 20nm (inexpensive)
 Cuvettes (sample containers):
1. Virtually all UV spectra are recorded solution-phase
2. Cells can be made of plastic, glass or quartz
3. Only quartz is transparent in the full 200-700 nm
range; plastic and glass are only suitable for visible
spectra
Radiation detectors
Phototube - electrons produced by irradiation of cathode
travel to anode. response depends on cathode material
(200-1000 nm)

Photomultiplier tube (PMT) - irradiation of

cathode produces electrons, series of anodes
(dynodes) increases gain to 105-107 electrons
per photon.
 Types of instruments
.

Single Beam Instruments

Two sources are used, a tungsten halogen and a deuterium lamp

The wavelength selector is a grating or prism monochromators
the detector is usually a vacuum phototube or a photomultiplier tube in higher cost
instruments
Double Beam Instruments
a. UV-Vis Double Beam Spectrophotometer (in space) Two sources are existent
namely a tungsten halogen
lamp and a deuterium lamp.
A lamp selection mechanism
is present.

The beam from the source is split into two beams by a beam splitter which is a
semipermeable mirror.
The detector is usually a pair of photomultiplier tubes connected to a difference amplifier
and the wavelength selector is a grating or prism monochromators.
 ABSORBING SPECIES
The absorption of UV/Vis. radiation by a molecular/atomic/ionic
species M can be considered to be a two-step process,

Excitation of Absorbing species: Relaxation / deexcitation of excited

-Represented as: M + hn  M* (excited - Via loss of heat M*  M + heat (to
sps.)
- Due to electronic transition.
-Lifetime of M* is ≈ 10-8-10-9 sec
- Bonding or non bonding electron from
valence cell is excited.
- l max of abs. peak is related to nature
of bond or functional gr. Which can be
identified it in a molecule.
- not limited in identification/
quantitative determination can be done.
species:
surrounding)
- Via decomposition i.e. photo chemical
reaction. Further heat loss to
surrounding
- Via re-emission fluorescence and
phosphorescence. (Extra evidence for
detail identification)
 Types of electronic Transitions
From the nature of electrons in a molecule, of transition are categorized into three
types. Which includes the involvement of;
(a) Transition of electrons between orbital's
 Three types of orbital electrons (, and n)
 it may be bonding (, ), non bonding( lone pair) & anti bonding(*, *)
 The loosely held electron (valence) will get excited by radiation
 High energy transition (labsorption < 185nm --vacuum UV)
 Low energy transition (labsorption > 185nm -- UV/Vis)- Chromophores
 The electronic spectra of molecule containing chromophores is complex since vib.
energy levels are superimposed on electronic energy levels (simultaneous rot, vib
and electronic transition). Thus molecular absorption is continuous broad band
spectrum. Thus UV/vis. Absorption is mostly used for qualitative and semi
qualitative analysis.
 Head to head overlapping –  bond (strong)/ Side wise overlapping –  bond
(weak)
 n electrons are localized to single atom (O,N,S,X), &electrons to many bonded
atoms (MO)
 s Types of electrons ,& n in
r on s tron
s
H t
H  le ctro  lec ns
c n formaldehyde
 le n ctro
C x xO C  O n le
H H lectrolencstrons

Nature and structure of bonding non bonding and anti

bonding orbitals
Superposition of vib energy
levels on electronic level:
Orbita (fig)
l
* Orbital v3
v2
E1 v1
v0
Orbital

* Orbital v3
v2
v1
E0 v0
n bonding Orbital has the shape of p or hybridised orbital.
1. σ  σ* trastion:
It is the transition between extreme position
Large energy is required; (vacuum UV) l max < 185nm .
CH4, l max = 125nm, C2H6, l max = 135nm
The strength of C-C > C-H bond (due to hybridization/overlapping)
l absorption Falls below cut-off point of most solvents [200-700]
Anti bonding orbital
C C *
*

C C
Bonding orbital
C C 

1. Alkane:
Fig. Electronic trastion: σ  σ* trastion:
2. n  σ* trastion:
# A saturated organic compound must contain hetero atom/ with lone pair.
# The energy of transition, 150 nm < l max < 250nm
# maximum are below 200nm
#  ranges between 100-3000 Lcm-1 mol-1 .less intense peak
# l for transtion depends on the nature of the bond and not on the structure.
H2O (l max= 167nm), CH3OH (l max=184 nm),
(CH3)2O (lmax=184 nm), CH3I (l max=258 nm),
# Absorbance maxima will shift to short wavelength by polar solvents ?? and is less
accessible for UV-vis absorption (below cut-off point) .
. n => *
C N * CN
C N

n(sp3)
C N -OH , -NH2= l175-200nm)

-SH , -S-S= l200-220nm)

C N CN

2. Alcohol, ether, amines, s-compounds:

Fig. Electronic trastion: n  σ* trastion:
a) n  * require 3. n  * and  * transtion:
presence of  It is the most accessible transtion in
unsaturated bond
and bonding atom UV-vis absorption.
with lone pair of  l max will lie between 200 to 700 nm
electron. (considered as chromophores)
 It is forbidden
transition, C O * CN
l180-290nm,  =15
10 <  < 100Lcm-1mol-1
 lmax shift to short n => *
wavelength with C O * CO C O
polarity of solvent
hypso/blue shift.
Ground state will be
n(py)  => *
more stabilized by C O
solvent. H-bonding
will show drastic shift
of lmax by around C O  C O
30nm [lowering
energy of n electron ≈
CO l188nm,  =900
energy of H-bond] C O
 * excited state the
energy is not affected
by H –bond. No H- C O n(sp)
bond will occur by
single excited
electron 4. Carbonyl-compounds:
Fig. Electronic trastion: n  * and  * trastion:
b)  *
 Only presence of unsaturated bond is sufficient for  * transition.
 1000 <  < 10,000 Lcm-1mol-1 (is allowed transition give intense peak/ easily
observed);
 Has very less effect by solvent polarity ≈5nm or less, the shift in lmax is
commonly to high wavelength batho/red shift.
 Attractive polarization between solvent and analyte molecule will lower
energy level of both ground state and excited state. The effect is greater for
excited state. Thus the energy gap decreases with solvent polarity.
C C *

C C

Alkene = l75nm)
C C 
-Alkyne= l70nm)

3. Alkene, alkyne:
Fig. Electronic trastion:  * trastion:
 Organic chromophores and their approximate absorption maxima:

Note: This lmax is affected by solvent and structure details and also due to vibrational precise
determination of peak maxima is difficult. Also it is better to calculate max from cal. Curve
than the tabulated value.
Absorption spectra of acetone.
• λmax is the wavelength where maximum
absorption occurred
Chromophores:
• A chromophore (literally color-bearing) group is a functional group, not
conjugated with another group, which exhibits a characteristic absorption
spectrum in the ultraviolet or visible region. Like C=C, C=O, N=N, C=S
{n ⟶ 𝜎* , n ⟶ π* , π ⟶ π* }
• If the structural change occurs in the chromophore, the exact energy
and intensity of absorption are expected to be changed.
• When these groups are/is present the absorption pattern of the
compound give rough guidelines in identify the functional group.
• If any of the simple chromophores is conjugated with another (of the
same type or different type) a multiple chromophore is formed having a
new absorption band which is more intense and at a longer wavelength
that the strong bands of the simple chromophores.
 Auxochrome:
• Functional group (or substituent’s) that do not absorb itself in
UV/vis region (above 220nm – 800 nm) but shift the
chromophore to long wavelength and also increase the intensity
of absorption.
• Auxochrome should contain at least a lone pair e- , which stabilizes
*excited state. e.g. Phenolate is strong auxochrome than phenol (anion has
extra pair of e-)
• Aniline is auxochrome but converting it to anilium cation the
effect is lost.
• Some auxochromes are –OH, -NH2
Shift Effect

Bathochromic (red) shift:

longer wavelength / low energy
Hypsochromic (blue) shift:
shorter wavelength / high energy
Hyperchromic:
an increasing intensity
Hypochromic:
decrease in intensity
Effect of Conjugation on λmax

• Conjugation refers to delocalization of pie-electrons.

• Conjugation raises the energy of the HOMO and lowers the energy of the LUMO
[* orbital (give it less antibonding character)]

• With increase in conjugation the HOMO-LUMO gap will decrease, the absorption
maxima for n  * and  *transtion shifts to longer wavelength (batho)
• Absorption by multifunctional chromophores in a single organic molecule is
approximately additive if are separated from one another by more than one single
bond.
• The conjugation of chromophore not only produce bathochromic shift but also
increase the intensity.
 Alkene lmax 
Ethylene 175 15000
1,3 butene 217 21000
1,3,5 hexatriene 258 35000
b carotene(4 double 465 125000
bond)
ketones
Acetone  *189 300
n  * 280 12
213
3-butene-2-one * 7100
320 27
n  *

Fig: effect of conjugation in alkenes

 Solvents Common solvents
with cutoff line:
• The effect on the absorption spectrum of a compound
when diluted in a solvent will vary depending on the

Lower wave
length limit
chemical structures involved.

Solvents
• Polar molecules when interacted with polar solvents show
dramatic effect.
• Interaction between solute and solvent leads to Acetonitrile 190
absorption band broadening and a consequent reduction Chloroform 240
in structural resolution and ε max. Cyclohexane 195
• Figure illustrates the effect of iso-octane and ethanol on 1-4 dioxane 215
the spectrum of phenol, a change from hydrocarbon to
95% ethanol 205
hydroxylic solvent.
N hexane 201
• The loss of fine structure in the latter is due to broad band
h-bonded solvent-solute complexes replacing the fine Methanol 205
structure present in the iso-octane. Isooctane 195
• The fine structure in the latter solvent illustrates the Water 190
principle that non-solvating or nonchelating solvents Trimethyl 210
produce a spectrum much closer to that obtained in the phosphate
gaseous state.
Spectra of Phenol in Iso-octane and in Ethanol

Effect of solvent on Ultraviolet absorption by 1,2,4,-triazine in

absorption of vacuum phase, hexane solution, aqueous
ACETALDEHYDE
solution
 (b) Transtion of electrons between degenerate orbitals by ligand (i.e.
absorption including d and f electrons)
Transition metals, lanthanides and actinides/-ions also absorbs in UV/visible region

A. Lanthanides and actinides:

 The spectra are narrow, well defined &
are characteristic peak i.e. the unique
feature.
 Radiation is absorbed by excitation of
4f and 5f orbital electrons.
 The 5f electrons; situated in inner
orbital are largely screened by external
influence of outer electrons (occupying
high energy orbital's), thus bands are
more narrow and are relatively less
affected by the solvent & spec. bonded
by outer electrons.

Figure: spectra of actinides

B. I and II transtion elements: dz 2 dx 2 - y 2
dx 2 - y 2

dxy dxz dyz

 5 d orbitals has different geometry & possess dxy

equal energy in isolated gaseous state.  dz 2

Energy
dxz dyz

 In complex or in solution the energy of orbital dxy dxz dyz

dxy dxz dyz dz 2 d x 2 - y 2

get changed, due to presence of electron dz 2 dx 2 - y 2

Tetrahedral Square planar
No ligand Octahedral
donating species. field ligand field ligand field ligand field

 The absorption band is broad & is highly

influenced by chemical environmental factors;
for eg, aq. Cu2+ [Cu(H2O)4]2+ is pale blue and
ammonical Cu2+ [Cu(NH3)] 2+ is deep blue
 The magnitude of Crystal field splitting depends
on the:
 Nature of ligand field strength:
Spectrochemical series =>[I-< Br-<Cl-<F-
<OH-<C2O42-≈H2O<SCN-<NH <et diamine<o
phenonthroline<NO2-<CN-] ligand field
increases will decrease l max.
 Charge possessed by the metal i.e.
Oxidation state.
 Position of metal in the PT.
(c) Excitation of electron may cause charge transfer from one atom or
group of atom to next atom or group: Charge transfer absorption:
 Many organic complex are sensitive to charge transfer
process; most e electron transfer occurs from ligand to
metal (reverse is exceptional: o-phenonthroline complex of
Fe(II)/or Cu(I)
 In charge transfer, one component is electron donor and
next is electron acceptor , e- is donated from low energy
orbital to high energy orbital is internal redox process.
 Has high molar absorptivities ( max>10,000), (Fully allowed
transition/ high intensity) hence are important for
..analysis.
For eg. FeSCN2+, Fe(phen)32+, Starch-I5- are shown in fig.
Interesting charge transfer complex of organic compounds:
Quinhydrone (1:1 complex of quinine and hydroquinone)
Absorption by inorganic
&Iodine complex with amines, aromatics and sulfides. ions n – *
In: Fe(III)-SCN complex Nitrate-313nm
photons → electron is transformed from SCN- to Fe(III) → SCN Carbonate-217nm
+ Fe(II) → may relax to ordinary condition or dissociates to
Nitrite-300 & 280nm
give redox product.
Azide-230nm

Trithiocarbamate-500nm
 APPLICATION OF UV/vis Spectroscopy
Application to absorbing species:
Use table:
• The determination of organic compound containing one or more of
these chromophores is feasible (many examples are found in
literature)
• Many inorganic species are susceptible for quantitative determination.
Like transition metal ion, nitrate ion, nitrite, chromate ion, osmium and
ruthenium tetroxides, molecular iodine, ozone

Application to non absorbing species:

• Different selective color forming reagents are available that reacts with
non absorbing species, the product formed will strongly absorb in
UV/vis region. In this analysis either the reaction should be complete
very fast or abs. is to be taken at controlled time.
• The color forming reagents can also be applied to absorbing species
like transition metal ion if 𝜀 of product >> uncombined species.
• There are many selective complexing agent are available to determine
inorganic species of our interest.
– SCN- for Fe, Co, Mo; Peroxide ion for Ti, V, Cr; I2- for Bi, Pd, Te
– Other are organic chelating ligands giving colored complex o-phenonthroline
for Fe2+,DMG for Ni2+, diethylthiocarbonate for Cu2+, diphenyl thio carbazone
for Pb2+.
1. For qualitative analysis: The steps are;
A. Plotting Spectral Data/ presentation of data: Several different methods.
– The ordinate is most commonly %T, A, log 𝜀, etc.
– The abscissa is usually λ , 𝜈, wave number cm-1.
– A plot gives large difference in peak height than %T as a function of concentration.
– Log A plot has lost the spectral details but convenient for comparison of spectra of solution
of different concentration.
B. Criteria for solvents:
i. Must be transparent:
ii. The next criteria is effects in fine structures non polar/polar/gas.[H-bond ] i.e. polar solvent
will lost spectral fine structures (arising from vibrational effects),
iii. Third criteria is stabilization of either ground state or excited state:
Polar solvent => lower energy of n orbital => shift n-π* to short λ
Reverse effect may obtain if polar solvent form H-bond with excited state => shift to short
λ
– λ max can also be affected by solvent
C. Determination of functional group (i.e. type of chromophores).
Use table:
i. A weak absorption band around 280-290nm indicates presence of C=O group (displaces to
short wavelength with polarity of solvent)
ii. A weak absorption band around 260nm, with vib fine structure indicates existence of
aromatic ring)
iii. By comparing the effect of pH on spectra of solution containing sample will indicate
presence of aromatic amine or phenolic structure.
iv. If two or more benzene conjugate is existing, it shifts the band to more visible region.
2. Quantitative analysis:
It is the one most useful and widely used tool available to the chemist for
quantitative analysis. General features are,
i. Wide applicability to both organic and inorganic species.
ii. Highly sensitive from 10-4-10-5 or higher 10-7 (with simple modification)
iii. Moderate to high selectivity.
iv. Good accuracy (relative uncertainty 1-3%)
v. Ease and convenience of data acquisition.

Procedural detail:
i. Selection of wavelength.
ii. The variables which can alter absorbance should be controlled.
iii. Cleaning and handling of cell/ cuvettes.
iv. Plotting the calibration curve for standard.
v. Dealing with mixtures of absorbing species.
References:
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