UV-VISIBLE spectroscopy
Principle of UV-VISIBLE spectroscopy
UV-VISIBLE spectroscopy obeys the Beer-Lambert law, which states that: when a beam of monochromatic light is
passed through a solution of an absorbing substance, the rate of decrease of intensity of radiation with thickness of
the absorbing solution is proportional to the incident radiation as well as the concentration of the solution.
The expression of Beer-Lambert law is-
A = log (I0/I) = Ecl
Where, A = absorbance
I0 = intensity of light incident upon sample cell
I = intensity of light leaving sample cell
C = molar concentration of solute
L = length of sample cell (cm.)
E = molar absorptivity
From the Beer-Lambert law it is clear that greater the number of molecules capable of absorbing light of a given
wavelength, the greater the extent of light absorption. This is the basic principle of UV-VISIBLE spectroscopy.
Instrumentation and working of UV-VISIBLE spectroscopy
Instrumentation and working of the UV-VISIBLE spectrometers can be studied simultaneously. Most of the modern
UV-VISIBLE spectrometers consist of the following parts-
Light Source- Tungsten filament lamps and Hydrogen-Deuterium lamps are most widely used and suitable light
source as they cover the whole UV-VISIBLE region. Tungsten filament lamps are rich in red radiations; more
specifically they emit the radiations of 375 nm, while the intensity of Hydrogen-Deuterium lamps falls below 375
nm.
Monochromator- Monochromators generally composed of prisms and slits. The most of the spectrophotometers
are double beam spectrophotometers. The radiation emitted from the primary source is dispersed with the help of
rotating prisms. The various wavelengths of the light source which are separated by the prism are then selected by
the slits such the rotation of the prism results in a series of continuously increasing wavelength to pass through the
slits for recording purpose. The beam selected by the slit is monochromatic and further divided into two beams with
the help of another prism.
Sample and reference cells- One of the two divided beams is passed through the sample solution and second beam
is passé through the reference solution. Both sample and reference solution are contained in the cells. These cells are
made of either silica or quartz. Glass can't be used for the cells as it also absorbs light in the UV-VISIBLE region.
Detector- Generally two photocells serve the purpose of detector in UV-VISIBLE spectroscopy. One of the
photocell receives the beam from sample cell and second detector receives the beam from the reference. The
intensity of the radiation from the reference cell is stronger than the beam of sample cell. This results in the
generation of pulsating or alternating currents in the photocells.
Amplifier- The alternating current generated in the photocells is transferred to the amplifier. The amplifier is
coupled to a small servometer. Generally current generated in the photocells is of very low intensity, the main
purpose of amplifier is to amplify the signals many times so we can get clear and recordable signals.
Recording devices- Most of the time amplifier is coupled to a pen recorder which is connected to the computer.
Computer stores all the data generated and produces the spectrum of the desired compound.
, Applications of UV-VISIBLE spectroscopy
1. Detection of functional groups- UV-VISIBLE spectroscopy is used to detect the presence or absence of
chromophore in the compound. This is technique is not useful for the detection of chromophore in complex
compounds. The absence of a band at a particular band can be seen as an evidence for the absence of a particular
group. If the spectrum of a compound comes out to be transparent above 200 nm than it confirms the absence of –
a) Conjugation b) A carbonyl group c) Benzene or aromatic compound d) Bromo or iodo atoms.
2. Detection of extent of conjugation- The extent of conjugation in the polyenes can be detected with the help of
UV-VISIBLE spectroscopy. With the increase in double bonds the absorption shifts towards the longer wavelength.
If the double bond is increased by 8 in the polyenes then that polyene appears visible to the human eye as the
absorption comes in the visible region.
3. Identification of an unknown compound- An unknown compound can be identified with the help of UV-
VISIBLE spectroscopy. The spectrum of unknown compound is compared with the spectrum of a reference
compound and if both the spectrums coincide then it confirms the identification of the unknown substance.
4. Determination of configurations of geometrical isomers- It is observed that cis-alkenes absorb at different
wavelength than the trans-alkenes. The two isomers can be distinguished with each other when one of the isomers
has non-coplanar structure due to steric hindrances. The cis-isomer suffers distortion and absorbs at lower
wavelength as compared to trans-isomer.
5. Determination of the purity of a substance- Purity of a substance can also be determined with the help of UV-
VISIBLE spectroscopy. The absorption of the sample solution is compared with the absorption of the reference
solution. The intensity of the absorption can be used for the relative calculation of the purity of the sample
substance.
Atomic Absorption Spectroscopy
Meaning of Atomic Absorption Spectroscopy
Atomic Absorption Spectroscopy was invented by Alan Walsh in 1950’s for the qualitative determination of trace
metals in liquids. The superiority of the technique over other is based on the fact that by this technique 50-60
elements can be determined without any interference from trace to big quantities.All these elements can be detected
here which fail to yield satisfactory result in flame photometry. Thus, it is a successful instruments for detection and
estimation of metals and non-metals both types of pollution from factories.The technique has also proved very
helpful to both aqueous and non-aqueous solutions.
Principle of Atomic Absorption Spectroscopy:
When a solution having a mixture of metallic species is introduced into the flame, the solvent evaporates and vapour
of metallic species is obtained. Some of metal atoms can be raised to an energy level sufficiently high to emit
characteristics radiation of metal-a phenomenon that is used in flame photometry. Here a large amount of metal
atoms remain in non-emitting ground state.These ground state atoms of a particular element are receptive of light
radiation of their own specific resonance wavelength. In this way, when a light of this wavelength passes through a
flame, a part of light will be absorbed and this absorption will be proportional to the intensity of atoms in the
flame.So in atomic absorption spectroscopy the amount of light absorbed is determined because the absorption is
proportional to the concentration of the element.
Advantages of Atomic Absorption over Flame Photometry:
(1) It does not suffer from spectral interference, which occurs in flame emission spectroscopy.
(2) It is independent of flame temperature.
Principle of UV-VISIBLE spectroscopy
UV-VISIBLE spectroscopy obeys the Beer-Lambert law, which states that: when a beam of monochromatic light is
passed through a solution of an absorbing substance, the rate of decrease of intensity of radiation with thickness of
the absorbing solution is proportional to the incident radiation as well as the concentration of the solution.
The expression of Beer-Lambert law is-
A = log (I0/I) = Ecl
Where, A = absorbance
I0 = intensity of light incident upon sample cell
I = intensity of light leaving sample cell
C = molar concentration of solute
L = length of sample cell (cm.)
E = molar absorptivity
From the Beer-Lambert law it is clear that greater the number of molecules capable of absorbing light of a given
wavelength, the greater the extent of light absorption. This is the basic principle of UV-VISIBLE spectroscopy.
Instrumentation and working of UV-VISIBLE spectroscopy
Instrumentation and working of the UV-VISIBLE spectrometers can be studied simultaneously. Most of the modern
UV-VISIBLE spectrometers consist of the following parts-
Light Source- Tungsten filament lamps and Hydrogen-Deuterium lamps are most widely used and suitable light
source as they cover the whole UV-VISIBLE region. Tungsten filament lamps are rich in red radiations; more
specifically they emit the radiations of 375 nm, while the intensity of Hydrogen-Deuterium lamps falls below 375
nm.
Monochromator- Monochromators generally composed of prisms and slits. The most of the spectrophotometers
are double beam spectrophotometers. The radiation emitted from the primary source is dispersed with the help of
rotating prisms. The various wavelengths of the light source which are separated by the prism are then selected by
the slits such the rotation of the prism results in a series of continuously increasing wavelength to pass through the
slits for recording purpose. The beam selected by the slit is monochromatic and further divided into two beams with
the help of another prism.
Sample and reference cells- One of the two divided beams is passed through the sample solution and second beam
is passé through the reference solution. Both sample and reference solution are contained in the cells. These cells are
made of either silica or quartz. Glass can't be used for the cells as it also absorbs light in the UV-VISIBLE region.
Detector- Generally two photocells serve the purpose of detector in UV-VISIBLE spectroscopy. One of the
photocell receives the beam from sample cell and second detector receives the beam from the reference. The
intensity of the radiation from the reference cell is stronger than the beam of sample cell. This results in the
generation of pulsating or alternating currents in the photocells.
Amplifier- The alternating current generated in the photocells is transferred to the amplifier. The amplifier is
coupled to a small servometer. Generally current generated in the photocells is of very low intensity, the main
purpose of amplifier is to amplify the signals many times so we can get clear and recordable signals.
Recording devices- Most of the time amplifier is coupled to a pen recorder which is connected to the computer.
Computer stores all the data generated and produces the spectrum of the desired compound.
, Applications of UV-VISIBLE spectroscopy
1. Detection of functional groups- UV-VISIBLE spectroscopy is used to detect the presence or absence of
chromophore in the compound. This is technique is not useful for the detection of chromophore in complex
compounds. The absence of a band at a particular band can be seen as an evidence for the absence of a particular
group. If the spectrum of a compound comes out to be transparent above 200 nm than it confirms the absence of –
a) Conjugation b) A carbonyl group c) Benzene or aromatic compound d) Bromo or iodo atoms.
2. Detection of extent of conjugation- The extent of conjugation in the polyenes can be detected with the help of
UV-VISIBLE spectroscopy. With the increase in double bonds the absorption shifts towards the longer wavelength.
If the double bond is increased by 8 in the polyenes then that polyene appears visible to the human eye as the
absorption comes in the visible region.
3. Identification of an unknown compound- An unknown compound can be identified with the help of UV-
VISIBLE spectroscopy. The spectrum of unknown compound is compared with the spectrum of a reference
compound and if both the spectrums coincide then it confirms the identification of the unknown substance.
4. Determination of configurations of geometrical isomers- It is observed that cis-alkenes absorb at different
wavelength than the trans-alkenes. The two isomers can be distinguished with each other when one of the isomers
has non-coplanar structure due to steric hindrances. The cis-isomer suffers distortion and absorbs at lower
wavelength as compared to trans-isomer.
5. Determination of the purity of a substance- Purity of a substance can also be determined with the help of UV-
VISIBLE spectroscopy. The absorption of the sample solution is compared with the absorption of the reference
solution. The intensity of the absorption can be used for the relative calculation of the purity of the sample
substance.
Atomic Absorption Spectroscopy
Meaning of Atomic Absorption Spectroscopy
Atomic Absorption Spectroscopy was invented by Alan Walsh in 1950’s for the qualitative determination of trace
metals in liquids. The superiority of the technique over other is based on the fact that by this technique 50-60
elements can be determined without any interference from trace to big quantities.All these elements can be detected
here which fail to yield satisfactory result in flame photometry. Thus, it is a successful instruments for detection and
estimation of metals and non-metals both types of pollution from factories.The technique has also proved very
helpful to both aqueous and non-aqueous solutions.
Principle of Atomic Absorption Spectroscopy:
When a solution having a mixture of metallic species is introduced into the flame, the solvent evaporates and vapour
of metallic species is obtained. Some of metal atoms can be raised to an energy level sufficiently high to emit
characteristics radiation of metal-a phenomenon that is used in flame photometry. Here a large amount of metal
atoms remain in non-emitting ground state.These ground state atoms of a particular element are receptive of light
radiation of their own specific resonance wavelength. In this way, when a light of this wavelength passes through a
flame, a part of light will be absorbed and this absorption will be proportional to the intensity of atoms in the
flame.So in atomic absorption spectroscopy the amount of light absorbed is determined because the absorption is
proportional to the concentration of the element.
Advantages of Atomic Absorption over Flame Photometry:
(1) It does not suffer from spectral interference, which occurs in flame emission spectroscopy.
(2) It is independent of flame temperature.
