Spectroscopy and its possible branch in civil engineering
nafeesha2005
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Jul 14, 2024
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About This Presentation
Tecroscopic and its branches in civil engineering and its possible ways can be identified here it will be very useful for you so you can use it here which is very we would be specific for you this PPT would find your better way to findTecroscopic and its branches in civil engineering and its possibl...
Slide Content
Spectroscopy
+ It is the branch of science that deals with the
study of interaction of matter with light.
OR
+ It is the branch of science that deals with the
study of interaction of electromagnetic
radiation with matter.
+ It is the branch of science that deals with the
study of interaction of matter with light.
OR
+ It is the branch of science that deals with the
study of interaction of electromagnetic
radiation with matter.
\ Electromagnetic
\Radiation
\Radiation
Electromagnetic Radiation
+ Electromagnetic radiation consist of discrete
packages of energy which are called as
photons.
+ A photon consists of an oscillating electric field
(E) & an oscillating magnetic field (M) which
are perpendicular to each other.
+ Electromagnetic radiation consist of discrete
packages of energy which are called as
photons.
+ A photon consists of an oscillating electric field
(E) & an oscillating magnetic field (M) which
are perpendicular to each other.
Magnetic field
Electric field Direction
Electric field Direction
Electromagnetic Radiation
+ Frequency (v):
— It is defined as the number of times electrical field
radiation oscillates in one second.
— The unit for frequency is Hertz (Hz).
1 Hz = 1 cycle per second
+ Wavelength (A):
— It is the distance between two nearest parts of the
wave in the same phase i.e. distance between two
nearest crest or troughs.
+ Frequency (v):
— It is defined as the number of times electrical field
radiation oscillates in one second.
— The unit for frequency is Hertz (Hz).
1 Hz = 1 cycle per second
+ Wavelength (A):
— It is the distance between two nearest parts of the
wave in the same phase i.e. distance between two
nearest crest or troughs.
Electromagnetic Radiation
iw avelength >:
+ The relationship between wavelength &
frequency can be written as:
c=vA
+ As photon is subjected to energy, so
E=hv=hc/A
iw avelength >:
+ The relationship between wavelength &
frequency can be written as:
c=vA
+ As photon is subjected to energy, so
E=hv=hc/A
Electromagnetic Radiation
Electromagnetic Radiation
Dispersion
Angle
Violet 400 - 420 nm
Indigo 420 - 440 nm
Blue 440 - 490 nm
Green 490 - 570 nm
Dispersion
Angle
Violet 400 - 420 nm
Indigo 420 - 440 nm
Blue 440 - 490 nm
Green 490 - 570 nm
ultra —violet.
180,2 4000 9
180,2 4000 9
rinciples of —
Spectroscopy
Spectroscopy
Principles of Spectroscopy
+ The principle is based on the measurement of
spectrum of a sample containing atoms /
molecules.
+ Spectrum is a graph of intensity of absorbed or
emitted radiation by sample verses frequency
(v) or wavelength (A).
+ Spectrometer is an instrument design to
measure the spectrum of a compound.
+ The principle is based on the measurement of
spectrum of a sample containing atoms /
molecules.
+ Spectrum is a graph of intensity of absorbed or
emitted radiation by sample verses frequency
(v) or wavelength (A).
+ Spectrometer is an instrument design to
measure the spectrum of a compound.
Principles of Spectroscopy
1. Absorption Spectroscopy:
+ An analytical technique which concerns with
the measurement of absorption of
electromagnetic radiation.
+ e.g. UV (185 - 400 nm) / Visible (400 - 800 nm)
Spectroscopy, IR Spectroscopy (0.76 - 15 um)
1. Absorption Spectroscopy:
+ An analytical technique which concerns with
the measurement of absorption of
electromagnetic radiation.
+ e.g. UV (185 - 400 nm) / Visible (400 - 800 nm)
Spectroscopy, IR Spectroscopy (0.76 - 15 um)
Principles of Spectroscopy
2. Emission Spectroscopy:
+ An analytical technique in which emission
(of a particle or radiation) is dispersed
according to some property of the emission
&the amount of dispersion is measured.
* e.g. Mass Spectroscopy
2. Emission Spectroscopy:
+ An analytical technique in which emission
(of a particle or radiation) is dispersed
according to some property of the emission
&the amount of dispersion is measured.
* e.g. Mass Spectroscopy
“The study of interaction of electromagnetic radiation with
molecules/atoms ”.
The study of absorbed radiation by molecule , in the
orm of spectra.
Eg: UV, IR, NMR, colorimetry,
Atomic absorption spectroscopy
=
E
The radiation emitted by molecules can also be
studied to reveal the structure of molecule.
Eg:flame photometry, flourimetry
molecules/atoms ”.
The study of absorbed radiation by molecule , in the
orm of spectra.
Eg: UV, IR, NMR, colorimetry,
Atomic absorption spectroscopy
=
E
The radiation emitted by molecules can also be
studied to reveal the structure of molecule.
Eg:flame photometry, flourimetry
Study of ‘Speetroscapy
inersction J EMR+ATOMS
Changes in energy take place at atomic level
Eg: atomic absorption spectroscopy, flame
photometry:
Interaction of EMR + molsculs
Changes in energy take place at molecular level
Eg: UV, IR, colorimetry
Results in transitions between vibrational, 8
rotational energy levels
inersction J EMR+ATOMS
Changes in energy take place at atomic level
Eg: atomic absorption spectroscopy, flame
photometry:
Interaction of EMR + molsculs
Changes in energy take place at molecular level
Eg: UV, IR, colorimetry
Results in transitions between vibrational, 8
rotational energy levels
UV-visible spectroscopy measure
he response of a sample to ultra
violet and visible range of
electromagnetic radiation.
Molecules have either n,n or o
Electrons.These electrons absorb
UV radiation & undergoes
ransitions from ground state to
excited state.
he response of a sample to ultra
violet and visible range of
electromagnetic radiation.
Molecules have either n,n or o
Electrons.These electrons absorb
UV radiation & undergoes
ransitions from ground state to
excited state.
The absorption of uv radiation brings about the promotion
lof an electron from bonding to antibonding orbital.
The wavelength of radiation is slowly changed from
minimum to maximum in the given region, and the
absorbance at every wavelength is recorded. Then a plot of
lenergy absorbed Vs wavelength is called absorption
spectrum.
The significant features:
Amax (wavelength at which there is a maximum
absorption)
€max (The intensity of maximum absorption)
The UV spectrum depends on
solvents
concentration of solution
lof an electron from bonding to antibonding orbital.
The wavelength of radiation is slowly changed from
minimum to maximum in the given region, and the
absorbance at every wavelength is recorded. Then a plot of
lenergy absorbed Vs wavelength is called absorption
spectrum.
The significant features:
Amax (wavelength at which there is a maximum
absorption)
€max (The intensity of maximum absorption)
The UV spectrum depends on
solvents
concentration of solution
N = 270.5 nm
max
1
1
|
1
Absorption 1
|
|
1
1
|
|
I
1
1 1 1 L 1 1
220 240 260 280 300 320
wavelength in nm ———
fig:- UV spectrum of acetone
max
1
1
|
1
Absorption 1
|
|
1
1
|
|
I
1
1 1 1 L 1 1
220 240 260 280 300 320
wavelength in nm ———
fig:- UV spectrum of acetone
| Interaction of
EMR EE
EMR EE
Interaction of EMR with matter
1. Electronic Energy Levels:
+ At room temperature the molecules are in the
lowest energy levels E,.
+ When the molecules absorb UV-visible light
from EMR, one of the outermost bond / lone
pair electron is promoted to higher energy
state such as E,, E,, ...E,, etc is called as
electronic transition and the difference is as:
AE=hw=E, - Eg Where (n= 1, 2, 3, ... etc)
AE = 35 to 71 kcal/mole
1. Electronic Energy Levels:
+ At room temperature the molecules are in the
lowest energy levels E,.
+ When the molecules absorb UV-visible light
from EMR, one of the outermost bond / lone
pair electron is promoted to higher energy
state such as E,, E,, ...E,, etc is called as
electronic transition and the difference is as:
AE=hw=E, - Eg Where (n= 1, 2, 3, ... etc)
AE = 35 to 71 kcal/mole
Interaction of EMR with matter
2. Vibrational Energy Levels:
These are less energy level than electronic
energy levels.
The spacing between energy levels are
relatively small i.e. 0.01 to 10 kcal/mole.
e.g. when IR radiation is absorbed, molecules
are excited from one vibrational level to
another or it vibrates with higher amplitude.
2. Vibrational Energy Levels:
These are less energy level than electronic
energy levels.
The spacing between energy levels are
relatively small i.e. 0.01 to 10 kcal/mole.
e.g. when IR radiation is absorbed, molecules
are excited from one vibrational level to
another or it vibrates with higher amplitude.
Interaction of EMR with matter
3. Rotational Energy Levels:
+ These energy levels are quantized €: discrete.
+ The spacing between energy levels are even
smaller than vibrational energy levels.
AEvotationa < AE vibrationa < AE etectronic
3. Rotational Energy Levels:
+ These energy levels are quantized €: discrete.
+ The spacing between energy levels are even
smaller than vibrational energy levels.
AEvotationa < AE vibrationa < AE etectronic
Beer’s Law
« When a monochromatic radiation is passed
through a solution, the decrease in the
intensity of radiation with thickness of the
solution is directly proportional to the
intensity of the incident light as well as
concentration of the solution.
+ Let I be the intensity of incident radiation.
x be the thickness of the solution.
C be the concentration of the solution.
Then
« When a monochromatic radiation is passed
through a solution, the decrease in the
intensity of radiation with thickness of the
solution is directly proportional to the
intensity of the incident light as well as
concentration of the solution.
+ Let I be the intensity of incident radiation.
x be the thickness of the solution.
C be the concentration of the solution.
Then
Beer’s Law
Er CT
dx
So; at ikaw
dx
Integrate equation between limit
I= lo at x =O and
I=lIat x=1,
We get,
im ete
Lo
Er CT
dx
So; at ikaw
dx
Integrate equation between limit
I= lo at x =O and
I=lIat x=1,
We get,
im ete
Lo
Beer’s Law
Lo
2.303 log —)=K.C1
Lo K
log = ———Ci
74 2.303
Where, log ELE A Absorbance
7
KL | E Molar extinction
2.303 coefficient
A = E.CI Beer’s Law
Lo
2.303 log —)=K.C1
Lo K
log = ———Ci
74 2.303
Where, log ELE A Absorbance
7
KL | E Molar extinction
2.303 coefficient
A = E.CI Beer’s Law
Beer’s Law
Ar A 2
7 z OR log T log 4 A
I
o
From the equation it is seen that the absorbance
which is also called as optical density (OD) of a solution
in a container of fixed path length is directly
proportional to the concentration of a solution.
Ar A 2
7 z OR log T log 4 A
I
o
From the equation it is seen that the absorbance
which is also called as optical density (OD) of a solution
in a container of fixed path length is directly
proportional to the concentration of a solution.
PRINCIPLES OF
UV - VISIBLE
SPECTROSCOPY
UV - VISIBLE
SPECTROSCOPY
Principle
+ The UV radiation region extends from 10 nm
to 400 nm and the visible radiation region
extends from 400 nm to 800 nm.
Near UV Region: 200 nm to 400 nm
Far UV Region: below 200 nm
+ Far UV spectroscopy is studied under vacuum
condition.
« The common solvent used for preparing
sample to be analyzed is either ethyl alcohol
or hexane.
+ The UV radiation region extends from 10 nm
to 400 nm and the visible radiation region
extends from 400 nm to 800 nm.
Near UV Region: 200 nm to 400 nm
Far UV Region: below 200 nm
+ Far UV spectroscopy is studied under vacuum
condition.
« The common solvent used for preparing
sample to be analyzed is either ethyl alcohol
or hexane.
Electronic
Transitions
Transitions
The possible electronic transitions can
graphically shown as:
©” (anti-bonding)
x" (anti-bonding)
n (non-bonding)
>
a
1-2
pra]
2
a
ax (bonding)
GS (bonding)
graphically shown as:
©” (anti-bonding)
x" (anti-bonding)
n (non-bonding)
>
a
1-2
pra]
2
a
ax (bonding)
GS (bonding)
Observed electronic transitions
Here is a graphical representation
Atomic orbital Atomic orbital
S
Here is a graphical representation
Atomic orbital Atomic orbital
S
The possible electronic transitions are
* O — o* transition
* O — o* transition
* 6 — o* transition
+ & electron from orbital is excited to
corresponding anti-bonding orbital o*.
« The energy required is large for this
transition.
* e.g. Methane (CH,) has C-H bond only and
can undergo o > o* transition and shows
absorbance maxima at 125 nm.
+ & electron from orbital is excited to
corresponding anti-bonding orbital o*.
« The energy required is large for this
transition.
* e.g. Methane (CH,) has C-H bond only and
can undergo o > o* transition and shows
absorbance maxima at 125 nm.
* an — rn“ transition
- x electron in a bonding orbital is excited to
corresponding anti-bonding orbital nı*.
+ Compounds containing multiple bonds like
alkenes, alkynes, carbonyl, nitriles, aromatic
compounds, etc undergo n > rı* transitions.
+ e.g. Alkenes generally absorb in the region
170 to 205 nm.
- x electron in a bonding orbital is excited to
corresponding anti-bonding orbital nı*.
+ Compounds containing multiple bonds like
alkenes, alkynes, carbonyl, nitriles, aromatic
compounds, etc undergo n > rı* transitions.
+ e.g. Alkenes generally absorb in the region
170 to 205 nm.
* n — o* transition
+ Saturated compounds containing atoms with
lone pair of electrons like O, N, S and
halogens are capable of n > o* transition.
+ These transitions usually requires less energy
than o > o* transitions.
« The number of organic functional groups
with n > o* peaks in UV region is small (150
— 250 nm).
+ Saturated compounds containing atoms with
lone pair of electrons like O, N, S and
halogens are capable of n > o* transition.
+ These transitions usually requires less energy
than o > o* transitions.
« The number of organic functional groups
with n > o* peaks in UV region is small (150
— 250 nm).
| * n — nr“ transition
« An electron from non-bonding orbital is
promoted to anti-bonding rı* orbital.
+ Compounds containing double bond
involving hetero atoms (C=O, C=N, N=O)
undergo such transitions.
* n > n* transitions require minimum energy
and show absorption at longer wavelength
around 300 nm.
« An electron from non-bonding orbital is
promoted to anti-bonding rı* orbital.
+ Compounds containing double bond
involving hetero atoms (C=O, C=N, N=O)
undergo such transitions.
* n > n* transitions require minimum energy
and show absorption at longer wavelength
around 300 nm.
ls + o —>x* transition |
| & | * a — o* transition
«These electronic transitions are forbidden
transitions & are only theoretically possible.
-Thus, n > n* £ n > n* electronic transitions
show absorption in region above 200 nm
which is accessible to UV-visible
spectrophotometer.
«The UV spectrum is of only a few broad of
absorption.
| & | * a — o* transition
«These electronic transitions are forbidden
transitions & are only theoretically possible.
-Thus, n > n* £ n > n* electronic transitions
show absorption in region above 200 nm
which is accessible to UV-visible
spectrophotometer.
«The UV spectrum is of only a few broad of
absorption.
The transitions with the values of extinction co-
efficient more than 104 are usually called allowed
transitions.
They generally arise due to
TT-TT Trar on.
Eg: In 1,3-butadiene molar extinction co-efficient is
very high i.e.21000
efficient more than 104 are usually called allowed
transitions.
They generally arise due to
TT-TT Trar on.
Eg: In 1,3-butadiene molar extinction co-efficient is
very high i.e.21000
These transitions are as a result of the excitation of
one electron from the lone pair present on the hetero
atom to an anti bonding n* orbital.
Eg: carbonyl compounds
Molar extinction co-efficient value is 104
one electron from the lone pair present on the hetero
atom to an anti bonding n* orbital.
Eg: carbonyl compounds
Molar extinction co-efficient value is 104
Terms used
in
UV / Visible
Spectroscopy
in
UV / Visible
Spectroscopy
Chromophore
The part of a molecule responsible for imparting
color, are called as chromospheres.
OR
The functional groups containing multiple bonds
capable of absorbing radiations above 200 nm
due ton > n* &n > n* transitions.
e.g. NO), N=O, C=O, C=N, C=N, C=C, C=S, etc
The part of a molecule responsible for imparting
color, are called as chromospheres.
OR
The functional groups containing multiple bonds
capable of absorbing radiations above 200 nm
due ton > n* &n > n* transitions.
e.g. NO), N=O, C=O, C=N, C=N, C=C, C=S, etc
Chromophore
To interpretate UV — visible spectrum following
points should be noted:
ae
Non-conjugated alkenes show an intense
absorption below 200 nm & are therefore
inaccessible to UV spectrophotometer.
Non-conjugated carbonyl group compound
give a weak absorption band in the 200 - 300
nm region.
To interpretate UV — visible spectrum following
points should be noted:
ae
Non-conjugated alkenes show an intense
absorption below 200 nm & are therefore
inaccessible to UV spectrophotometer.
Non-conjugated carbonyl group compound
give a weak absorption band in the 200 - 300
nm region.
Chromophore
e.g. ff Acetone which has A,,,, = 279 nm,
nba re
and that cyclohexane has Aa = 291 nm.
When double bonds are conjugated in a
compound A,,,, is shifted to longer wavelength.
e.g. 1,5 - hexadiene has À,,,, = 178 nm
2,4 - hexadiene has A... = 227 nm
CH CH
FRE NN 2 He N mu 3
e.g. ff Acetone which has A,,,, = 279 nm,
nba re
and that cyclohexane has Aa = 291 nm.
When double bonds are conjugated in a
compound A,,,, is shifted to longer wavelength.
e.g. 1,5 - hexadiene has À,,,, = 178 nm
2,4 - hexadiene has A... = 227 nm
CH CH
FRE NN 2 He N mu 3
Chromophore
3. Conjugation of C=C and carbonyl group shifts
the Amax of both groups to longer wavelength.
e.g. Ethylene has Ama, = 171 nm =
max = 279 nm N
H,C—=CH, HAC cu,
Crotonaldehyde has A „ax = 290 nm
Acetone has A
O
Il
Cc
H20—= EN
A CH;
3. Conjugation of C=C and carbonyl group shifts
the Amax of both groups to longer wavelength.
e.g. Ethylene has Ama, = 171 nm =
max = 279 nm N
H,C—=CH, HAC cu,
Crotonaldehyde has A „ax = 290 nm
Acetone has A
O
Il
Cc
H20—= EN
A CH;
Auxochrome
The functional groups attached to a
chromophore which modifies the ability of the
chromophore to absorb light , altering the
wavelength or intensity of absorption.
OR
The functional group with non-bonding electrons
that does not absorb radiation in near UV region
but when attached to a chromophore alters the
wavelength & intensity of absorption.
The functional groups attached to a
chromophore which modifies the ability of the
chromophore to absorb light , altering the
wavelength or intensity of absorption.
OR
The functional group with non-bonding electrons
that does not absorb radiation in near UV region
but when attached to a chromophore alters the
wavelength & intensity of absorption.
Auxochrome
e.g. Benzene Amax = 255 nm DI
OH
Phenol Amax = 270 nm ES
NH,
Aniline A, = 280 nm
max
e.g. Benzene Amax = 255 nm DI
OH
Phenol Amax = 270 nm ES
NH,
Aniline A, = 280 nm
max
Absorption
& Intensity
Shifts
& Intensity
Shifts
a Bathochromic Shift (Red Shift)
+ When absorption maxima A Of
compound shifts to longer wavelength, it is
known as bathochromic shift or red shift.
- The effect is due to presence of an auxochrome
or by the change of solvent.
* e.g. An auxochrome group like —OH, -OCHz
causes absorption of compound at longer
wavelength.
+ When absorption maxima A Of
compound shifts to longer wavelength, it is
known as bathochromic shift or red shift.
- The effect is due to presence of an auxochrome
or by the change of solvent.
* e.g. An auxochrome group like —OH, -OCHz
causes absorption of compound at longer
wavelength.
&-
+ In alkaline medium, p-nitrophenol shows red
shift. Because negatively charged oxygen
delocalizes more effectively than the unshared
pair of electron.
ASEO
medium =
o
Sitio baña]
Amax = 255 nm Amax = 265 nm
max
+ In alkaline medium, p-nitrophenol shows red
shift. Because negatively charged oxygen
delocalizes more effectively than the unshared
pair of electron.
ASEO
medium =
o
Sitio baña]
Amax = 255 nm Amax = 265 nm
max
ES Hypsochromic Shift (Blue Shift)
+ When absorption maxima An.) of a
compound shifts to shorter wavelength, it is
known as hypsochromic shift or blue shift.
- The effect is due to presence of an group
causes removal of conjugation or by the
change of solvent.
+ When absorption maxima An.) of a
compound shifts to shorter wavelength, it is
known as hypsochromic shift or blue shift.
- The effect is due to presence of an group
causes removal of conjugation or by the
change of solvent.
2 l ae Shift (Blue Shift) >
3 ” :
+ Aniline shows blue shift in acidic medium, it
loses conjugation.
NH, im NH ,° CI”
H
co Acidic
medium
Aniline
A = 280 nm A = 265 nm
max max
3 ” :
+ Aniline shows blue shift in acidic medium, it
loses conjugation.
NH, im NH ,° CI”
H
co Acidic
medium
Aniline
A = 280 nm A = 265 nm
max max
Par Hyperchromic Effect
« When absorption intensity (e) of a compound is
increased, it is known as hyperchromic shift.
elf auxochrome introduces to the
compound, the intensity of absorption
increa AS =
| = =
N N CH,
Pyridine 2-methyl pyridine
Amax = 257 nm Amax = 260 nm
« When absorption intensity (e) of a compound is
increased, it is known as hyperchromic shift.
elf auxochrome introduces to the
compound, the intensity of absorption
increa AS =
| = =
N N CH,
Pyridine 2-methyl pyridine
Amax = 257 nm Amax = 260 nm
El Hypochromic Effect
+ When absorption intensity (£) of a compound is
decreased, it is known as hypochromic shift.
Naphthalene 2-methyl naphthalene
€ = 19000 e = 10250
+ When absorption intensity (£) of a compound is
decreased, it is known as hypochromic shift.
Naphthalene 2-methyl naphthalene
€ = 19000 e = 10250
Shifts and Effects
Hyperchromic shift
Blue
shift
Red
shift
Absorbance (A)
Hypochromic shift
1
A
max
Wavelength (A)
Hyperchromic shift
Blue
shift
Red
shift
Absorbance (A)
Hypochromic shift
1
A
max
Wavelength (A)
INSTRUMENTATION OF UV-VISIBLE
SPECTROPHOTOMETRY
SPECTROPHOTOMETRY
Content
» Introduction
» Components of spectrophotometry.
» Instrument design.
en
» Introduction
» Components of spectrophotometry.
» Instrument design.
en
INTRODUCTION
» Absorption spectrophotometry in the ultraviolet and visible
region is considered to be one of the oldest physical method
for quantitative analysis and structural elucidation,
» Wavelength
+ UV- 200-400nm
+ VISIBLE- 400-800nm
nn...
» Absorption spectrophotometry in the ultraviolet and visible
region is considered to be one of the oldest physical method
for quantitative analysis and structural elucidation,
» Wavelength
+ UV- 200-400nm
+ VISIBLE- 400-800nm
nn...
INSTRUMENTS
» PHOTOMETER
» SPECTOPHOTOMETER
» COLORIMETER
2 PHOTOMETER: An instrument for measuring the
intensity of light or the relative intensity of a pair of
lights. Also called an illuminometer. It utilizes filter
to isolate a narrow wavelength region.
en
» PHOTOMETER
» SPECTOPHOTOMETER
» COLORIMETER
2 PHOTOMETER: An instrument for measuring the
intensity of light or the relative intensity of a pair of
lights. Also called an illuminometer. It utilizes filter
to isolate a narrow wavelength region.
en
= SPECTOPHOTOMETER: An instrument measures the
ratio, or a function of the two, of the radiant power of two
EM beams over a large wavelength region. It utilizes
dispersing element (Prisms/Gratings) instead of filters, to
scan large wavelength region.
= COLORIMETER: An instrument which is used for
measuring absorption in the visible region is generally
called colorimeter.
ratio, or a function of the two, of the radiant power of two
EM beams over a large wavelength region. It utilizes
dispersing element (Prisms/Gratings) instead of filters, to
scan large wavelength region.
= COLORIMETER: An instrument which is used for
measuring absorption in the visible region is generally
called colorimeter.
COMPONENTS OF UV-VIS SPECTROPHOTOMETER
» source of radiant energy.
» Collimating system
» monochromator system.
» sample holder or container to hold sample.
» detector system of collecting transmitted radiation.
» suitable amplifier or readout device.
en
» source of radiant energy.
» Collimating system
» monochromator system.
» sample holder or container to hold sample.
» detector system of collecting transmitted radiation.
» suitable amplifier or readout device.
en
collimating dispersing
light lens element
un container detector
readout
light lens element
un container detector
readout
SOURCE OF RADIANT ENERGY
REQUIREMENTS OF AN IDEAL SOURCE
¥ It should be stable and should not allow fluctuations,
+ It should emit light of continuous spectrum of high and
uniform intensity over the entire wavelength region in which
it’s used.
+ It should provide incident light of sufficient intensity for the
transmitted energy to be detected at the end of optic path.
+ It should not show fatigue on continued use.
en
REQUIREMENTS OF AN IDEAL SOURCE
¥ It should be stable and should not allow fluctuations,
+ It should emit light of continuous spectrum of high and
uniform intensity over the entire wavelength region in which
it’s used.
+ It should provide incident light of sufficient intensity for the
transmitted energy to be detected at the end of optic path.
+ It should not show fatigue on continued use.
en
FOR VISIBLE RADIATION
TUNGSTEN HALOGEN LAMP
» Its construction is similar to a house hold lamp.
» The bulb contains a filament of Tungsten fixed in evacuated
condition and then filled with inert gas.
» The filament can be heated up to 3000 k, beyond this
Tungsten starts sublimating.
» Itis used when polychromatic light is required. To prevent this
introduced
pme amount of halogen i
TUNGSTEN HALOGEN LAMP
» Its construction is similar to a house hold lamp.
» The bulb contains a filament of Tungsten fixed in evacuated
condition and then filled with inert gas.
» The filament can be heated up to 3000 k, beyond this
Tungsten starts sublimating.
» Itis used when polychromatic light is required. To prevent this
introduced
pme amount of halogen i
y Sublimated form of tungsten reacts with Iodine to
form Tungsten —Iodine complex.
Which migrates back to the hot filament where it
decomposes and Tungsten get deposited.
vy DEMERIT:
y It emits the major portion of its radiant energy in
near IR region of the spectrum.
| e E
form Tungsten —Iodine complex.
Which migrates back to the hot filament where it
decomposes and Tungsten get deposited.
vy DEMERIT:
y It emits the major portion of its radiant energy in
near IR region of the spectrum.
| e E
OURCE FOR UV RADIATIO
HYDROGEN DISCHARGE LAMP:
» In Hydrogen discharge lamp pair of electrodes is enclosed in a
glass tube (provided with silica or quartz window for UV
radiation to pass trough) filled with hydrogen gas.
» When current is passed trough these electrodes maintained at
high voltage, discharge of electrons occurs which excites
hydrogen molecules which in tum cause emission of UV
radiations in near UV region.
r They are stable and robust.
HYDROGEN DISCHARGE LAMP:
» In Hydrogen discharge lamp pair of electrodes is enclosed in a
glass tube (provided with silica or quartz window for UV
radiation to pass trough) filled with hydrogen gas.
» When current is passed trough these electrodes maintained at
high voltage, discharge of electrons occurs which excites
hydrogen molecules which in tum cause emission of UV
radiations in near UV region.
r They are stable and robust.
NON DISCHARGE LAMP:
» It possesses two tungsten electrodes separated by some distance.
» These are enclosed in a glass tube (for visible) with quartz or fused
silica and xenon gas is filled under pressure.
» An intense are is formed between electrodes by appl
high
voltage. This is a good source of continuous plus additional intense
radiation. Its intensity is higher than the hydrogen discharge lamp.
DEME
» The lamp since operates at high voltage becomes very hot during
on.
77
operation and hence needs thermal insula
» It possesses two tungsten electrodes separated by some distance.
» These are enclosed in a glass tube (for visible) with quartz or fused
silica and xenon gas is filled under pressure.
» An intense are is formed between electrodes by appl
high
voltage. This is a good source of continuous plus additional intense
radiation. Its intensity is higher than the hydrogen discharge lamp.
DEME
» The lamp since operates at high voltage becomes very hot during
on.
77
operation and hence needs thermal insula
MERCURY ARC LAMP
In mercury arc lamp, mercury vapor is stored under high
pressure and excitation of mercury atoms is done by electric
discharge.
DEMERIT:
Not suitable for continuous spectral studies,(because it doesn’t
give continuous radiations).
In mercury arc lamp, mercury vapor is stored under high
pressure and excitation of mercury atoms is done by electric
discharge.
DEMERIT:
Not suitable for continuous spectral studies,(because it doesn’t
give continuous radiations).
COLLIMATING SYSTE
The radiation emitted by the source is collimated (made
parallel) by lenses, mirrors and slits. )
(
LENSES: ==>
> Materials used for the lenses must be transparent to the
radiation being used.
> Ordinary silicate glass transmits between 350 to 3000 nm
and is suitable for visible and near IR region.
or fused
¡sed as a material for lenses to work
00nm,
The radiation emitted by the source is collimated (made
parallel) by lenses, mirrors and slits. )
(
LENSES: ==>
> Materials used for the lenses must be transparent to the
radiation being used.
> Ordinary silicate glass transmits between 350 to 3000 nm
and is suitable for visible and near IR region.
or fused
¡sed as a material for lenses to work
00nm,
MIRRORS
» These are used to reflect, focus or collimate light beams in
spectrophotometer.
» To minimize the light loss, mirrors are aluminized on their
front surfaces.
» These are used to reflect, focus or collimate light beams in
spectrophotometer.
» To minimize the light loss, mirrors are aluminized on their
front surfaces.
SLITS:
» Slit is an important device in resolving polychromatic
radiation into monochromatic radiation.
» To achieve this, entrance slit and exit slit are used.
» The width of slit plays an important role in resolution of
polychromatic radiation.
» Slit is an important device in resolving polychromatic
radiation into monochromatic radiation.
» To achieve this, entrance slit and exit slit are used.
» The width of slit plays an important role in resolution of
polychromatic radiation.
MONOCHROMATORS
It is a device used to isolate the radiation of the desired
wavelength from wavelength of the continuous spectra,
Following types of monochromatic devices are used.
1. Filters
Prisms:
3 Gratings
It is a device used to isolate the radiation of the desired
wavelength from wavelength of the continuous spectra,
Following types of monochromatic devices are used.
1. Filters
Prisms:
3 Gratings
FILTERS
Selection of filters is usually done on a compromise between
peak transmittance and band pass width; the former should be as
high as possible and latter as narrow as possible.
1. Absorption filters. works by selective absorption of
unwanted radiation and transmits the radiation which is
required.
Examples- Glass and Gelat
Selection of filters is usually done on a compromise between
peak transmittance and band pass width; the former should be as
high as possible and latter as narrow as possible.
1. Absorption filters. works by selective absorption of
unwanted radiation and transmits the radiation which is
required.
Examples- Glass and Gelat
Selection of absorption filter is done according to
the following procedure:
» Draw a filter wheel.
430 nm
» Write the color VIBGYOR in clockwise or anticlockwise
manner, omitting Indigo.
the following procedure:
» Draw a filter wheel.
430 nm
» Write the color VIBGYOR in clockwise or anticlockwise
manner, omitting Indigo.
» If solution to be analyzed is BLUE in color a filter having a
complimentary color ORANGE is used in the a
alysis.
» Similarly, we can select the required filter in colorimeter, based
upon the color of the solution,
complimentary color ORANGE is used in the a
alysis.
» Similarly, we can select the required filter in colorimeter, based
upon the color of the solution,
» An Absorption glass filter is made of solid sheet of glass that
has been colored by pigments which Is dissolved or dispersed
in the glass.
» The color in the glass filters are produced by incorporating
metal oxides like (V, Cr, Mn, Fe, Ni, Co, Cu etc.).
has been colored by pigments which Is dissolved or dispersed
in the glass.
» The color in the glass filters are produced by incorporating
metal oxides like (V, Cr, Mn, Fe, Ni, Co, Cu etc.).
» Gelatin filter is an example of absorption filter prepared by
adding organic pigments; here instead of solid glass sheets thin
gelatin sheets are used. Gelatin filters are not use now days.
» Ittends to deteriorate with time and gets affected by the heat and
moisture. The color of the dye gets bleached.
adding organic pigments; here instead of solid glass sheets thin
gelatin sheets are used. Gelatin filters are not use now days.
» Ittends to deteriorate with time and gets affected by the heat and
moisture. The color of the dye gets bleached.
MERIT:
» Simple in construction
» Cheaper
» Selection of the filter is easy
ss accurate
» Band pass (bandwidth) is more (+20-30nm) ie. if we have to
measure at 400n: e get radiation from 370-430nm. Hence
less accurate results are obtained.
| e ;
» Simple in construction
» Cheaper
» Selection of the filter is easy
ss accurate
» Band pass (bandwidth) is more (+20-30nm) ie. if we have to
measure at 400n: e get radiation from 370-430nm. Hence
less accurate results are obtained.
| e ;
TERFERENCE FILTERS
> Works on the interference phenomenon, causes rejection of
unwanted wavelength by selective reflection
> It is constructed by using two parallel glass plates, which are
silvered internally and separated by thin film of dielectric
material of different (CaF, SiO, MgF,) refractive index. These
filters have a band pass of 10-15nm with peak transmittance of
40-60%.
en
> Works on the interference phenomenon, causes rejection of
unwanted wavelength by selective reflection
> It is constructed by using two parallel glass plates, which are
silvered internally and separated by thin film of dielectric
material of different (CaF, SiO, MgF,) refractive index. These
filters have a band pass of 10-15nm with peak transmittance of
40-60%.
en
EAU
Merits -
> Provide greater transmittance and narrower band pass (10-
15nm) as compare to absorption filter.
> Inexpensive
> Additional filters can be used to cut off undesired wavelength.
Merits -
> Provide greater transmittance and narrower band pass (10-
15nm) as compare to absorption filter.
> Inexpensive
> Additional filters can be used to cut off undesired wavelength.
PRISM
Prism is made from glass, Quartz or fused silica.
Quartz or fused silica is the choice of material of UV
spectrum.
When white light is passed through glass prism, dispersion
of polychromatic light in rainbow occurs. Now by rotation
of the prism different wavelengths of the spectrum can be
made to pass through in exit slit on the sample.
The effective wavelength depends on the dispersive power
of prism material and the optical angle of the prism.
Prism is made from glass, Quartz or fused silica.
Quartz or fused silica is the choice of material of UV
spectrum.
When white light is passed through glass prism, dispersion
of polychromatic light in rainbow occurs. Now by rotation
of the prism different wavelengths of the spectrum can be
made to pass through in exit slit on the sample.
The effective wavelength depends on the dispersive power
of prism material and the optical angle of the prism.
PRISM
White
Light
Dispersion
Angle
White
Light
Dispersion
Angle
+ There are two types of mounting in an instrument one is called
“Cornu type'(refractive), which has an optical angle of 60°
and its adjusted such that on rotation the emerging light is
A
allowed to fall on exit slit.
wee
Cama pe
+ The other type is called “Littrow type”(reflective), which has
optical angle 30° and its one surface is aluminized with
reflected light back to pass through prism and to emerge on the
same side of the light source i.e. light doesn’t pass through the
prism on other side. minos
| Fe mm
“Cornu type'(refractive), which has an optical angle of 60°
and its adjusted such that on rotation the emerging light is
A
allowed to fall on exit slit.
wee
Cama pe
+ The other type is called “Littrow type”(reflective), which has
optical angle 30° and its one surface is aluminized with
reflected light back to pass through prism and to emerge on the
same side of the light source i.e. light doesn’t pass through the
prism on other side. minos
| Fe mm
GR.
GS
> Are most effective one in converting a polychromatic light to
monochromatic light. As a resolution of +/- 0.Inm could be
achieved by using gratings, they are commonly used in
spectrophotometers.
> Gratings are of two types.
1. Diffraction grating.
2. Transmission gratings.
GS
> Are most effective one in converting a polychromatic light to
monochromatic light. As a resolution of +/- 0.Inm could be
achieved by using gratings, they are commonly used in
spectrophotometers.
> Gratings are of two types.
1. Diffraction grating.
2. Transmission gratings.
Diffraction Grating
> More refined dispersion of light is obtained by means of
diffraction gratings.
> These consist of large number of parallel lines ( grooves)
about 15000-30000/ inch is ruled on highly polished surface of
aluminum,
» these gratings are replica made from master gratings by
coating the original master grating with a epoxy resin and are
r removed after setting
> More refined dispersion of light is obtained by means of
diffraction gratings.
> These consist of large number of parallel lines ( grooves)
about 15000-30000/ inch is ruled on highly polished surface of
aluminum,
» these gratings are replica made from master gratings by
coating the original master grating with a epoxy resin and are
r removed after setting
» To make the surface reflective, a deposit of aluminum is made
on the surface. In order to minimize to greater amounts of
scattered radiation and appearance of unwanted radiation of
other spectral orders, the gratings are blazed to concentrate the
radiation into a single order.
on the surface. In order to minimize to greater amounts of
scattered radiation and appearance of unwanted radiation of
other spectral orders, the gratings are blazed to concentrate the
radiation into a single order.
Transmission grating
» It is similar to diffraction grating but refraction takes place
instead of reflection, Refraction produces reinforcement. this
occurs when radiation transmitted through grating reinforces
with the partially refracted radiation.
» It is similar to diffraction grating but refraction takes place
instead of reflection, Refraction produces reinforcement. this
occurs when radiation transmitted through grating reinforces
with the partially refracted radiation.
Advantages
> Grating gives higher and linear dispersions compared to
prism monochromator.
> Can be used over wide wavelength ranges.
> Gratings can be constructed with materials like
aluminium which is resistant to atmospheric moisture,
> Provide light of narrow wavelength.
> No loss of energy due to absorption.
en
> Grating gives higher and linear dispersions compared to
prism monochromator.
> Can be used over wide wavelength ranges.
> Gratings can be constructed with materials like
aluminium which is resistant to atmospheric moisture,
> Provide light of narrow wavelength.
> No loss of energy due to absorption.
en
[Comparison
Prism
Grating
Made of ise Visible Grooved on highly polished
Quanz/fused silica-: UV |surfacelike alumina
Alkali halide:- IR.
Working Principle | Angle of Incident Taw of diffraction
4 (sinissind)
> Prisms
dispersion hence
overlap of spectral order
> It can't be used over
consideration wavelength|
ranges,
> Prisms are not sturdy and
long lasting.
rating gives Tier disper
hhence overlap of spectral
order
> It cn be used over
considerable wavelength
ranges.
> Grating are sturdy and long
lasting
Prism
Grating
Made of ise Visible Grooved on highly polished
Quanz/fused silica-: UV |surfacelike alumina
Alkali halide:- IR.
Working Principle | Angle of Incident Taw of diffraction
4 (sinissind)
> Prisms
dispersion hence
overlap of spectral order
> It can't be used over
consideration wavelength|
ranges,
> Prisms are not sturdy and
long lasting.
rating gives Tier disper
hhence overlap of spectral
order
> It cn be used over
considerable wavelength
ranges.
> Grating are sturdy and long
lasting
SAMPLE HOLDERS/CUVETTES
> The cells or cuvettes are used for handling liquid samples.
> The cell may either be rectangular or cylindrical in nature.
> For study in UV region; the cells are prepared from quartz or
fused silica whereas color corrected fused glass is used for
visible region
> The surfaces of absorption cells must be kept scrupulously
clean. No fingerprints or blotches should be present on cells.
> Cleaning is carried out washing with distilled water or with
> The cells or cuvettes are used for handling liquid samples.
> The cell may either be rectangular or cylindrical in nature.
> For study in UV region; the cells are prepared from quartz or
fused silica whereas color corrected fused glass is used for
visible region
> The surfaces of absorption cells must be kept scrupulously
clean. No fingerprints or blotches should be present on cells.
> Cleaning is carried out washing with distilled water or with
Sample holder
DETECTORS
> Device which converts light energy into electrical signals, that
are displayed on readout devices.
> The transmitted radiation falls on the detector which
determines the intensity of radiation absorbed by sample
The following types of detectors are employed in instrumentation
of absorption spectrophotometer
1. Barrier layer cell/Photovoltaic cell
3. Photomultiplier tube
Phototubes/ Photo emissive tube
> Device which converts light energy into electrical signals, that
are displayed on readout devices.
> The transmitted radiation falls on the detector which
determines the intensity of radiation absorbed by sample
The following types of detectors are employed in instrumentation
of absorption spectrophotometer
1. Barrier layer cell/Photovoltaic cell
3. Photomultiplier tube
Phototubes/ Photo emissive tube
Requirements of an ideal detector:-
» I should give quantitative response.
» It should have high sensitivity and low noise level.
» It should have a short response time.
» It should provide signal or response quantitative to wide
spectrum of radiation received.
» I should give quantitative response.
» It should have high sensitivity and low noise level.
» It should have a short response time.
» It should provide signal or response quantitative to wide
spectrum of radiation received.
Barrier layer cell/Photovoltaic cell
> The detector has a thin film metallic layer coated with silver or
gold and acts as an electrode.
> Italso has a metal base plate which acts as another electrode.
> These two layers are separated by a semiconductor layer of
selenium.
| | | cotector ring (- ve)
Hypothetical D transparent layer
‘aver
B seientom layer
A base plate
> The detector has a thin film metallic layer coated with silver or
gold and acts as an electrode.
> Italso has a metal base plate which acts as another electrode.
> These two layers are separated by a semiconductor layer of
selenium.
| | | cotector ring (- ve)
Hypothetical D transparent layer
‘aver
B seientom layer
A base plate
> When light radiation falls on selenium layer, electrons become
mobile and are taken up by transparent metal layer.
> This cre
es a potential difference between two electrodes $e
causes the flow of current
> When it is connected to galvanometer, a flow of current
observed which is proportional to the intensity and wavelength
of light falling on it.
haa > | sate
mobile and are taken up by transparent metal layer.
> This cre
es a potential difference between two electrodes $e
causes the flow of current
> When it is connected to galvanometer, a flow of current
observed which is proportional to the intensity and wavelength
of light falling on it.
haa > | sate
Photo Tubes/Photoemissive Tubes
Photo tubes
Photo Tubes/Photoemissive Tubes
> Consists of a evacuated glass tube with a photocathode and a
collector anode.
> The surface of photocathode is coated with a layer of elements
like cesium, silver oxide or mixture of them.
> When radiant energy falls on photosensitive cathode, electrons
are emitted which are attracted to anode causing current to
flow.
> More sensitive compared to barrier layer cell and therefore
widely used.
> Consists of a evacuated glass tube with a photocathode and a
collector anode.
> The surface of photocathode is coated with a layer of elements
like cesium, silver oxide or mixture of them.
> When radiant energy falls on photosensitive cathode, electrons
are emitted which are attracted to anode causing current to
flow.
> More sensitive compared to barrier layer cell and therefore
widely used.
Photo Multiplier Tubes
» The principle employed in this detector i
that, multiplication
of photoelectrons by secondary emission of electrons.
» In a vacuum tube, a primary photo-cathode is fixed which
receives radiation from the sample.
» Some eight to ten dynod
are fixed each with increasing
potential of 75-100V hig!
er than preceding one.
» Near the last dynode is fixed an anode or electron collector
electrode.
» Photo-multiplier is extremely sensitive to light and is best
weaker or low radiation is received
» The principle employed in this detector i
that, multiplication
of photoelectrons by secondary emission of electrons.
» In a vacuum tube, a primary photo-cathode is fixed which
receives radiation from the sample.
» Some eight to ten dynod
are fixed each with increasing
potential of 75-100V hig!
er than preceding one.
» Near the last dynode is fixed an anode or electron collector
electrode.
» Photo-multiplier is extremely sensitive to light and is best
weaker or low radiation is received
Photo Multiplier Tubes
STRUMENT DESIG)
> Depending upon the monochromators (filters or dispersing
device) used to isolate and transmit a narrow beam of radiant
energy from the incident light determines whether the
instrument is classified as Photometer or a Spectrophotometer.
> Spectrophotometers used here detects the percentage
transmittance of light radiation, when light of certain
intensity & frequency range is passed through the sample.
> Both can be a single beam or double beam optical system.
> Depending upon the monochromators (filters or dispersing
device) used to isolate and transmit a narrow beam of radiant
energy from the incident light determines whether the
instrument is classified as Photometer or a Spectrophotometer.
> Spectrophotometers used here detects the percentage
transmittance of light radiation, when light of certain
intensity & frequency range is passed through the sample.
> Both can be a single beam or double beam optical system.
SINGLE BEAM_SPECTROPHOTOMETER
+ Light from the source is carried through lens and/or through
aperture to pass through a suitable filter.
+ The type of filter to be used is governed by the colour of the
solution,
+ The sample solution to be analysed is placed in cuvettes.
| e
+ Light from the source is carried through lens and/or through
aperture to pass through a suitable filter.
+ The type of filter to be used is governed by the colour of the
solution,
+ The sample solution to be analysed is placed in cuvettes.
| e
Single beam instrument
LENS
SOURCE ENTRANCE SUIT
DECLUDER ==" Lens
RATING
LENS
SOURCE ENTRANCE SUIT
DECLUDER ==" Lens
RATING
» After passing through the solution, the light strikes the surface
of detector (barrier-layer cell or phototube) and produces
electrical current,
» The output of current is measured by the deflection of needle
of light-spot galvanometer or micro ammeter. This meter is
calibrated in terms of transmittance as well as optical density.
The readings of solution of both standard and unknown are
recorded in optical density units after adjusting instrument to a
reagent blank.
of detector (barrier-layer cell or phototube) and produces
electrical current,
» The output of current is measured by the deflection of needle
of light-spot galvanometer or micro ammeter. This meter is
calibrated in terms of transmittance as well as optical density.
The readings of solution of both standard and unknown are
recorded in optical density units after adjusting instrument to a
reagent blank.
Single beam instrument
DOUBLE BEAM UV-VIS
'ECTROPHOTOMETER
» Double beam instrument is the one in which two beams are
formed in the space by a U shaped mirror called as beam
splitter or beam chopper .
» Chopper is a device consisting of a circular disc. One third of
the disc is opaque and one third is transparent, remaining one
third is mirrored. It splits the monochromatic beam of light
into two beams of equal intensities.
'ECTROPHOTOMETER
» Double beam instrument is the one in which two beams are
formed in the space by a U shaped mirror called as beam
splitter or beam chopper .
» Chopper is a device consisting of a circular disc. One third of
the disc is opaque and one third is transparent, remaining one
third is mirrored. It splits the monochromatic beam of light
into two beams of equal intensities.
Reference
Detector 2
-|-1,
Detector 1
A
Fi
Mirror envers
¡ire N
Viens 1
Half mirar
Mirror 2 Sample
ave
Mirar 3 = Viens 2
Detector 2
-|-1,
Detector 1
A
Fi
Mirror envers
¡ire N
Viens 1
Half mirar
Mirror 2 Sample
ave
Mirar 3 = Viens 2
Double Beam
NS
NS
Advantages of single & double beam spectrophotometer
Single bean
» Simple in construction, Easy to use and economical
Double beam-
» It facilitates rapid scanning over wide À region.
» Fluctuations due to radiation source are minimised.
» It doesn’t require adjustment of the transmittance at 0% and
100% at each wavelength.
» It gives ratio of intensities of sample & reference beams
simultaneously.
Single bean
» Simple in construction, Easy to use and economical
Double beam-
» It facilitates rapid scanning over wide À region.
» Fluctuations due to radiation source are minimised.
» It doesn’t require adjustment of the transmittance at 0% and
100% at each wavelength.
» It gives ratio of intensities of sample & reference beams
simultaneously.
Disadvantages
Single beam
» Any fluctuation in the intensity of radiation sources affects the
absorbance.
» Continuous spectrum is not obtained.
Double beam
» Construction is complicated.
» Instrument is expensive.
| e
Single beam
» Any fluctuation in the intensity of radiation sources affects the
absorbance.
» Continuous spectrum is not obtained.
Double beam
» Construction is complicated.
» Instrument is expensive.
| e
COMPARISON:
SL.
NO
SINGLE BEAM
INSTRUMENT
DOUBL BEAM
INSTRUMENT
Calibration should be
done with blank every
time, before measuring
the absorbance or
transmittance of sample
Calibration is done
only in the beginning.
|
SL.
NO
SINGLE BEAM
INSTRUMENT
DOUBL BEAM
INSTRUMENT
Calibration should be
done with blank every
time, before measuring
the absorbance or
transmittance of sample
Calibration is done
only in the beginning.
|
2] Radiant energy intensity
changes with fluctuation
It permits a large degree
of inherent
amount of transmitted
light reaching the detector
of voltage. compensation for
fluctuations in the
intensity of the radiant
energy.
3 | It measure the total It measures the
percentage of light
absorbed by the sample.
Ee
changes with fluctuation
It permits a large degree
of inherent
amount of transmitted
light reaching the detector
of voltage. compensation for
fluctuations in the
intensity of the radiant
energy.
3 | It measure the total It measures the
percentage of light
absorbed by the sample.
Ee
ES
In single beam it's not
possible to compare blank
and sample together.
In double beam it’s
possible to do direct one
step comparison of sample
in one path with a standard
in the other path.
a
In single beam radiant
energy wavelength has to
be adjusted every time.
In this scanning can be
done over a wide
wavelength region
a
D consumin,
Working on single beam is
tedious and time
Working on double beam is
fast and non tedious.
In single beam it's not
possible to compare blank
and sample together.
In double beam it’s
possible to do direct one
step comparison of sample
in one path with a standard
in the other path.
a
In single beam radiant
energy wavelength has to
be adjusted every time.
In this scanning can be
done over a wide
wavelength region
a
D consumin,
Working on single beam is
tedious and time
Working on double beam is
fast and non tedious.
REFERENCE
» Instrumental Analysis, Skoog, Fifth edition, Page no. 312-316
» Instrumental methods of chemical analysis, Gurdeep R
chatwal. Page n02.116-2.122
» Elementary organic analysis, Principles and chen
applications , Y R Shrama, page no12-14
» Atextbook of pharmaceutical analysis, kasturi A V, Vol 3 10%
ed., 169-81
en
» Instrumental Analysis, Skoog, Fifth edition, Page no. 312-316
» Instrumental methods of chemical analysis, Gurdeep R
chatwal. Page n02.116-2.122
» Elementary organic analysis, Principles and chen
applications , Y R Shrama, page no12-14
» Atextbook of pharmaceutical analysis, kasturi A V, Vol 3 10%
ed., 169-81
en
INSTRUMENTATION
Components of spectrophotometer
“4 Source
= Monochromator
= Sample compartment
Detector
= Recorder
Ml...
Components of spectrophotometer
“4 Source
= Monochromator
= Sample compartment
Detector
= Recorder
Ml...
INSTRUMENTATION
Fig.-block diagram of instrumentation of UV-spectrophotometer
Ml...
Fig.-block diagram of instrumentation of UV-spectrophotometer
Ml...
amplifier
Monochromator Detector
Exit slit
Dispersion
Source device
Entrance
slit
ck diagrammatic representation of UV-spectrophotometer
Monochromator Detector
Exit slit
Dispersion
Source device
Entrance
slit
ck diagrammatic representation of UV-spectrophotometer
RADIATION SOURCE
It is important that the power of the radiation source does
not change abruptly over its wavelength range. The
electrical excitation of deuterium or hydrogen at low
pressure produces a continuous UV spectrum.
Both Deuterium and Hydrogen lamps emit radiation in
the range 160 - 375 nm.
Problem-
- Due to evaporation of tungsten life period decreases.
- Itis overcome by using tungsten-halogen lamp.
» Halogen gas prevents evaporation of tungsten.
Ml...
It is important that the power of the radiation source does
not change abruptly over its wavelength range. The
electrical excitation of deuterium or hydrogen at low
pressure produces a continuous UV spectrum.
Both Deuterium and Hydrogen lamps emit radiation in
the range 160 - 375 nm.
Problem-
- Due to evaporation of tungsten life period decreases.
- Itis overcome by using tungsten-halogen lamp.
» Halogen gas prevents evaporation of tungsten.
Ml...
RADIATION SOURCE
For ultra violet region-
Hydrogen discharge lamp
» consist of two electrode contain in deuterium filled silica
envelop.
UV-Vis spectrophotometer have both deuterium & tungsten
lamps.
4 Selection of lamp is made by moving lamp mounting or
mirror to cause the light fall on Monochromator.
Deuterium lamps:-
>» Radiation emitted is 3-5 times more than the hydrogen
discharge lamps.
Xenon discharge lamp:-
r Xenon stored under pressure in 10-30 atmosphere.
For ultra violet region-
Hydrogen discharge lamp
» consist of two electrode contain in deuterium filled silica
envelop.
UV-Vis spectrophotometer have both deuterium & tungsten
lamps.
4 Selection of lamp is made by moving lamp mounting or
mirror to cause the light fall on Monochromator.
Deuterium lamps:-
>» Radiation emitted is 3-5 times more than the hydrogen
discharge lamps.
Xenon discharge lamp:-
r Xenon stored under pressure in 10-30 atmosphere.
FILTERS OR MONOCHROMATORS
All Monochromators contain the following component parts;
+ An entrance slit
- A collimating lens
+ A dispersing device (a prism or a grating)
+ A focusing lens
+ An exit slit
Reflection
ance grating Exit
slit
All Monochromators contain the following component parts;
+ An entrance slit
- A collimating lens
+ A dispersing device (a prism or a grating)
+ A focusing lens
+ An exit slit
Reflection
ance grating Exit
slit
O Filters —
a)Glass filters- Made from pieces of colored glass which
transmit limited wave length range of spectrum. Wide band
width 150nm.
b)Gelatin filters- Consist of mixture of dyes placed in gelatin
& sandwiched between glass plates. Band width 25nm.
c)Inter ferometric filters- Band width 15nm
OPrisms-
-Prism bends the monochromatic light.
-Amount of deviation depends on wavelength
-They produce non linear dispersion.
Ml...
a)Glass filters- Made from pieces of colored glass which
transmit limited wave length range of spectrum. Wide band
width 150nm.
b)Gelatin filters- Consist of mixture of dyes placed in gelatin
& sandwiched between glass plates. Band width 25nm.
c)Inter ferometric filters- Band width 15nm
OPrisms-
-Prism bends the monochromatic light.
-Amount of deviation depends on wavelength
-They produce non linear dispersion.
Ml...
Fig.-mechanism of working of prism.
SAMPLE CONTAINERS OR SAMPLE CELLS
A variety of sample cells available for UV region. The
choice of sample cell is based on
a) the path length, shape, size
b) the transmission characteristics at the desired
wavelength
c) the relative expense
- The cell holding the sample should be transparent to the
wavelength region to be recorded. Quartz or fused silica
cuvettes are required for spectroscopy in the UV region.
Silicate glasses can be used for the manufacture of
cuvettes for use between 350 and 2000nm. The
thickness of the cell is generally 1 cm. cells may be
| in shape or cylindrical with flat ends.
A variety of sample cells available for UV region. The
choice of sample cell is based on
a) the path length, shape, size
b) the transmission characteristics at the desired
wavelength
c) the relative expense
- The cell holding the sample should be transparent to the
wavelength region to be recorded. Quartz or fused silica
cuvettes are required for spectroscopy in the UV region.
Silicate glasses can be used for the manufacture of
cuvettes for use between 350 and 2000nm. The
thickness of the cell is generally 1 cm. cells may be
| in shape or cylindrical with flat ends.
DETECTORS
Three common types of detectors are used
|. Barrier layer cell
ll. Photo cell detector
IN. Photomultiplier , Photo voltaic cells
barrier layer cells
It consist of flat Cu or Fe electrode on which semiconductor such
as selenium is deposited. on the selenium a thin layer of silver or
gold is sputtered over the surface.
ae
= Semitransparent
Silver
Semiconductor
= — Metal Base
Three common types of detectors are used
|. Barrier layer cell
ll. Photo cell detector
IN. Photomultiplier , Photo voltaic cells
barrier layer cells
It consist of flat Cu or Fe electrode on which semiconductor such
as selenium is deposited. on the selenium a thin layer of silver or
gold is sputtered over the surface.
ae
= Semitransparent
Silver
Semiconductor
= — Metal Base
CONTINUED
Photomultiplier tube
It is generally used as detector in UV- spectrophotometer It is the
combination of photodiode & electron multipli
It consist of evacuated tube contains photo- cathode. 9-16 electrodes
known as dynodes.
Photosensitive Cathode
Photomultiplier tube
It is generally used as detector in UV- spectrophotometer It is the
combination of photodiode & electron multipli
It consist of evacuated tube contains photo- cathode. 9-16 electrodes
known as dynodes.
Photosensitive Cathode
Recorder
The construction of a traditional UV-VIS spectrometer is very similar to an
IR, as similar functions — sample handling, irradiation, detection and
output are required
Here is a simple schematic that covers most modern UV spectrometers:
log(Ip/!) =A
UV-VIS sources
e 7
monochromator/
beam splitter optics
IR, as similar functions — sample handling, irradiation, detection and
output are required
Here is a simple schematic that covers most modern UV spectrometers:
log(Ip/!) =A
UV-VIS sources
e 7
monochromator/
beam splitter optics
UV Spectroscopy
Two sources are required to scan the entire UV-VIS band:
. Deuterium lamp — covers the UV — 200-330
. Tungsten lamp — covers 330-700
As with the dispersive IR, the lamps illuminate the entire band of UV
or visible light; the monochromator (grating or prism) gradually
changes the small bands of radiation sent to the beam splitter
The beam splitter sends a separate band to a cell containing the
sample solution and a reference solution
The detector measures the difference between the transmitted light
through the sample (I) vs. the incident light (I) and sends this
information to the recorder
Two sources are required to scan the entire UV-VIS band:
. Deuterium lamp — covers the UV — 200-330
. Tungsten lamp — covers 330-700
As with the dispersive IR, the lamps illuminate the entire band of UV
or visible light; the monochromator (grating or prism) gradually
changes the small bands of radiation sent to the beam splitter
The beam splitter sends a separate band to a cell containing the
sample solution and a reference solution
The detector measures the difference between the transmitted light
through the sample (I) vs. the incident light (I) and sends this
information to the recorder
UV Spectroscopy
Il. “Instrumentation and Spectra
A. Instrumentation
7. As with dispersive IR, time is required to cover the entire UV-VIS band
due to the mechanism of changing wavelengths
8. A recent improvement is the diode-array spectrophotometer - here a
prism (dispersion device) breaks apart the full spectrum transmitted
through the sample
Each individual band of UV is detected by a individual diodes on a silicon
wafer simultaneously — the obvious limitation is the size of the diode, so
some loss of resolution over traditional instruments is observed
UV-VIS sources |
Polychromator
— entrance slit and dispersion device
Il. “Instrumentation and Spectra
A. Instrumentation
7. As with dispersive IR, time is required to cover the entire UV-VIS band
due to the mechanism of changing wavelengths
8. A recent improvement is the diode-array spectrophotometer - here a
prism (dispersion device) breaks apart the full spectrum transmitted
through the sample
Each individual band of UV is detected by a individual diodes on a silicon
wafer simultaneously — the obvious limitation is the size of the diode, so
some loss of resolution over traditional instruments is observed
UV-VIS sources |
Polychromator
— entrance slit and dispersion device
UV Spectroscopy
Il. “Instrumentation and Spectra
B. Instrumentation — Sample Handling
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
4. Concentration (we will cover shortly) is empirically determined
A typical sample cell (commonly called a ):
Il. “Instrumentation and Spectra
B. Instrumentation — Sample Handling
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
4. Concentration (we will cover shortly) is empirically determined
A typical sample cell (commonly called a ):
DESCRIPTION OF UV- SPECTROPHOTOMETER
Advantage of double beam spectrophotometer:- It is not necessary to
continually replace the blank with the sample or to adjust the auto zero.
The ratio of the powers of the sample & reference is constantly obtained.
It has rapid scanning over the wide wavelength region because of the
above two factors.
did VA
UV-VIS sources
SS 7
monochromator/
beam splitter optics
Advantage of double beam spectrophotometer:- It is not necessary to
continually replace the blank with the sample or to adjust the auto zero.
The ratio of the powers of the sample & reference is constantly obtained.
It has rapid scanning over the wide wavelength region because of the
above two factors.
did VA
UV-VIS sources
SS 7
monochromator/
beam splitter optics
Single beam spectrophotometer
ene Entrance slit
Col ating lens
Lamp
À Grating
Faire Wavelength
+ control cam
knob
Exit slit
Meter Cuvette
Phototube
a
ene Entrance slit
Col ating lens
Lamp
À Grating
Faire Wavelength
+ control cam
knob
Exit slit
Meter Cuvette
Phototube
a
Double beam colorimeter
Display device
Display device
APPLICATIONS OF
UV / VISIBLE
SPECTROSCOPY
UV / VISIBLE
SPECTROSCOPY
Applications
+ Qualitative & Quantitative Analysis:
— It is used for characterizing aromatic compounds
and conjugated olefins.
— It can be used to find out molar concentration of the
solute under study.
+ Detection of impurities:
— It is one of the important method to detect
impurities in organic solvents.
+ Detection of isomers are possible.
+ Determination of molecular weight using Beer’s
law.
+ Qualitative & Quantitative Analysis:
— It is used for characterizing aromatic compounds
and conjugated olefins.
— It can be used to find out molar concentration of the
solute under study.
+ Detection of impurities:
— It is one of the important method to detect
impurities in organic solvents.
+ Detection of isomers are possible.
+ Determination of molecular weight using Beer’s
law.
APPLICATIONS:
A. APPLICATIONS IN ORGANIC COMPOUNDS
1.1 is helps to show the relationship between different groups, it is useful to
detect the conjugation of the compounds.
2.Detection of geometrical isomers, In case of geometrical isomers compounds,
that trans isomers exhibits Amax at slightly longer wavelength and have larger
extinction coefficient then the cis isomers
3.Detection of functional groups, it is possible to detect the presence of certain
functional groups with the help of UV Spectrum.
GENERAL APPLICATIONS:
1.Quaitative analysis, UV absorption spectroscopy can characterizes those type of
compounds which absorb UV radiation. Identification is done by comparing the
absorption spectrum withthe spectra of known compound
2. Itis useful in Quantitative analysis of the compounds.
3. Detection of impurities, UV absorption spectroscopy is the one of the best
for detecting impurities in organic compounds.
A. APPLICATIONS IN ORGANIC COMPOUNDS
1.1 is helps to show the relationship between different groups, it is useful to
detect the conjugation of the compounds.
2.Detection of geometrical isomers, In case of geometrical isomers compounds,
that trans isomers exhibits Amax at slightly longer wavelength and have larger
extinction coefficient then the cis isomers
3.Detection of functional groups, it is possible to detect the presence of certain
functional groups with the help of UV Spectrum.
GENERAL APPLICATIONS:
1.Quaitative analysis, UV absorption spectroscopy can characterizes those type of
compounds which absorb UV radiation. Identification is done by comparing the
absorption spectrum withthe spectra of known compound
2. Itis useful in Quantitative analysis of the compounds.
3. Detection of impurities, UV absorption spectroscopy is the one of the best
for detecting impurities in organic compounds.
Tautomeric equilibrium, UV spectroscopy can be used to determine the
percentage of various keto and enol forms present in tautomeric equilibrium.
5. Chemical kinetics, UV spectroscopy can be used to study the kinetics of
reactions.
6. Molecular weight determination, molecular weights of compounds can be
measured by spectroscopy.
7. Analysis of inorganic compounds.
8. Measuring concentration of solution, absorption band can also used to
determine the concentration of compounds in a solution.
9. Inorganic chemistry, absorption spectra have been used in connection with
many problems in inorganic chemisty.
10. Itis useful to determine the structure of the chloral
percentage of various keto and enol forms present in tautomeric equilibrium.
5. Chemical kinetics, UV spectroscopy can be used to study the kinetics of
reactions.
6. Molecular weight determination, molecular weights of compounds can be
measured by spectroscopy.
7. Analysis of inorganic compounds.
8. Measuring concentration of solution, absorption band can also used to
determine the concentration of compounds in a solution.
9. Inorganic chemistry, absorption spectra have been used in connection with
many problems in inorganic chemisty.
10. Itis useful to determine the structure of the chloral
REFERENCES
Reference Books
« Introduction to Spectroscopy
— Donald A. Pavia
+ Elementary Organic Spectroscopy
— Y. R. Sharma
+ Physical Chemistry
— Puri, Sharma & Pathaniya
« Introduction to Spectroscopy
— Donald A. Pavia
+ Elementary Organic Spectroscopy
— Y. R. Sharma
+ Physical Chemistry
— Puri, Sharma & Pathaniya
Resources
+ http://www2.chemistry.msu.edu/faculty/reu
sch/VirtTxtJmI/Spectrpy/UV-
Vis/spectrum.htm
+ http://en.wikipedia.org/wiki/Ultraviolet%E2
%80%93visible spectroscopy
+ http://teaching.shu.ac.uk/hwb/chemistry/tut
orials/molspec/uvvisab1.htm
+ http://www2.chemistry.msu.edu/faculty/reu
sch/VirtTxtJmI/Spectrpy/UV-
Vis/spectrum.htm
+ http://en.wikipedia.org/wiki/Ultraviolet%E2
%80%93visible spectroscopy
+ http://teaching.shu.ac.uk/hwb/chemistry/tut
orials/molspec/uvvisab1.htm
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