1

LAQ:
1) Describe diff components of UV-Vis Spectrophotometer and draw its schematic diagram
2) a) Construction & working principle of detectors used in UV-Vis spectroscopy.
3) a) explain spectrophotometric titration with examples?

1) Describe diff components of UV-Vis Spectrophotometer and draw its

schematic diagram
Introduction
The essential parts of a spectrophotometer are:
1) Radiation Source: Both the tungsten and D₂ lamp are present in the UV-visible
spectrophotometer.
2) Wavelength Selector: It constitutes three essential parts:
i) Filter: Absorption and interference filters are mainly used.
ii) Monochromator: It gives the desired wavelength in the entire UV or visible
region.
iii) Slits: There are two slits, i.e., entrance slit and exit slit.
3) Cells or Cuvettes: For holding the sample solution and the pure solvent (reference).
4) Detector: The most commonly used detectors are photo-emissive cells or phototubes and
photomultiplier tubes.
5) Recording System: Recording is done by recorder pen.
6) Power Supply
Spectrophotometers are of two types:
1) Single-beam spectrophotometers, and
2) Double-beam spectrophotometers.

Sources of Radiation
 2

The best source of light is the one which is more stable, more intense and which gives range of
spectrum from 180-360nm (up to 400nm). The different sources available are:
1) Hydrogen Discharge Lamp: In these lamps, hydrogen gas is stored under relatively high
pressure. When an electric discharge is passed through the lamp, excited hydrogen
molecules will be produced which emit UV radiations. The high pressure in the hydrogen
lamps causes the hydrogen to emit a continuum rather than a simple hydrogen spectrum.
Hydrogen lamps cover the range of 3500-1200Å. These lamps are stable, robust, and
widely used. If deuterium (D2) is used instead of hydrogen, the emission intensity is
increased by as much as a factor of 3 at the short-wavelength end of the UV range.
2) Deuterium Lamp: It is similar to hydrogen discharge lamp, but filled with deuterium in
the place of hydrogen. It offers 3-5 times more intensity than other types. Deuterium
lamps are more expensive than hydrogen lamps but are used when higher intensity is
required.
3) Xenon Discharge Lamp: In this lamp, xenon at 10-30 atmospheric pressure is filled in
and it has two tungsten electrodes. The intensity is greater than the hydrogen discharge
lamp.
4) Mercury Arc: This contains mercury vapour and offers bands which are sharp. The
spectrum is not continuous. Hence, it is not widely used.
5) Tungsten Lamp: This lamp is similar in its functioning to an electric light bulb. It is a
tungsten filament heated electrically to white heat. It has two shortcomings. The intensity
of radiation at short wavelengths (<350nm) is small. Furthermore, to maintain a constant
intensity, the electrical current to the lamp must be carefully controlled.

Wavelength Selectors
Wavelength selectors consist of three parts:
1) Filters,
2) Monochromators, and
3) Slits.
Filters
Filters provide high radiation throughout, approximately 50-80% efficiency. The two types of
filters are:
1) Absorption Filters: These filters derive their effects from bulk interactions of radiation
within the material. Absorption filters are produced in a variety of host materials, like
gelatin, glass, liquid, and plastic. Glass filters are used in automated chemical analysis
equipment and colorimetry. The scattering type depends on Scattering crystals formed
within the glass mass through a reduction and thermal treatment. Shorter wavelengths are
scattered and absorbed, while longer wavelengths are unaffected. Cut-on and cut-off (or
sharp-cut) filters are widely used as blocking filters to suppress unwanted spectral orders
from interference filters and diffraction gratings.
 3

There are a wide variety of plastic filters, of both sharp cut and intermediate bandwidth
types. Plastic filters may be produced either by bulk colourants Introduced into the basic
batch or through subsequent dye treatments of clear base stock. Cut-on types, unlike their
glass counterparts, exhibit no fluorescence in the visible region.
2) Interference Filters: These filters, as the name implies( are based on the phenomenon of
optical interference.The thickness of dielectric film is controlled to be only one, two, or
three half-waves thick. These are referred to as first, second-, or third-order filters,
respectively.
Monochromators (Prisms and Gratings)
The monochromator is used to disperse the radiation according to the wavelength. The essential
elements of a monochromator are an entrance slit, a dispersing element, and an exit slit. The
entrance slit sharply defines the incoming beam of heterochromatic radiation. The dispersing
element disperses the heterochromatic radiation into its component wavelengths whereas exit slit
allows the nominal wavelength together with a band of wavelengths on either side of it. Position
of the dispersing element is always adjusted by rotating it to vary the nominal wavelength
passing through the exit slit.
The dispersing element may be a prism or grating. The prisms are generally made of glass,
quartz, or fused silica. Glass has the highest resolving power but it is not transparent to radiations
having the wavelength between 2000-3000A because glass absorbs strongly in this region.
Quartz and fused silica prisms are transparent throughout the entire UV range and are widely
used in UV spectrophotometers. Fused silica prisms are little more transparent in the short
wavelength region than quartz prisms, and are used only when very intense radiation is required.
The mirrors in the optical system are front surfaced because glass starts to absorb in the UV
region.
Characteristics of a Monochromator
1) It allows the largest entrance slit width for the band pass required.
2) It has the highest dispersion.
3) The largest optics is affordable.
4) Longest focal length is affordable.
5) Highest groove density will accommodate the spectral range.
6) Optics and coatings are appropriate for specific spectral range.
7) Entrance optics will optimise endues.
8) If the instrument is to be used at a single wavelength in a non-scanning mode, it must be
possible to adjust the exit slit to match the size of the entrance slit image.

Types of Monochromator
1) Prism Monochromator:
2) Grating Monochromator: Gratings provide an alternative means of producing
monochromatic light
 4

Slits
There are two slits, i.e., entrance slit and exit slit. The main function of the entrance slit is to
provide a narrow source of light so that there should be no overlapping of monochromatic
images. From this, the exit slit selects a narrow band of dispersed spectrum for observation
by the detector.
Sample Cells or Cuvettes
Sample containers, which are usually called cells or cuvettes, must have windows that are
ransparent in the spectral region of interest. Quartz or fused silica is required for the UV region
(wavelengths less than 350nm) and may be used in the visible region. Silicate glass is ordinarily
used for 375-2000nm region because of its low cost compared to quartz. Plastic cells are also
used in the visible region.The best cells have windows that are perpendicular to the direction of
the beam in order o minimise reflection losses. The most common cell path length for studies in
the UV und visible regions is of 1cm; matched, calibrated cells of this size are available from
several commercial sources. Many other cells with shorter and longer path lengths can be
purchased.
Detectors
Detectors used in UV-visible spectrophotometers can be called as photometriec detectors. The
most commonly used detectors are:
1) Phototubes or photo-emissive cells,
2) Photomultiplier Tubes (PMT),
3) Photovoltaic cell or barrier-layer cells, and
4) Photomultiplier Silicon photodiode array detector.

2) a) Construction & working principle of detectors used in UV-Vis

spectroscopy.
Detectors are the devices which convert light energy into electrical signals, that are displayed on
the readout devices. After passing through the sample cell, a part of the radiation is absorbed by
the sample and the remaining is transmitted. The transmitted radiation falls on the detector which
determines the intensity of the radiation absorbed by the sample.
(a) Phototubes
 5

Construction
It consists of a hollow glass tube with a photocathode and a collector anode. The surface of the
photocathode is coated with a layer of elements like cesium, potassium, silver oxide or a mixture
of these.
Working
When light falls on the photocathode, electrons are produced that travel towards the collector
anode and generate current. The amount of current generated is directly proportional to the
intensity of light radiation. Compared to barrier layer cell, phototubes are more sensitive and
therefore widely used.

(b) Photomultiplier Tube

Construction
It consist of a light sensitive cathode (photocathode) and 10 anodes (dynodes) maintained at a
potential of 75-100 volts. Photomultiplier tube being sensitive can detect extremely weak signals
also, therefore it is used to intricate instruments. It should be protected from stray light in order
to to obtain accurate results.
 6

Working
It works on the principle of multiplication of the photoelectrons by secondary emission of
electrons. When a beam stage of falls on the photocathode, photoelectrons are generated which
are accelmission of electrons. When a beach due to eleration free one anode to another, the
emission of electrons is end towards the anodes of do due to secondary emission of electrons.
Thus, several electrons are liberated by each original beta facts a result o secondary emission.
Due to this, a net factor of 10 is achieved from the last electrode. is achieved from the last
electrode.
(c )Photovoltaic Cell

Construction
It consists of a photocathode which is a thin metallic layer coated with gold or silver. It also
contains a metal base (usually iron) which acts as anode. Between these two electrodes is a
semiconductor layer of selenium.
Working
When light rays fall on the selenium layer, electrons are generated and taken up by the
photocathode. Because of the poor electrical conductivity of the selenium, the electrons get
accumulated on the cathode leading to the development of potential difference across the two
electrodes which results in the generation of electric current. The current flow causes deflection
in the galvanometer which gives the measure of the intensity of radiation, i.e., greater the
intensity of radiation, greater is the current produced hence greater the deflection in
galvanometer.
Advantage
It is economical, hence commonly used.
Disadvantages
 7

1. For signal amplification, the resistance of the external circuit should be low.
2. It is less responsive towards light except for blue and red.
d) Silicon Photodiode
These are the semiconductor detectors made up of silicon that immediately respond to high
energy particles and protons. These are enabled by absorption of either photons or charged
particles and generate a flow of current in the external circuit which is proportional to the
incident power.
These detectors are used to detect the presence or absence of very small proportions of light in
UV, visible or infrared region and also used in calibration of extremely accurate measurements
from the intensities below 1pW/cm² to the intensities above 100 mW/cm². These are used in
various devices such as analytical instruments, beam alignment, laser range finders, medical
imaging instruments, optical communications, optical position sensors, photography,
spectroscopy and surface characterization.
There exits two different types of semiconductors used in silicon photodiodes.
p-type Semiconductor
A specific incorporation of group III elements in photodiode forms a p-type semiconductor
which is mostly rich in holes.
n-type Semiconductor
A specific incorporation of group V elements in photodiode forms an n-type semiconductor
which is mostly rich in electrons.

3) a) explain spectrophotometric titration with examples?

Spectrophotometric Titrations – Single Component and Multicomponent Analysis
Photometric or spectrophotometric titration signifies the equivalence point provided the
condition that analyte, the equivalence point of the titration is given out by the the reagent or the
titration product absorbs radiant energy, absorbing indicator which shows the change in
absorbance.A graph is plotted by taking absorbance (2) on Y-axis and volume of the titrant on X-
axis. Most of the photomet- nc titrations demonstrate two linear points having different slopes.
One of the slopes occur early in the titration and the other one appears after the equivalence point
region. The linear portions of the curve are extrapolated and inter- gcted to get the end point. At
a fixed absorbance, the titra- bon provides the end points. Moreover the end points are
 8

determined by conversion of the linear-segment curve to a sgmoid-shaped curve with the help of
a derivative.
 9

Figure (i) shows the curve for the titration of a nonabsorbing species with an absorbing titrant,
that reacts with the analyte to form a product that is ‘nonabsorbing’. For instance, the titration of
thiosulfate ion with triiodide ion.
The figure (ii) demonstrates the curve for the titration of iodide ion with a standard iodate ion
solution to form triiodide. The figures (iii), (iv), (v) and (vi) demonstrate different curves that are
obtained with varying combinations of analytes, titrants and products. The titration curves with
linear portions are obtained provided the absorbing species obey Beer’s law. Moreover
corrections are performed on absorbances for volume changes. They are achieved by multiplying
the absorbed observed absorbance by Where, (V + h)/V
V = Original volume of the solution V = Volume of the added titrant.
Beer’s law is not necessarily obeyed when end points are determined by various techniques by
the use of changes in only absorbance in most of the methods.
Instrumentation
The titrations of spectrophotometric titrations are carried out using a spectrophotometer or a
photometer that is modified to the extent that the titration vessel is kept stationary in the high
path. A probe type cell can be used alternatively. Firstly, the instrument is set to a suitable
wavelength, otherwise appropriate filter is selected and inserted. Then the 0% T adjustment is
performed. The radiations are allowed to pass through the analyte solution to the transducer,
accordingly the instrument is adjusted to a convenient absorbance reading. This is achieved by
varying the source intensity or the transcriber sensitivity. The advantage over here is that the
‘end point’ is detected using the ‘relative values of absorbances’ rather than the true absorbance.
The criteria for a photometric titration are the power of the radiation source and the response of
the transducer are kept constant and the path length is kept constant by maintaining the cells at a
fixed position.

Applications
1. Spectrophotometric titrations involve several measurements to determine the end
point, therefore give accurate results.
2. The titrations involve measurements of a change in absorbance hence the other
absorbing species if present do not interfere in the titrations.
3. It is used to titrate more dilute solutions. The end points are determined from
linear-segment titration curves. At the equivalence point regions, the absorbance
changes gradually. The data collected to determine the end points are collected
away from this equivalence point region. This leads to a smaller equilibrium
constant when compared to sigmoid titration curve. (Here the data is collected
near the equivalence point).
 10

4. Standard oxidizing agents have characteristic absorption spectra. Hence show

photometrically detectable end points. Standard acids or bases do not absorb,
therefore, acid-base indicators are introduced that helps in photometric
neutralization titrations.
5. Photometric end point is used in titration with ethylenediamine tetraacetic acid
(EDTA) and complexing agents