Full research about Spectrophotometer

2. A spectrophotometer is an instrument that measures the
amount of light absorbed by a sample.
Spectrophotometer techniques are used to measure the
concentration of solutes in solution by measuring the
amount of the light that is absorbed by the solution in a
cuvette placed in the spectrophotometer .

3. The spectrophotometer technique is to measures

light intensity as a function of wavelength. It
does this by diffracting the light beam into a
spectrum of wavelengths, detecting the intensities
with a charge-coupled device, and displaying the
results as a graph on the detector and then the
display device .

4. 1)Measure the concentration of the solution
A spectrophotometer optically determines the
absorbance or transmission of characteristic
wavelengths of radiant energy (light) by a chemical
species in solution. Each molecule absorbs light at
certain wavelengths in a unique spectral pattern
because of the number and arrangement of its
characteristic functional groups, such as double
bonds between carbon atoms.
According to the Beer-Lambert law, the amount of light
absorbed at these wavelengths is directly
proportional to the concentration of the chemical
species.

5. 2) Identify organic compounds by determining
the absorption maximum.
Spectrophotometers are used to identify organic
compounds by determining the absorption
maxima (which for most compounds and
groups of compounds have very distinct
fingerprints (that's what the absorption curves
and peaks are called).
3) Used for color determination within the
spectral range
If one is working in the range of 380 to 700 nm,
the spectrophotometers can also be used for color
determination within this spectral range

6. Example
-In the Figure below the red part of the spectrum has been
almost completely absorbed by CuSO4 and blue light
has been transmitted. Thus, CuSO4 absorbs little blue
light and therefore appears blue.
-We will get better sensitivity by directing red light
through the solution because CuSO4 absorbs strongest
at the red end of the visible spectrum. But to do this, we
have to isolate the red wavelengths

8. 1)Light source
The function of the light source is to provide a
sufficient of light which is suitable for marking a
measurement. The light source typically yields a
high output of polychromatic light over a wide
range of the spectrum.

9. I) Tungsten Lamp
Tungsten Halogen Lamp, it is the most common light source
used in spectrophotometer. This lamp consists of a tungsten
filament enclosed in a glass envelope, with a wavelength
range of about 330 to 900 nm, are used for the visible region.
They are generally useful for measuring moderately dilute
solutions in which the change in color intensity varies
significantly with changes in concentration . It has long life
about 1200h.

10. II) Hydrogen / Deuterium Lamps
For the ultraviolet region, hydrogen or deuterium
lamps are frequently used.
their range is approximately 200 to 450 nm.
Deuterium lamps are generally more stable and has
long life about 500h.This lamp generates continuous
or discontinuous spectral.

11. III) Xenon flash lamps
Xenon flash lamps have several advantages as the
following :
1)Their range between ( 190nm - 1000 nm)
2) Emit both UV and visible wavelengths
3) Long life
4) Do not heat up the instrument
5) Reduce warm up time

12. 2) Dispersion devices
*Monochromator
Accepts polychromatic input light from a lamp
and outputs monochromatic light.
Monochromator consists of three parts:
I) Entrance slit
II) Exit slit
III) Dispersion device

13. Monochromator

14. Dispersion devices :
Dispersion devices causes a different wavelength of light
to be dispersion at different angles monochromators used
for function.
*Types of dispersion devices :
1)Prism
Prism is used to isolate different wavelength .If a parallel beam
of radiation falls on a prism , the radiation of two different
wavelength will be bent through different angles.
Prism may be made of glass or quartz. The glass prisms are
suitable for radiation essentially in the visible range whereas
the quartz prism can cover the ultraviolet spectrum also.
It is found that the dispersion given by
glass is about three
times that of quartz.

15. 2)Filter
Filters separate different parts of the electromagnetic
spectrum by absorbing or reflecting certain wavelengths
and transmitting other wavelengths.
*Absorption filters are glass substrates containing absorbing
species that absorb certain wavelength. A typical example is a cut
on color filter, which blocks short wavelength light such as an
excitation source, and transmits longer wavelength light such as
fluorescence that reaches a detector.
*Interference filters are made of multiple dielectric thin films
on a substrate. They use interference to selectively transmit or
reflect a certain range of wavelengths.
A typical example is a Bandpass interference filter that
transmits a narrow range of wavelengths, and can isolate
a single emission line from a discharge lamp.

17. 3) Diffraction gratings
Diffraction grating is an optical component with
a regular pattern, which splits (diffracts) light
into several beams travelling in different
directions. The directions of these beams
depend on the spacing of the grating and the
wavelength of the light so that the grating acts
as a dispersive element.
The diffraction grating disperses the light into a
linear spectrum of its component
wavelengths, which is then directed, in whole
or in part along the light path of the
instrument.

19. 3)Focusing devices
Combinations of lenses, slits, and mirrors. Variable
slits also permit adjustments in the total radiant energy
reaching the detector. The Ebert and Czerny-Turner
monochromators and their variations are combinations of
prisms or gratings and focusing devices .
Ebert and Czerny-Turner
Monochromator.

20. *Optical Materials
1)Mirrors
Type of rays Mirror material
X-rays – Ultraviolet(UV) Aluminum

Visible Aluminum

Near infrared Gold

Infrared (IR) Copper or Gold


21. 2)Lenses
Rays Material
X-rays Ultraviolet Fused silica , Sapphire

Visible Glass

Infrared Glass


22. 4)Absorption cells(Cuvettes)
A cuvette is a kind of cell (usually a small square
tube) sealed at one end, made of Plastic, glass or
optical grade quartz and designed to hold samples
for spectroscopic experiments. Cuvette should be
as clear as possible, without impurities that might
affect a spectroscopic reading. Like a test-tube, a
cuvette may be open to the atmosphere on top or
have a glass or Teflon cap to seal it shut.

23. Cuvettes are chosen for transparency in the spectral
wavelengths of interest.
For measurements in the visible region, cuvettes of optical
glass are sufficient; however, optical glass absorbs light
below 350 nm , and more expensive quartz or fused silica
must be used for these wavelengths. The sample cuvettes
are placed in a darkened analysis chamber; some chambers
have rotating carousels that can hold several cuvettes.

24. 5)Detectors
Any photosensitive device can be used as a detector
of radiant energy .The photocell and phototube are
the simplest photodetectors, producing current
proportional to the intensity of the light striking
Them .

25. *Types of detectors
1) Silicon PIN Photodiodes Photovoltaic V-Series
Blue enhanced for spectral range from 350nm to
1100nm; designed for low-noise, D.C. to
medium bandwidth applications. Active areas
range from .31mm² to 100mm². Applications
include: low light level measurements, particle
counting, chemical and analytical measurement
and detection.

26. 2)Gallium Nitride (GaN) UV Detectors
This family of Gallium Nitride (GaN) UV Detectors
are Schottky processed fully passivated U.V.
photodiodes. Spectral range from 200 nm to 365
nm and is ideal for UVA or UVB sensing
applications and is packaged with a quartz
window.

27. 6)Display devices
The data from a detector are displayed by a readout
device, such as an analog meter, a light beam
reflected on a scale, or a digital display , Or liquid
crystal display(LCD) .The output can also be
transmitted to a computer or printer.

28. First we but the sample into a Cuvette then the light source
generates light at a specific wave length or wave lengths , the
light passes through the dispersion devices that separate the
light into its components wavelengths .
Slits then isolate the wave lengths needed for measurement with
a * Bandpass filter to improve its purity . Next , the light
passes through the sample ,and a portion of radiant energy
absorbed . The remaining light is transmitted to the
Photometer ,which converts light energy to electrical energy
can be registered on a meter or digital readout.
The amount of light absorbed depends on the nature of the
concentration of the sample .
*Bandpass filter is a device that passes frequencies within a
certain range and rejects frequencies outside that range.

30. Used Laws :

33. There are two classes of spectrophotometers:
1)Single beam
The single beam spectrophotometer was the first
invented, and all the light passes through the
sample. In this case, to measure the intensity of
the incident light, the sample must be removed
so all the light can pass through. This type is
cheaper because there are less parts and the
system is less complicated.

34. The advantages of the single beam design
are low cost, high throughput, and hence high
Sensitivity , because the optical system is simple.
The disadvantage is that an appreciable amount of
Time elapses between taking the reference (I) and
Making the sample measurement (Io) so that there
can be problems with drift. This was certainly true of
Early designs but modern instruments have better
electronics and more stable lamps, so stability with
single beam instruments is now more than adequate
for the vast majority of application.

36. 2)Double beam
The double beam instrument design aims to eliminate

drift by measuring blank and sample virtually
simultaneously. A quot;chopperquot; alternately transmits and
reflects the light beam so that it travels down the blank
and the sample optical paths to a single detector. The
chopper causes the light beam to switch paths at about 50
Hz causing the detector to see a quot;saw toothquot; signal of Io
and I which are processed in the electronics to give either
transmittance or absorbance as output.
To measure a spectrum with a double beam instrument

the two cuvettes, both containing solvent are place in the
sample and reference positions and a quot;balancequot;
measurement is made. This is the difference between the
two optical paths and is subtracted from all subsequent
measurement. The sample is then placed in the sample
cuvette and the spectrum is measured. I and Io are
measured virtually simultaneously as described above.

37. The advantage of the double beam design

Is high stability because reference and sample are
measured virtually at the same moment in time.
The disadvantages are higher cost, lower
sensitivity because throughput of light is poorer
because of the more complex optics and lower
reliability because of the greater complexity.

39. 3)Split Beam
 The split beam spectrophotometer is similar to the
double beam spectrophotometer but it uses a beam
splitter instead of a chopper to send light along the
blank and sample paths simultaneously to two
separate but identical detectors. Thus blank and
sample measurements can be made at the same
moment in time. Spectra are measured in the same
way as with a double beam spectrophotometer.
 The advantage of this design is good stability, though
not as good as a double beam instrument because two
detectors can drift independently, and good
noise, though not as good as a single beam instrument
because the light is split so that less than 100% passes
through the sample.

40. Understanding the types of instruments

available for measuring color is important when
choosing the instrument to purchase or use for
your application . The quot;colorimeterquot; and
quot;spectrophotometerquot; cause some confusion . both
types of instruments provide data obtained over
the same range of visible wavelengths
(about400-700nm) but may treat this data
differently.
Spectrophotometers and colorimeters are

instruments that measure color intensities of
solutions by applying a light source to the
solution.