UV SPECTROSCOPY
Introduction
UV Spectroscopy is very widely used technique in labs and in
life science life department. Many different experiments to relay on the idea
of UV visible Spectroscopy but the first question is why we need this technique
than what this technique gives us the answer to that is in any situation we
need to find concentration of different substance of different molecule in
solution and that we can measure perfectly with UV spectrophotometry. In some
case also concentration of cell during the microbiology it also tells us the idea
of the two things the wavelength and frequency of that the electromagnetic wave
and the frequency of a wave.
More frequency means more energy and if something has high
frequency it will have a lesser or smaller wavelength. So, a very longer wavelength
wave will have less frequency and less energy. If we divided this UV visible
spectrum into the small part and fraction, we would find out that it is simply
ranging from 200 nm to 800 nm that's kind of the range of the UV-visible light.
Although it can be extended to 10-20 nanometer in the wavelength.
From 200 to 400nm it is UV range and from 400 to 800nm it belongs to the
visible range that we usually use for the UV visible Spectroscopy.
When we move towards x-ray, gamma ray the wavelength become
more smaller and frequency gets higher, and the energy will also be greater.
The visible light further can be sub divided into seven different colors. This
can be seen when a light ray passes through a water droplet or a prism. That’s
why we see a beautiful rainbow after rain. The colors of rainbow or the white
light has the following sub colors in ordered form:
1-
Red
2-
Orange
3-
Yellow
4-
Green
5-
Blue
6-
Indigo
7-
violet
If we look at the instrument the filament act as a light
source. It also has the separate unit known as the monochromator.
UV Spectroscopy is an analytical technique that measure the number
of different wavelengths of UV or visible light that are absorbed by or
transmitted through a sample in comparison to a or reference or blank sample.
Every sample has its own lambda max value which we can also find through a
graph. The UV can provide information on what is in the sample and the
concentration.
Since this Spectroscopy technique works using the light,
let's first consider the properties of light. UV Spectroscopy refers to
absorption to absorption Spectroscopy or reflectance spectroscopy.
What is Spectroscopy and how it is use?
Spectroscopy is the branch of science in which we study
about the interaction of light with the matter. UV Spectroscopy is the one of
the techniques used to measure how much a chemical reaction absorbs light. This
is done by measuring the intensity of light. It is referred to transmit
Spectroscopy in part of the ultraviolet and the full near to the visible region
of the electromagnetic spectrum. In UV Spectroscopy a sun light travel through
a solution in a cuvette with wavelength ranging between 180 and 1100nm.
The sample transmit this UV or visible radiation in the
cuvette. Cuvette can be made up of quartz or any other substance. It should be
clean while using it, make sure to wash the cuvette with distilled water so
that you can observe a clear result. Make sure to test run water at the start
before putting a sample in the cuvette. The distilled water line which you get
on the computer will act as a standard for you to compare the results or we can
say is it will act as a baseline.
Invention of UV Spectroscopy
In July 1941, it is invented by Arnold Beckman, founder of
his eponymous company, introduced his DU UV - vis spectrometer.
Range of UV Spectroscopy
The UV range extends from 100-400nm and the visible spectrum
ranges from 400-700nm. However, most spectrophotometers don not operate in the
deep UV range of 100-200nm, as light sources in this range are not economical
and difficult to operate.
Uses of UV Spectroscopy
UV-Vis Spectroscopy is a quantitative technique used to
measure how much a chemical substance absorbs light. This is done by measuring
the intensity of light that passes through a sample with respect to the
intensity of light through a reference sample or blank. It is commonly used
technique in many areas of science ranging from bacterial culturing, drug
identification and nucleic acid purity unmixed and quantitation, to quality
control in the beverage industry and chemical research. Pharmaceutical
industries are the most common industries where UV spectrophotometer is being
used.
It is a quantitative technique used to measure how much a
chemical substance absorbs light. This is done by measuring the intensity of light
that passes through a sample with respect to the intensity of light through blank
reference sample. UV spectroscopy is also used in semiconductor industry to
measure the thickness and optical properties of thin film on a wafer.
UV spectrometers are used to measure the reflectance of
light and can be action. UV spectroscopy like fitter is a technique which is
useful in finding out the purity of the drug. Many molecules contain
chromophores which will absorb specific wavelength of ultraviolet. According to
the Beer Lambert Law the absorption of spectra produces from these sample of
given wavelength can be related directly to the concentrated to these sample.
The equation of lamber-beer law is as follow:
A = Absorbance
ℇ = Molar absorptivity
B = path length
C = concentration
Principle of UV-Vis Spectroscopy
The principle of UV-Vis Spectroscopy is based on the
absorption of ultraviolet light or visible light by chemical compounds, which
result in the production of different spectra. Spectroscopy is based on the
interaction between light and matter. As a result of absorption of
electromagnetic radiation molecular change occur. These changes occur between
the electronic energy level. As molecule absorb energy, an energy is to change
of position from occupied orbital to an unoccupied orbital of greater potential
energy
Instrumentation in UV Spectroscopy
I. Light source
II. Monochromator
III. Detector
IV. Amplifiers
V. Recorder
.jpg)
Light source in UV Spectroscopy
The basic important things are that light source which
passes polychromatic radiation to monochromator. we know light source as it
will have visible and UV range light. we will have different light bulbs as a
source of light depending upon the wavelength we need, and it is like a head
light of our motor bike. Mostly in experiments two light sources are used, for
ultraviolet light Deuterium is used and for visible light tungsten lamp is
used.
There are many things which we should keep in mind while
using the source of light firstly, the output light should be uniform over a
specific wavelength secondly, it should be stable for a long period of time
thirdly, the usage should be long and lastly the light source should be
economical. Some of the light sources are listed below.
i.
Halogen Lamps
ii.
Deuterium Lamps
iii. Xenon Lamp (Xenon Arc Lamp)
iv. Xenon Flash Lamp
v.
Low-Pressure Mercury Lamp
Monochromator
Monochromators are used to break up large bands of
polychromatic light. The light source is separated by a monochromator to obtain
monochromatic light of certain wavelength, which is irradiated to the sample
solution in the sample chamber. It has three types.
i.
Filters
ii.
Prisms
iii.
Gratings
Prism monochromator is made up of quartz, fused silica,
both, and glass. The spectrophotometer has two types, single monochromator type
which has only one monochromator and second is double type monochromator which
has two monochromators, and you easily guess it by their name.
Sample holder
It is device which is used as a sample holder and the sample
is kept here. UV spectrophotometer’s holder is made up of glass, Quartz, iron,
and bentonite. It is generally called as cuvette holder which is a customized
light measuring component intended to keep liquid transmission measurements as
accurate as possible. It keeps the sample in a stable position throughput the
process.
Modern cuvette holders can additionally include a
temperature-controlled setting for testing how well a cuvette transmits light
under various heat conditions and in reaction to fiber optic. These light
measurement accessories are strong and adaptable, and they have a staggering
array of uses in the study of light measurement and the characterization of
materials.
Detectors
Detector will detect the light signal which then will be
converted to the signal displayed on the device. The UV-Vis spectroscopy
apparatus employs a variety of detectors. The analysis needs to be
straightforward and inexpensive. Depending on the requirements, every detector
may be used to various scenarios and samples.
The choice of
detectors is influenced by the volume of the sample that is present, the
analyte concentration, and the reaction mixture. In the case of
ultraviolet-visible spectroscopy, detection should be carried out at a very
smaller sampling concentration. They ought to be smaller and of greater quality
as well.
The ability of the normal eye to perceive fluctuations in
color makes it a useful tool for colorimetric equipment that match colors.
Although the human visual system achieved the intended function, the
examination was subject to personal judgment and perception, which made alternative
sensitive detection choices necessary.
With the passage of time, electrical charge transfer
properties-based detectors were created. There are basically four types of
detectors in any spectrophotometer.
Following is the type of detectors
1)barrier layer cell
2)Photo tube
3) photomultiplier tube
4) Thermocouple
Amplifier and recorder
Amplifier: Amplifiers amplifies signal coming from detector
Recorder: Recorder
records them which is displayed on the information shown on a computer.
.jpg)
Application of UV Spectroscopy
UV Spectroscopy is one of the best methods for to find out
impurities in organic molecule. Addition of the most extreme possible amount
can be observed due to the possible amount in the sample and can be compared
with that of raw material. By also measuring the absorbance of specific wavelength
the impurities can be detect.
Some power of UV Spectroscopy makes it so great for many
application
o
This technique is non causing a very large amount
of damage which can allow the sample to be reused for further processing or
analysis.
o
Measurement can be performed very quickly.
o
Less user training is need as instrument is
easier to use and handle.
o
Only minimal processing is need for data
analysis.
o The instrument is not high price as it is easy
to need and to be able to use in laboratory.
o
The application of UV Spectroscopy is enormous. The
following are the main field in which UV visible Spectroscopy is used
o
DNA or RNA analysis
o
Pharmaceutical analysis
o
Bacterial culture
o
Beverage analysis
o
Environmental monitoring and control
o
Biotech
o
Life science
o
Gas and oil
o
Geology
o
Metallurgy
Disadvantage of UV Spectroscopy
The main disadvantage of using UV spectrometer is the time.
It takes time to prepare to use one with UV visible spectrophotometer, set up
is key. It must be clear the area of any outside light electronic noise or
other outside. The operator of the spectrophotometer should be well trained, in
addition to that the system or the computer and the recorder should be
connected properly to get accurate results. If cuvette is not properly clean it
may affect the results. Spending a lot of time under UV light may harm your skin
as well as your eyes.
The room where system operate should be a dark space and it
should not have any kind of vibration of any electronic or anything. Furthermore,
it can only analyze the molecules which contains chromophores. External and
internal factors such as pH, temperature, contaminants, and impurities can also
affect the results. We cannot analyze solid or a gas sample, and the sample
needs to in liquid form.
Advantage of UV Spectroscopy
The biggest advantage for chemist and astronomers who use UV
visible spectrometer is the accuracy of the device. Even small UV visible
spectrometer can give extremely occur ate reading which is extremely important
when we are preparing chemical solution. In addition to that it is easy to use,
to handle and is easy to operate. We can easily buy it from the market at a
very low price and even small laboratories can afford it.
It can easily cover the entire region of the UV-vis light
and can be used for the quantitative and the qualitative relationship. It can
also provide a derivative graph which is important in any research. It also can
be used for the degradation of the study of the degradation of the dyes and
drugs.
Conclusion
The spectroscopy in the visible ultraviolet region. In a
very useful technique for qualitative studies related to characterization of
organic and inorganic compounds, in food material. In this context the
scientific community has used this technique more research area of food science
and technology sector, it’s application in quality laboratories of food
industries has been very important, because it satisfies both the scope and
public health problem. It allows quality verification of different product
widely compare and little around the world.
No comments:
Post a Comment