UV spectroscopy by group 5

Group members
 Afsana Azeez
 Arjun Battula
 Angel Benny
 Munavar Chembukandy
 Sai Pooja Sri Chittepu

Spectroscopy
• It is the branch of science that deals with the study of interaction of matter
with light or the interaction of electromagnetic radiation with matter.
• UV spectroscopy comes under the spectroscopy analysis which is an
instrumental method of analytical technique.

Absorption spectroscopy
• Absorption spectroscopy refers to spectroscopic techniques that measure the
absorption of radiation, as a function of frequency or wavelength, due to its
interaction with a sample. The sample absorbs energy, i.e., photons, from the
radiating field. Absorption spectroscopy is performed across the
electromagnetic spectrum.
• Absorption spectroscopy is employed as an analytical chemistry tool to
determine the presence of a particular substance in a sample and, in many
cases, to quantify the amount of the substance present. Infrared and
ultraviolet–visible spectroscopy are particularly common in analytical
applications.

Principles &definition
Definition
UV and visible absorption spectroscopy is the measurement of the attenuation of a
beam of light after it passes through a sample or after reflection from the sample
surface. Absorption wavelength can be at a single wavelength or over an extended
spectral range.
Principle of UV-vis spectroscopy
 Ultraviolet absorption spectra arise from transition of electron with in a
molecule from a lower level to a higher level.
 A molecule absorbs ultraviolet radiation of frequency (ϑ ¿, the electron in that
molecule undergo transition from lower to higher energy level.
The energy can be calculated by the equation,
Ε 1−Ε 0=hϑ
E total =E electronic +E vibrational +E rotational

UV radiation
• It is a form of electromagnetic radiation with wavelength from 10nm to 400nm,
shorten than that of visible light but longer than x- rays. UV radiation is
present in sunlight, and constitutes about 10% of the total electromagnetic
radiation output from the sun.
• The region beyond red is called infra-red while that beyond violet is called as
ultra – violet. The wavelength range of UV radiation starts at blue end of
visible light (4000A) & ends at 2000A.
Why we use?
• Detection of functional groups.
• Detection of impurities.
• Qualitative analysis.
• Quantitative analysis.
• Single compound without chromophore.
• Drugs with chromophore reagent.
• Helps to slow the relationship between different groups, it is useful to detect
the conjugation of the compounds.

STRENGTHS OF UV SPECTROSCOPY
 The technique is non-destructive, allowing the sample to be reused or
proceed to further processing or analyses.
 Measurements can be made quickly, allowing easy integration into
experimental protocols.
 Instruments are easy to use, requiring little user training prior to use.
 Data analysis generally requires minimal processing, again meaning little user
training is required.
 The instrument is generally inexpensive to acquire and operate, making it
accessible for many laboratories.

LIMITATIONS OF UV SPECTROSCPY
 The main disadvantage of using a UV-VIS spectrometer is the time it takes to
prepare to use one. With UV-VIS spectrometers, setup is key. You must clear
the area of any outside light, electronic noise, or other outside contaminants
that could interfere with the spectrometer's reading.
 If the space has been properly prepared ahead of time, UV-VIS spectrometers
are simple to use and give accurate results. However, if the space has not
been properly prepared, even a small bit of outside light or vibration from a
small electronic device could interfere with the results you are hoping to
achieve in using a UV-VIS spectrometer

Instrumentation
• Components of spectrophotometer
• Source
 • Monochromator
• Sample compartment
• Detector
• Recorder

APPLICATIONS
1. Pharmaceutical analysis
 One of the most common uses of UV-Vis spectroscopy is in the
pharmaceuticals industry. In particular, processing UV-Vis spectra using
mathematical derivatives allows overlapping absorbance peaks in the original
spectra to be resolved to identify individual pharmaceutical compounds.
 For example, benzocaine, a local anesthetic, and chlortetracycline, an
antibiotic, can be identified simultaneously in commercial veterinary powder
formulations by applying the first mathematical derivative to the absorbance
spectra. Simultaneous quantification of both substances was possible on a
microgram per milliliter concentration range by building a calibration function
for each compound.
2. Bacterial culture
 UV-Vis spectroscopy is often used in bacterial culturing. OD measurements
are routinely and quickly taken using a wavelength of 600 nm to estimate the
cell concentration and to track growth. 600 nm is commonly used and
 preferred due to the optical properties of bacterial culture media in which they
are grown and to avoid damaging the cells in cases where they are required
for continued experimentation.
3. Beverage analysis
 The identification of particular compounds in drinks is another common
application of UV Vis spectroscopy. Caffeine content must be within certain
legal limits, for which UV light can facilitate quantification. Certain classes of
colored substances, such as anthocyanin found in blueberries, raspberries,
blackberries, and cherries, are easily identified by matching their known peak
absorbance wavelengths in wine for quality control using UV Vis absorbance
4. It is used in characterization of atomic compound and in detection of conjugation.
5. UV–Vis spectroscopy is also used in the semiconductor industry to measure the
thickness and optical properties of thin films on a wafer.
6. UV/Vis can be applied to determine the kinetics or rate constant of a chemical
reaction. The reaction, occurring in solution, must present color or brightness shifts
from reactants to products in order to use UV/Vis for this application. For example,
the molecule mercury dithizonate is a yellow-orange color in diluted solution, and
turns blue when subjected with particular wavelengths of visible light (and UV) via a
conformational change.
7. The rate constant of a particular reaction can be determined by measuring the
UV/Vis absorbance spectrum at specific time intervals.
8. Ultraviolet-visible (UV-Vis) spectroscopy is a widely used technique in many areas
of science ranging from bacterial culturing, drug identification and nucleic acid purity
checks and quantitation, to quality control in the beverage industry and chemical
research.
9. An equilibrium constant can also be calculated with UV/Vis spectroscopy. After
determining optimal wavelengths for all species involved in equilibria, a reaction can
be run to equilibrium, and the concentration of species determined from
spectroscopy at various known wavelengths
Conclusion
UV-visible spectroscopy is a valid, simple and cost-effective method for determining
the concentration of absorbing species if applied to pure compounds. From above all
we can conclude that UV/Vis spectroscopy is best method which routinely used in
analytical chemistry for the quantitative determination of different analytes, such as
transition metal ions, highly conjugated organic compounds, and biological
macromolecules. that’s all about our presentation. hope u all understood. thank you
for listening.

Thankyou