Applications of UV- Visible Spectroscpoy.pptx

1. Applications of UV-
Visible Spectroscpoy
SUBMITTED BY: HAFIZA REHANA BATOOL
ROLL NUMBER: 29777
SUBMITTED TO: DR. FREEHA HAFEEZ

2. INTRODUCTION
 UV-Vis spectroscopy is an analytical technique that
measures the amount of discrete wavelengths of UV or
visible light
 This light is absorbed by or transmitted through a
sample in comparison to a
reference or blank sample.

3. UV/VIS SPECTROPHOTOMETER
 A UV-vis spectrophotometer is an analytical instrument
that measures the amount of ultraviolet (UV) and visible
light that is absorbed by a sample.
 It is a widely used technique in chemistry, biochemistry,
and other fields, to identify and quantify compounds in a
variety of samples.

4. Applications of UV-Visible
Spectroscopy
 UV-Vis spectroscopy has many different applications in
organic and biological chemistry.
 One of The most basic of these applications is the use of
the Beer - Lambert Law to determine the concentration of
a substance

5. 1-Application of UV/VIS in water
analysis
Basic principle
 The Lambert-Beer law is the basis of quantitative analysis
of water quality parameters using UV-Vis spectroscopy.

6. Beer - Lambert Law
 According to the Beer-Lambert Law the absorbance is
proportional to the concentration of the substance in
solution
A = εLc
 So UV-visible spectroscopy can also be used to measure
the concentration of a sample.

7. Determination of Pollutants
 Pollutants differ by absorption characteristics and yielded
spectral curves but most of them absorb light in the UV-
Vis region.
 UV-Vis spectroscopy is based on the fact that pollutant
molecules in the water can absorb UV-Vis light of a
specific wavelength.
 Light of specific wavelength is absorbed by electron
movement from the ground state to an excited state.

9.  Determination of COD in Water
 Detection of Heavy Metal Ions in Water
 Detection of Nitrate Nitrogen in Water
 Detection of DOC in Water

10. 2-Determination the
concentration of a protein using
ultraviolet (UV) spectroscopy
 Estimation of protein concentration in a given protein
preparation is one of the most commonly performed
tasks in a biochemistry lab.
 If a protein is pure, UV spectroscopic quantitation is the
method of choice
 It is easy and less time consuming to perform
furthermore, the protein sample can be recovered back.

11.  Aromatic amino acids, tryptophan, tyrosine, and
phenylalanine and the disulfide linkage
constitute the chromophores that absorb in the near UV
region
 Their absorbance values are 250,279.280 nm respectively

13. ABSORBANCE OF PEPTIDE BOND
 The proteins and peptides that lack aromatic residues
and disulfide linkage do not absorb the near UV
radiation.
 The concentration of such proteins and peptides can be
estimated using far UV radiation.
 Peptide bond is the major chromophore in the far
UV region with a strong absorption band around 190 nm
(π → π* transition) and a weak band around 220 nm (n →
π* transition)

14. DETERMINATION OF DNA AND RNA
 DNA or RNA purity can also be determined by measuring
the absorption peaks of 260 nm
 This is because the nucleic acids that make up DNA and
RNA absorb strongly at 260 nm.

16. Advantages of UV/VIS
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