U.V Spectroscopy

U.V Spectroscopy by shagufta farooqui

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2. 2 Content

3. Spectroscopy: • It is the branch of science that deals with the study of interaction of matter with light. OR • It is the branch of science that deals with the study of interaction of electromagnetic radiation with matter. 3

4. Electromagnetic Radiation: Electromagnetic radiation consist of discrete packages of energy which are called as photons. A photon consists of an oscillating electric field (E) & an oscillating magnetic field (M) which are perpendicular to each other 4

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6. Principles of Spectroscopy: The principle is based on the measurement of spectrum of a sample containing atoms /molecules. Spectrum is a graph of intensity of absorbed or emitted radiation by sample verses frequency (ν) or wavelength ( λ). Spectrometer is an instrument design to measure the spectrum of a compound. 6

7. Principles of Spectroscopy 1. Absorption Spectroscopy: • An analytical technique which concerns with the measurement of absorption of electromagnetic radiation. • e.g. UV (185 - 400 nm) / Visible (400 - 800 nm) Spectroscopy, IR Spectroscopy (0.76 - 15 μ m) 2. Emission Spectroscopy: • An analytical technique in which emission (of a particle or radiation) is dispersed according to some property of the emission & the amount of dispersion is measured. • e.g. Mass Spectroscopy 7

8. Principle: 8 Its is the absorption spectroscopy in which sample solution is placed in light path the sample will absorb the particular wavelength which is characteristic to the functional group present in the light path. T he excitation of electron takes place from higher energy level to lower energy level.The spectrum will recorded which is the graph consist of absorption against wavelength.

9. Spectrophotometer: Components of optical instruments 9

10. A single beam spectrophotometer 10

11. 11 1- Sources of light Sources used in UV-Vis Spectrophotometers are continuous sources. Continuous sources emit radiation of all wavelengths within the spectral region for which they are to be used. Sources of radiation should also be stable and of high intensity.

12. 12 Filters permit certain bands of wavelength (bandwidth of ~ 50 nm) to pass through. The simplest kind of filter is absorption filters , the most common of this type of filters is colored glass filters . They are used in the visible region. The colored glass absorbs a broad portion of the spectrum (complementary color) and transmits other portions (its color). Disadvantage They are not very good wavelength selectors and can’t be used in instruments utilized in research. This is because they allow the passage of a broad bandwidth which gives a chance for deviations from Beer’s law. They absorb a significant fraction of the desired radiation . i- Filters

13. ii- Monochromators They are used for spectral scanning (varying the wavelength of radiation over a considerable range ). They can be used for UV/Vis region. All monochromators are similar in mechanical construction. All monochromators employ slits, mirrors, lenses, gratings or prisms. 13

14. 14 1- Prism monochromators Dispersion by prism depends on refraction of light which is wavelength dependent Violet color with higher energy (shorter wavelength) are diffracted or bent most While red light with lower energy (longer wavelength are diffracted or bent least As a result, the poly- chromatic white light is dispersed to its individual colors.

15. 15 3- Sample compartment (cells) For Visible and UV spectroscopy, a liquid sample is usually contained in a cell called a cuvette . Glass is suitable for visible but not for UV spectroscopy because it absorbs UV radiation. Quartz can be used in UV as well as in visible spectroscopy 1 cm 1 cm Opaque Face Transparent Face Long pathlength Short pathlength (b) 1 cm pathlength cuvet

16. 16 4- Detectors The detectors are devices that convert radiant energy into electrical signal . A Detector should be sensitive , and has a fast response over a considerable range of wavelengths. In addition, the electrical signal produced by the detector must be directly proportional to the transmitted intensity ( linear response ). h  e - -V Photosensitive cathode amplifier i- Phototube anode Phototube emits electrons from a photosensitive, negatively charged cathode when struck by visible or UV radiation The electrons flow through vacuum to an anode to produce current which is proportional to radiation intensity.

17. 17 ii. Photomultiplier tube It is a very sensitive device in which electrons emitted from the photosensitive cathode strike a second surface called dynode which is positive with respect to the original cathode. Electrons are thus accelerated and can knock out more than one electrons from the dynode. If the above process is repeated several times, so more than 10 6 electrons are finally collected for each photon striking the first cathode.

18. Double Beam Spectrophotometer Double beam instrument is the one in which two beams are formed in the space by a U shaped mirror called as beam splitter or beam chopper . Chopper is a device consisting of a circular disc. One third of the disc is opaque and one third is transparent, remaining one third is mirrored. It splits the monochromatic beam of light into two beams of equal intensities. 18

19. 19 Double Beam Spectrophotometer Slit Beam Chopper Reference (Blank) Mirror Mirror Semi-transparent Mirror Tungsten Lamp Grating Photo- multiplier Quartz Cuvette Sample Mirror

20. 20 Advantages of double beam instruments over single beam instruments Single beam spectrophotometer is inconvenient because The sample and blank must be placed alternately in the light path. For measurements at multiple wavelengths, the blank must be run at each wavelength. In double beam instruments The absorption in the sample is automatically corrected for the absorption occurring in the blank, since the readout of the instrument is log the difference between the sample beam and the blank beam. Automatic correction for changes of the source intensity and changes in the detector response with time or wavelength because the two beams are compared and measured at the same time. Automatic scanning and continuous recording of spectrum (absorbance versus wavelength).

21. 21 Applications of U.V Spectroscopy

22. 1. Detection of Impurities  UV absorption spectroscopy is one of the best methods for determination of impurities in organic molecules. Additional peaks can be observed due to impurities in the sample and it can be compared with that of standard raw material. By also measuring the absorbance at specific wavelength, the impurities can be detected. U.V. SPECTRA OF PARACETAMOL (PCM) 22

23. 2.Structure elucidation of organic compounds.  UV spectroscopy is useful in the structure elucidation of organic molecules, the presence or absence of unsaturation, the presence of hetero atoms.  From the location of peaks and combination of peaks, it can be concluded that whether the compound is saturated or unsaturated, hetero atoms are present or not etc . 23

24. 3.QUANTITATIVE ANALYSIS :  UV absorption spectroscopy can be used for the quantitative determination of compounds that absorb UV radiation. This determination is based on Beer’s law which is as follows. A = log I0 / It = log 1/ T = – log T = abc = εbc Where : ε - is extinction co-efficient, c- is concentration, and b- is the length of the cell used in UV spectrophotometer. 24

25. QUALITATIVE ANALYSIS;  UV absorption spectroscopy can characterize those types of compounds which absorbs UV radiation.Identification is done by comparing the absorption spectrum with the spectra of known compounds U.V. SPECTRA'S OF IBUPROFEN 25

26. 4.DETECTION OF FUNCTIONAL GROUPS :  This technique is used to detect the presence or absence of functional group in the compound  Absence of a band at particular wavelength regarded as an evidence for absence of particular group TOLUENE 26

27. REFERENCES: 1. Sharma. Y.R. Elementary Organic Spectroscopy. First edition . S.Chand Publisher; 2010. 2. Chatwal G.R. Instrumental methods of chemical analysis. First edition. Himalaya Publisher; 2010 . 3.Textbook: Principles of instrumental analysis, Skoog et al., 5 th edition, chapter 7, 13. 4.Quantitative chemical analysis, Daniel C. Harris, 6 th edition , chapter 20. 5. Ultra-violet Visible Spectroscopy by Alain Martelli . 27

28. 28 6. M. Hesse, H. Meier, B. Zeeh, Spektroskopische Methoden in der organischen Chemie, 7th ed., Georg Thieme Verlag, Stuttgart, 2005. 7. P. W. Atkins, J. D. Paula, Physikalische Chemie, 5th ed., Wiley - VCH, Weinheim, 2013

29. 29 Thank you