IR SPECTROSCOPY / biophysics/biophysical techniques
- 2. • Infrared radiations refers broadly to the part of electromagnetic spectrum between visible and microwave region. • Infrared spectroscopy or vibrational spectroscopy is concerned with the study of absorption of infrared radiation, which results in vibrational transitions. • Infrared spectroscopy is used in identification of functional groups in pure compounds.
- 3. PRINCIPLE The principle of IR spectroscopy is related to the vibrational and rotational energy of a molecule. When the frequency of the IR radiation is equal to the natural frequency of vibration, the molecule absorb IR radiation. • Absorption of IR radiation causes an excitation of molecule from a lower to the higher vibrational level. • Therefore the IR spectroscopy is also called as “vibrational-rotational spectroscopy”
- 4. FUNDAMENTAL VIBRATIONS • The vibrations are arising when molecule promoted from ground state to lower excited state • The fundamental vibrations for linear and non-linear molecules are determined by following way: Linear molecule : 3n-5 Non-linear molecule :3n-6 Where, ‘n’ is number of atom present in the molecule
- 5. Types of fundamental vibrations: a)Stretching vibration: Distance between two atom increase and decrease but bond angle remains constant. Types of stretching vibrations i) Symmetric stretching vibration: In this case both the atoms stretched or compressed in same direction. ii) Asymmetric stretching vibration: In this vibration one atom undergoes stretching and other atom undergoes compression and vice versa.
- 6. Bending vibrations: Distance between two atom remains constant but bond angle changes. These vibrations can occur either in plane or out of plane. Types of bending vibrations 1) In plane bending vibrations: i) Scissoring: both the atom move towards each other just like scissor. ii) Rocking: both the atoms move in same direction, either in left side or right side.
- 7. 2) Out of plane bending vibration i) Wagging: both the atom move up and down with respect to central atom. ii) Twisting: one atom move up and other atom move down withrespect to central atom.
- 10. Zones of the IR region Three main zones can be identified in the IR region: 1.The far-IR (FIR, 400–10 cm−1, 25–300 μm) 2.The mid-IR (MIR, 4000–400 cm−1, 2.5–25 μm) 3.The near-IR (NIR, 14,000–4000 cm−1, 0.7–2.5 μm)
- 11. The MIR region can be further divided into : The fingerprint region (400–1400 cm−1) and The functional groups’ region (1400–4000 cm−1). • MIR region is the most commonly used where most compounds would have a signature absorption/emission in this region.
- 12. IR ACTIVE AND IR INACTIVE In IR spectroscopy, some of the fundamental vibrations are IR active and some are IR inactive. # There are some selection rules: 1) In a molecule with a centre of symmetry, the vibrations symmetrical about the centre of symmetry are IR-inactive. 2) The vibrations which are not symmetrical about the centre of symmetry are IR-active
- 13. Vibrations which are associated with the change in the dipole moment of the molecule are called IR- active molecules. • All other vibrations are called IR-inactive.
- 14. All the symmetrical diatomic molecules such as H2, N2 and Cl2 etc are IR-inactive. The symmetrical stretching of the C=C bond in ethylene (centre of symmetry) is IR-inactive. • The symmetrical stretching in CO2 is IR-inactive, whereas asymmetric stretching is IR-active.
- 16. FINGERPRINT REGION In IR, the region rich in many absorption bands and the region is known as fingerprint region. Here the number of bending vibrations are usually more than the number of stretching vibrations. In this region, small difference in the structure and constitution of a molecule results significant changes in the absorption bands. • Many compounds show unique absorption bands in this region and which is very useful for the identification of the compound.
- 17. FUNCTIONAL GROUP’S REGION Function group region contain relatively few peaks which are associated with the stretching vibration of functional group.
- 19. OVERTONES & COMBINATION BANDS • When molecule absorbed electromagnetic radiation in IR region, and then molecule promoted from ground state to second, third or even fourth vibrational excited state. These bands are known as Overtones. The intensity of these bands is very weak. It is helpful in characterization of aromatic compounds. • When two fundamental vibrational frequencies (ν1 + ν2) in a molecule couple to give rise to a new vibrational frequency within the molecule, it is known as combination band.
- 21. FOURIER TRANSFORM IR SPECTROMETER (FTIR) • Fourier transform spectrometers have recently replaced dispersive instruments for most applications due to their superior speed and sensitivity. • They have greatly extended the capabilities of infrared spectroscopy and have been applied to many areas that are very difficult or nearly impossible to analyze by dispersive instruments. • Instead of viewing each component frequency sequentially, as in a dispersive IR spectrometer, all frequencies are examined simultaneously in Fourier transform infrared (FTIR) spectroscopy
- 22. APPLICATIONS • Identification of functional group and structure elucidation. • Identification of substances. • Studying the progress of the reaction. • Quantitative analysis • Detection of impurities.