Spectroscopy

Applied biochemistry

1. spectroscopy By, Lokeswari b bsc -biotechnology Bon secours college for women thanjavur

2. About spectroscopy; #Spectroscopy is a branch of science which studies the interaction of electromagnetic radiation with matter where the interaction of radiation with chemical species is measured to obtain characteristics quality and quantity of the species.

3. a)ATOMIC SPECTROSCOPY Atomic spectroscopy is based upon the absorption and emission of electromagnetic radiation by atomic particle. a)The first step in all atomic spectroscopic procedures is atomization. *Atomization is a process in which a sample is volatilized and decomposed to produce gas-phase atoms and ions. *It is a critical step in all atomic spectroscopy.

4. a) 1)atomic absorption spectroscopy: It is a type of elemental analysis based on excitation of electron of atom. 2 fundamentals during application of AAS: *sample preparation and introduction, *sample atomization #Many samples like solid,animal tissue,plant leaves,minerals etc.,are not directly used as sample and rather they should be prepared as solution by extensive preliminary treatment. #After sample is prepared from of clear solution ,if it introduced to the instrument for atomization process.In the atomization process sample is nebulized(conversion of samples to mist, ie small droplets of solution) by a flow of gaseous oxidant ( eg ; air oxygen,nitrogen ) mixed with gaseous fuel( eg:naturalgas,hydrogen,ethylene )

5. COMPARISON OF AAS AND AES AAS PROCESS MEASURED # Absorption (light absorbed by unexcited atom) USE OF FLAME #Atomization INSTRUMENTATION #Uses of light source BEER’S LAW #Applicable AES #Emission (light emitted by exited atoms) #Atomization and excitation #Do not use light source #Not applicable

6. Molecular spectroscopy; Molecular spectroscopy  involves the interaction of electromagnetic radiation with materials in order to produce an absorption pattern (i.e. a spectrum) from which structural or compositional information can be deduced. l

7. Ultra violet visible spectroscopy Ultraviolet–visible spectroscopy  or  ultraviolet–visible spectrophotometry  ( UV–Vis  or  UV/Vis ) refers to absorption spectroscopy or reflectance spectroscopy in part of the ultraviolet and the full, adjacent visible spectral regions. This means it uses light in the visible and adjacent ranges. The absorption or reflectance in the visible range directly affects the perceived color of the chemicals involved. In this region of the electromagnetic spectrum ,   atoms and   molecules undergo  electronic transitions . Absorption spectroscopy is complementary to fluorescence spectroscopy, in that fluorescence deals with transitions from the excited state to the ground state, while absorption measures transitions from the ground state to the excited state .

8. PRINCIPLE OF ULTRAVIOLET-VISIBLE ABSORPTION Molecules containing bonding and non-bonding electrons (n-electrons) can absorb energy in the form of ultraviolet or visible light to excite these electrons to higher anti-bonding molecular orbitals .  The more easily excited the electrons (i.e. lower energy gap between the HOMO and the LUMO), the longer the wavelength of light it can absorb . There are four possible types of transitions (π–π*, n–π*, σ–σ*, and n–σ*), and they can be ordered as follows :σ–σ* > n–σ* > π–π* > n–π *.

9. APPLICATIONS ; UV/Vis spectroscopy is routinely used in analytical chemistry for the quantitative determination of different analytes , such as transition metal ions, highly conjugated organic compounds, and biological macromolecules. Spectroscopic analysis is commonly carried out in solutions but solids and gases may also be studied.

10. Infra red spectroscopy; Infrared spectroscopy  ( IR spectroscopy ) is the  spectroscopy  that deals with the  infrared  region of the electromagnetic spectrum, that is light with a longer wavelength and lower frequency than visible light. It covers a range of techniques, mostly based on absorption  spectroscopy.

11. Principles of Ir spectroscopy; The   IR spectroscopy  theory utilizes the concept that molecules tend to absorb specific   frequencies  of light that are characteristic of the corresponding structure of the molecules.

12. Applications 0f ir spectroscopy;   Infrared spectroscopy  is widely used in industry as well as in research. It is a simple and reliable technique for measurement, quality control and dynamic measurement. It is also employed in forensic analysis in civil and criminal analysis .

13. Nmr spectroscopy; Nuclear Magnetic Resonance   is an analytical chemistry technique used in quality control and reserach for determining the content and purity of a sample as well as its molecular structure. For example,  NMR  can quantitatively analyze mixtures containing known compounds .

14. Applications; Nuclear  magnetic resonance spectroscopy  is widely used to determine the structure of organic molecules in solution and study molecular physics, crystals as well as non-crystalline materials.  NMR  is also routinely used in advanced medical imaging techniques, such as in  magnetic resonance  imaging (MRI).

15. Mass spectrometry; Mass spectrometry is an analytical technique that measures the mass-to-charge ratio of ions. The results are typically presented as a mass spectrum, a plot of intensity as a function of the mass-to-charge ratio. 

16. Principle of mass spectroscopy ; A  mass spectrometer generates multiple ions from the sample under investigation, it then separates them according to their specific mass-to-charge ratio (m/z), and then records the relative abundance of each ion type.

17. Components of mass spectrometry

18. Uses of mass spectroscopy; A  mass  spectrum is a plot of the ion signal as a function of the  mass -to-charge ratio. These  spectra  are  used to  determine the elemental or isotopic signature of a sample, the masses of particles and of molecules, and to elucidate the chemical identity or structure of molecules and other chemical compounds.