Md Rashed Hassan
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UV-visible spectroscopy involves using light in the ultraviolet and visible spectral regions to analyze organic compounds. It works by measuring how much light is absorbed by a compound at different wavelengths, which provides information about the compound's structure and functional groups. Key principles are that UV radiation extends from 10-400 nm while visible is 400-800 nm. Absorption is governed by Lambert's and Beer's laws. Different types of electronic transitions that can occur include σ→σ*, n→σ*, π→π*, and n→π*. UV-visible spectroscopy has many uses such as detection of impurities, structure elucidation, quantitative analysis based on Beer's law, identification of unknown compounds by comparing spectra, and studying reaction kinetics
Md Rashed Hassan
- 1. ASSIGNMENT ON UV-Visible Spectroscopy Course Title : Advanced Pharmaceutical Analysis & Quality Control Course Code: BPH 414 Submitted To: Sabiha Enam Spriha Lecturer Department of Pharmacy Daffodil International University Dateofsubmission: 10/06/2021 Submitted By: Name: Md Rashed Hassan ID: 181-29-1203 Department of Pharmacy Section: 19th AC-B
- 2. UV-Visible Spectroscopy Definition: 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. History of spectroscopy: • Spectroscopy began with Isaac Newton's optics experiments (1666–1672). Newton applied the word "spectrum" to describe the rainbow of colors. • During the early 1800s, Joseph von Fraunhofer made experimental advances with dispersive spectrometers that enabled spectroscopy to become a more precise and quantitative scientific technique. Principles of UV-visible spectroscopy: 1. The UV radiation region extends from 10 nm to 400 nm and the visible radiation region extends from 400 nm to 800 nm. • Near UV Region: 200 nm to 400 nm • Far UV Region: below 200 nm 2. Far UV spectroscopy is studied under vacuum condition. 3. The common solvent used for preparing sample to be analyzed is either ethyl alcohol or hexane. Laws of spectrometry: The absorption of light by any absorbing material is govern by two laws. 1. Lambert's Law. 2. Beer's Law.
- 3. Types of electronic Transition: According to molecular orbital theory when a molecule is excited by the absorption of UV or visible light its electrons are changed from one orbital to another orbital of higher energy. The electrons can be σ, π or an n electron. The various transitions involved in ultraviolet-visible spectroscopy are: (i)σ→ σ* transition: This type of transition takes place in saturated centers of the molecules. Here, an electron from a stable σ orbital goes to an unstable or anti bonding σ orbital. (ii) n →σ* transition: This type of transition takes place in saturated compounds containing a heteroatom with unshared pair of electrons (n electrons). Here, an electron from an unshared pair goes to an unstable or anti bonding σ orbital. (iii) π → π* transition: This type of transition takes place in unsaturated compounds containing double or triple bonds. Here, an electron from stable or π orbital goes to an unstable or antibonding π orbital. Alkenes, alkynes, carbonyl compounds, azo compounds etc. show π » π* transition. (iv) n → π* transition: This type of transition takes place in unsaturated compounds containing a heteroatom with unshared pair of electrons (n electrons). Here, an electron from an unshared pair goes to an unstable π orbital. How do conjugated double bond affects lambda max of a compound? When a double-bonded molecule such as ethene (common name ethylene) absorbs light, it undergoes a π - π* transition. Because π- π* energy gaps are narrower than σ - σ* gaps, ethene absorbs light at 165 nm - a longer wavelength than molecular hydrogen.
- 4. Longer frequency = smaller energy, this means that the energy gap ΔE between the highest- occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) decreases as the number of conjugated pi bonds Increases. Energy gap π- π* transition is smaller than for isolated double bond and thus the wavelength absorbed longer. Uses of UV-visible Spectroscopy: Some uses of Uv-visible spectroscopy are given below: 1. Detection of Impurities: It is one of the important methods to detect impurities in organic solvents. Additional peaks can be observed due to impurities in the sample and it can be compared with that of standard raw material. 2. Structure elucidation of organic compounds: The presence or absence of unsaturation, the presence of hetero atoms like S, N, O or halogens can be determined. 3. Structural analysis of organic compounds: Effect of conjugation: Extended conjugation shifts the max to longer x (Bathochromic shift) and reduction of the compound or saturation of double bonds leads to the opposite effect. I.e., hypochromic shift.
- 5. 4. 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 that is used in UV spectrophotometer. 5. 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 5. Chemical kinetics: Kinetics of reaction can also be studied using UV spectroscopy. 6. Fingerprint Region: • Small differences in structure & constitution of molecule result in significant changes in the peak in region. • Hence this region helps to identify an unknown compound. 7. Examination of poly nuclear hydrocarbons: Benzene and Polynuclear hydrocarbons have characteristic spectra in ultraviolet and visible region. Thus, identification of Polynuclear hydrocarbons can be made by comparison with the spectra of known Polynuclear compounds. Polynuclear hydrocarbons are the Hydrocarbon molecule with two or more closed rings; examples are naphthalene, C10H8, with two benzene rings side by side, or diphenyl, (C6H5)2, with two bond- connected benzene rings. Also known as polycyclic hydrocarbon. 8. Molecular weight determination: Molecular weights of compounds can be measured spectrophotometrically by preparing the suitable derivatives of these compounds. For example, if we want to determine the molecular weight of amine then it is converted in to amine picrate. Then known concentration of amine picrate is dissolved in a liter of solution and its optical density is measured at λ max 380 nm.
- 6. 9. Chemical kinetics: Kinetics of reaction can also be studied using UV spectroscopy. The UV radiation is passed through the reaction cell and the absorbance changes can be observed. 10. Identification of Substances: • To compare spectrums. • No to sample will have identical Ir spectrum. • Criteria: Sample and reference must be tested in identical conditions, like physical state, temperature, solvent etc. 11. Determination of Molecular Structure: • Used along with other spectroscopic techniques. • Identification is done based on position of absorption bands in the spectrum. e.g: C=O at 1717 cm-1 • Absence of band of a particular group indicates absence of that group in the compound. 12. To detect functional group: 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. 13. Spectroscopy in Biomedical Sciences: The biomedical use of light comprises numerous diagnostic and therapeutic applications. Photon time-of-flight spectroscopy may assist certain therapeutic methods (such as photodynamic therapy) by supplying them with the data on the optical properties governing tissue response. 14. As HPLC detector: A UV/Vis spectrophotometer may be used as a detector forHPLC.
- 7. References: 1. https://www.wikipedia.com 2. https://www.google.com/url?q=https://www2.chemistry.msu.edu/faculty/reusch/virttxtjml/spectrpy/uv- vis/spectrum.htm&sa=U&ved=2ahUKEwjR7tTJhJnvAhXvwzgGHTyKASsQFjAJegQIChAB&usg=AOvVaw3brhnc mw3vB4c0O_F1qsBz 3. www.chem.agilent.com/Library/applications/uv31.pd f 4. www.uvitron.com/Applications.html. 5. https://images.app.goo.gl/ubjU1jyMHvWUAVq778