UV-Basic Principles Lecture Notes PDF

Lecture Title UV- basic principles 1 Intended learning objectives At the end of the lecture students will be able to State Lambert and Beer’s laws Derive the fundamental equation of quantitative spectroscopy Explain the terms absorbance and transmittance Distinguis...

Lecture Title UV- basic principles 1 Intended learning objectives At the end of the lecture students will be able to State Lambert and Beer’s laws Derive the fundamental equation of quantitative spectroscopy Explain the terms absorbance and transmittance Distinguish between absorption coefficient, specific absorption coefficient and molar absorption coefficient 2 Principle 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 Far UV spectroscopy is studied under vacuum condition. The common solvent used for preparing sample to be analyzed is either ethyl alcohol or hexane. 3 Interaction of light and matter 4 Beer’s Law Beer’s law states that when monochromatic light is passed through a solution of uniform concentration, the rate of decrease in intensity of light is proportional to the intensity of light 𝑑𝐼 𝑑𝐼 − ∝𝐼 i.e − = 𝑘′𝐼 −𝑑𝐼/𝐼=𝑘′dc 𝑑𝑐 𝑑𝑐 Where K’= Absorption coefficient 5 Beer’s Law 𝑑𝐼 𝑑𝐼 − ∝ 𝐼 i.e − = 𝑘′𝐼 𝑑𝑐 𝑑𝑐 𝑑𝐼  − = 𝑘 ′ dc 𝐼 𝐼𝑡 𝑑𝐼 𝑐 − 𝐼 = 𝑘′ 0 𝑑𝑐 0 𝐼 Equation shows that the intensity of a monochromatic beam of light decreases exponentially when the concentration of solution increase −(𝑙𝑛𝐼𝑡 − 𝑙𝑛𝐼0 ) = k’ c Changing natural logarithm to the base 10, equation becomes, 𝐼0 𝑘′  log = 𝑐 𝐼𝑡 2.303 𝐼 𝐴 = log 0 = 𝐾′𝑐 The term log(Io/It) is called absorbance(A) 𝐼𝑡 Where A = Absorbance, K’= Absorption coefficient 6 Lambert’s Law Lambert’s law states that when monochromatic light passes through a transparent medium of uniform thickness, the rate of decrease in the intensity of light is directly proportional to the intensity of light I is the intensity of radiation b is pathlength (length of the medium through which light travelled) dI and db are the differences in intensity and pathlength respectively indicates decrease - indicates decrease 7 Lambert’s law 𝑑𝐼 − = 𝑘 𝐼 𝑑𝑏 𝑑𝐼 − = 𝑘 𝑑𝑏 𝐼 𝐼𝑡 𝑑𝐼 𝑏 − 𝐼 = 𝑘 0 𝑑𝑏 0 𝐼 Equation shows that the intensity of a monochromatic beam of light decreases exponentially when the thickness of the medium and intensity increase. −(𝑙𝑛𝐼𝑡 − 𝑙𝑛𝐼0 ) = k b Changing natural logarithm to the base 10, equation becomes, 𝐼 𝑘 log 0 = 𝑏 𝐼𝑡 2.303 𝐼0 𝐴 = log = 𝐾𝑏 𝐼𝑡 Where A = Absorbance, K= Absorption coefficient 8 Beer- lambert’s Law Combining both the equations, 𝑰𝟎 𝑨= 𝐥𝐨𝐠 = 𝒂𝒃𝒄 𝑰𝒕 where 𝑎 is specific absorption coefficient, the value of which is dependent on the way concentration is expressed and on the unit of path length. This is the fundamental equation of spectroscopy Also, A=log(1/T) or A=-log( T ) Hence, the absorbance is equal to the negative logarithm of transmittance 9 Terms used in spectrophotometry Transmittance is the ratio of intensity of transmitted light to that of incident light Absorbance: logarithmic ratio of intensity of incident light to that of transmitted light Synonyms: Optical density, Extinction Absorption coefficient: Absorbance per unit path length Classical definition: the reciprocal of path length in cm that is required to reduce the intensity of incident light to 1/10th of its value Synonyms: Absorptivity, extinction coefficient 10 Terms used in spectrophotometry Specific absorption coefficient : Absorbance per unit concentration per unit path length 𝐴 𝑎= 𝑏𝑐 𝐴1%1cm : A specific absorption coefficient when concentration is expressed as % solution and path length in 1 cm 11 Molar absorption coefficient and Molecular weight Molar absorption Coefficient (ɛ): Specific absorption coefficient when concentration is expressed as moles per liter solution and path length in cm Molecular weight in g = 1 Mole Assume Concentration of the solution is c % solution, c g is c/M.wt moles Concentration of the solution is 10c/M.wt moles/litre 𝐴 𝑀.𝑤𝑡 𝜀= 𝑏𝑐 10 Molar absorption coefficient and Molecular weight 12 Beer- Lamberts Law: Beer Lamberts Law: A=εbc A = absorbance ε = molar absorptivity with units of L /mol.cm b = path length of the sample (cuvette) c = Concentration of the compound in solution, (mol /L) 13 Deviations from Beer’s Law Beer's law is subjected to certain real and apparent deviations. Real deviations are most usually encountered in relatively concentrated solutions of the absorbing compound (>0,01 M). These deviations are due to interactions between the absorbing species and to alterations of the refractive index of the medium 14 Deviations from Beer’s Law Most common are the apparent deviations These deviations are due to: chemical reasons arising when the absorbing compound, dissociates, associates, or reacts with a solvent to produce a product having a different absorption spectrum presence of stray radiation polychromatic radiation 15 Limitations of the Beer-Lambert law The linearity of the Beer-Lambert law is limited by chemical and instrumental factors. Causes of nonlinearity include: Deviations in absorptivity coefficients at high concentrations (>0.01M) due to electrostatic interactions between molecules in close proximity Interaction with solvent: hydrogen bonding Scattering of light due to particulates in the sample Fluoresecence or phosphorescence- a positive deviation in % T and negative deviation for A 16 Limitations of the Beer-Lambert law Changes in refractive index at high analyte concentration Shifts in chemical equilibria as a function of concentration Non-monochromatic radiation, deviations can be minimized by using a relatively flat part of the absorption spectrum such as the maximum of an absorption band Stray light 17 Isosbestic point Isosbestic point is a specific wavelength, wavenumber or frequency at which the total absorbance of a sample does not change during a chemical reaction or a physical change of the sample. The word derives from two Greek words: "iso", meaning "equal", and "sbestos", meaning "extinguishable". Isobestic point of methyl orange 18 Electronic Transitions 19 Bonding orbitals i) σ (bonding) molecular as in ii) π (bonding) molecular orbital as in iii) n (non-bonding) atomic orbital as in 20 Antibonding orbitals In addition, two types of antibonding orbitals may be involved in the transition: i) σ* (sigma star) orbital ii) π* (pi star) orbital (There is no such thing as an n* antibonding orbital as the n electrons do not form bonds) 21 The possible electronic transitions are 1 σ → σ* transition 2 π → π* transition 3 n → σ* transition 4 n → π* transition 5 σ → π* transition 6 π → σ* transition 22 The possible electronic transitions can graphically shown as: 23  1 σ → σ* transition σ electron from orbital is excited to corresponding anti-bonding orbital σ* The energy required is large for this transition e.g. Methane (CH4) has C-H bond only and can undergo σ → σ* transition and shows absorbance maxima at 125 nm 24  2 π → π* transition K-band (Konjugerite-Conjugated systems) B-band (Benzenoid)- Chromopohre shifts B band at longer than K band E-band (Benzenoid of 3 ethylenic bonds) π electron in a bonding orbital is excited to corresponding anti- bonding orbital π* Allowed transitions Compounds containing multiple bonds like alkenes, alkynes, carbonyl, nitriles, aromatic compounds, etc undergo π → π* transitions. e.g. Alkenes generally absorb in the region 170 to 205 nm 25  3 n → σ* transition Saturated compounds containing atoms with lone pair of electrons like O, N, S and halogens are capable of n → σ* transition. Forbidden transition These transitions usually requires less energy than σ → σ* transitions. The number of organic functional groups with n → σ* peaks in UV region is small (150 – 250 nm). 26  4 n → π* transition R-bands (Radiclartig) An electron from non-bonding orbital is promoted to anti-bonding π* orbital. Compounds containing double bond involving hetero atoms (C=O, C≡N, N=O) undergo such transitions. n → π* transitions require minimum energy and show absorption at longer wavelength around 300 nm. 27  5 σ → π* transition 6 π → σ* transition These electronic transitions are forbidden transitions & are only theoretically possible. Thus, n → π* & π → π* electronic transitions show absorption in region above 200 nm which is accessible to UV-visible spectrophotometer. The UV spectrum is of only a few broad of absorption. 28 Transitions Both s to σ* and n to σ* transitions require a great deal of energy and therefore occur in the far ultraviolet region or weakly in the region 180-240nm. Consequently, saturated groups do not exhibit strong absorption in the ordinary ultraviolet region. Transitions of the n to π* and π to π* type occur in molecules with unsaturated centers; they require less energy and occur at longer wavelenghts 29 Summary Lambert’s law explains the effect of path length on the intensity of light Beer’s law explains the effect of concentration of the solution on the intensity of light Both laws assume monochromatic light Absorbance is logarithmic ratio of intensity of incident light to that of transmitted light Transmittance is the ratio of intensity of transmitted light to that of incident light The combined law gives the fundamental equation of quantitative spectroscopy 30 Summary The fundamental equation of spectroscopy is 𝐼0 𝐴= log = 𝑎𝑏𝑐 𝐼𝑡 where 𝑎 is specific absorption coefficient, the value of which is dependent on the way concentration is expressed and on the unit of path length. 31 Thank you 32