UV Spectroscopy Lecture Notes PDF

SPECTROSCOPY  GENERAL INTRODUCTION  Use of the radiations for quantitative and qualitative study of sample.  Absorption spectroscopy can be better understood from consideration of Electromagnetic spectrum and how molecules absorb radiation.  Electromagnetic radiatio...

SPECTROSCOPY  GENERAL INTRODUCTION  Use of the radiations for quantitative and qualitative study of sample.  Absorption spectroscopy can be better understood from consideration of Electromagnetic spectrum and how molecules absorb radiation.  Electromagnetic radiation posses a certain amount of energy.  This energy travels in waves, each wave has small particles of energy called PHOTONS.  The energy of each PHOTON is not the same.  The energy contained in each photon is directly proportional to the frequency of the electromagnetic wave.  The total energy in a beam (No of waves) of E.M.R is the sum of energies of its PHOTONS (oscillating electric and magnetic field; orthogonal).  Light moves in the form of waves along its line of travel. [fig 1]  Definitions Wave length (λ): Distance of one complete cycle. Wave number(v): Reciprocal of wave length; the number of waves in a unit length or (distance per cycle). Frequency(V): The No of cycles passing a fixed point per unit time : ie No. of cycles/ second.  The E.M.R consists of different size wavelengths ranging from short wave length (high energy) to long wave length (low energy).  These wave lengths cover: Gamma, X‐ray, UV‐Visible, I.R, microwaves and lastly the radio wave (lowest energy). -rays X-rays UV IR Microwave Radio Visible The formula that relates frequency and the wave‐length to the amount of energy is: λ = c/V E = hV i.e E= hc /λ where h is the plank’ constant ( 6.63x 10 Js, 6.63 x 10 Kjsor 4.132 x 10 eVs), c= velocity of light (3x10 cm/s or 3x10 m s 1.  ΔE is directly proportional tov and inverselyproportional toλ.  In numerical values, and ifλ = 300 nm.: λ in cm. = 300 x 10‐7cm. ΔE (kjs)= 6.63 x 10 x (kjs)x 3 x cm/s 300 x (cm)  Toobtain ΔE in terms of Kcal, we use conversion value 4.184 to convert Kjs into kcal (divide by).  To express the energy absorbed/ mole we multiply by AVOGADOR’S number ( 6.02 x )  Important ranges for Pharmaceutical analysis 1. 10‐190nm (σ – σ, less useful) 2. 200‐390nm (useful‐ UV –range) 3. 390‐700nm (visible range) and 4. 700‐1060 nm I.R range.  The visible range (390‐700) has the violet as shortest λ and red as the longest wave length.  Processes that determine the color of an object or a solution are : 1. Light emission. 2. Light reflection. 3. Light transmission or absorption. i) Light emission: Absorption of energy from a light source ;excitation of the object by the gained energy; released energy as emitted photons.  This process is the basis of Flame emission and Fluorescence. ii) Light reflection : A non transparent object absorbs all the wave length in the spectrum with the exception of the ones which we can see. i.e a black object absorbs all visible light. A white object reflects all. iii) Light transmission or absorption An object of a certain color transmits that color we see and absorbs the rest of the spectrum. The transmitted color is complementary to the absorbed color i.e a blue color indicates the absorption of a yellow colour etc  The absorption of radiation by molecules There are three processes by which a molecule can absorb radiation energy. The three types are quantized i.e they exist at discrete levels. These processes or transitions are: (1)The electronic transition (major): The electrons of a molecule absorb suitable energy and become raised to a higher energy level. (2) Vibrational transition: A toms or group of atoms within a molecule vibrate relative to each other and the energy of this vibration occurs at definite quantized levels. The molecule may be then raised to a higher vibrational energy level. (3)Rotational transitions: Molecules rotate about various axis, the energy of rotation being at definite energy levels. The molecule will be raised to a higher rotational energy level. Electrons in a molecule are classified as : 1/ Closed shell electrons : not involved in bonding. i.e require very high energy for excitation :. not suitable for UV‐Visible range. 2/ Covalent single bond electrons (σ, sigma). Require very high energy :. not suitable for UV‐ Vis(‐CH2‐CH2: Single valence bonds in saturated hydrocarbons).  3/ Paired non‐ bonding outer shell electrons (n electrons) such as N,O,S and halogens. These are less tightly held than sigma and can be excited by UV‐Vis. Radiation i.e useful. (These give n‐ π * transitions and associated with groups such as C=O , thio group, C=S, nitroso N=O. They give much lower intensity than π→ π*).  4/ Electrons in π (pi) orbitals. i.e Double or triple bonds. These are readily excited by UV‐Vis radiation and are responsible for majority of electronic spectra in UV‐ Vis.(These give π – π* transitions and usually associated with the multiple bonds of C=C,C ≡ C, etc. They give HIGH INTENSITY absorption) A molecule also possess unoccupied orbitals called ANTIBONDING orbitals or excited б* or π* levels.: Absorption of radiation results in an electronic transition from a bonding (б or π) or non –bonding (n) to antibonding б*or π*. The most common and useful transitions are π → π* or n → π*, these correspond to λs above 200nm; while n→ б* occur at λs below 200 nm. Examples: Ketones show both π →π* and n →π* The probability of π→ π* transitions is greater than n→ π* transitions. Molar absoropitivites for π→π* transitions are about 1000‐ 100,000 compared to 1000 for n→ π* transitions. 1. б → б* Transition : Required energy of 150 nm (Hydrocarbons). 2. n → б* Transition : Required energy of 175 nm (Alcohols, ketones, aldehydes& water). For this reasons we can use the above as solvents in UV‐ Spectrophotometry because it is out of range (200‐800 nm). 3. π → π* Transition: Required energy more than 200 nm (double and triple bonds and Aromatic compounds). 4. n → π* Transition: Required more than 200 nm (Conjugation in some compounds like carbonyle compounds). No 1 & 2 these transitions are forbidden transitions and are only theoretically possible. No 3 & 4 transitions shows absorptions at wave lengths above 200 nm which is accessible for the UV‐Visible spectrophotometer. Therefore, prolonged exposure of the sample to UV - radiation during measurement should be avoided to minimize possible decomposition of a proportion of the sample.  Energies of these magnitudes are associated with the promotion of an electron from a non‐bonding (n) orbital or a π bonding orbital to an anti‐ bonding orbital (π*) excited state or to an anti bonding sigma excited σ* state