Spectroscopy Part 1 PDF

## Introduction - Spectroscopy is the study of the interaction between matter and electromagnetic radiation as a function of the wavelength or frequency of the radiation. - It is the measurement of electromagnetic radiation (EMR) absorbed or omitted when a molecule or ions or atoms of a sample move...

## Introduction - Spectroscopy is the study of the interaction between matter and electromagnetic radiation as a function of the wavelength or frequency of the radiation. - It is the measurement of electromagnetic radiation (EMR) absorbed or omitted when a molecule or ions or atoms of a sample move from one energy state to another. ## Electromagnetic Radiation - Electromagnetic radiation consist of discrete packets of energy which are called as photons. - A photon consists of an oscillating electric field (E) & an oscillating magnetic field (M) which are perpendicular to each other. ## Properties of Electromagnetic Radiation - **Wave length:** The distance between two successive maxima on an electromagnetic wave. Units are: m, cm, mm, nm, and micrometer. - **Frequency:** The number of wavelength units pass through a given point in unit time is called frequency. Denoted by "v", the units are cycles per second, Hertz. - **Wave number:** The number of waves per cm in a vacuum. $v=1/wavelength$ - **Velocity:** The product of wavelength and frequency and is equal to the velocity of the wave in the medium. $V = η χλ$ - The relationship between wavelength & frequency can be written as: $c = ν λ$. - As a photon is subjected to energy, $E = h v=hc/λ$. ## Electronic Transitions - **σ→ σ* transition:** An electron in a bonding s-orbital is excited to the corresponding anti-bonding orbital and observed with saturated compounds. The energy required is large. For example, methane (which has only C-H bonds, and can only undergo σ→ σ* transition transitions) shows an absorbance maximum at 125 nm. Absorption maxima due to σ→ σ* transition are not seen in typical UV-VIS spectra (200 - 700 nm) but in UV-region (125-135nm). - **n → σ* transition:** Saturated compounds containing atoms with lone pairs (non-bonding electrons) like O, N, S and halogens are capable of n→ σ* transition. These transitions usually need less energy than n → σ* transition. They can initiated by light whose wavelength in range 150-250 nm. The number of organic functional groups with n → σ* peak in the UV region is small. - **π→ π* transition:** π electron in a bonding orbital is excited to corresponding anti-bonding orbital π* and observed in conjugated compounds. Compounds containing multiple bonds like alkenes, alkynes, carbonyl, nitriles, aromatic compounds, etc undergo π - π* transitions. For example, alkenes generally absorb in the region 170 to 205 nm. - **n → π* transition:** An electron from non-bonding orbital is promoted to anti-bonding π* orbital and required lower energy. 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. | Electronic Transition | Description | Energy Required | Wavelength | | :------------------------- | :---------- | :----------------- | :---------- | | σ→ σ* | An electron is excited from a bonding s-orbital to the corresponding anti-bonding orbital. Observed with saturated compounds. | Large | 125-135 nm | | n → σ* | An electron lone pair is excited to an anti-bonding orbital. Observed with saturated compounds containing atoms with lone pairs like O, N, S, and halogens. | Smaller | 150-250 nm | | π→ π* | An electron is excited from a bonding orbital to an anti-bonding orbital π*. Observed in conjugated compounds. | Medium | 170-205 nm | | n → π* | An electron from non-bonding orbital is promoted to anti-bonding π* | Lower | Around 300 nm | ## Deviations - When a non-linear curve is obtained, the system is said to undergo** deviation**. There are two types of deviations: **positive deviation** and **negative deviation**. - **Positive deviation** results when a small change in concentration produces a greater change in absorbance. - **Negative deviation** results when a large change in concentration produces smaller change in absorbance. **Reasons** 1. **Instrumental deviations:** stray radiation, improper slit width fluctuations in single beam and monochromatic light is not used. 2. **Physiochemical changes in solutions:** factors like association, dissociation, ionization (change in pH), faulty development of color (incompletion of reaction). ## Chromophore - Any isolated covalently bonded group that shows a characteristic absorption in the UV/Visible region. - Examples: -C=C-, C=O - The functional groups containing multiple bonds capable of Absorbing radiations above 200 nm due to n→π* & π→π* transitions. - Examples: NO2, N=O, C=O, C=N, C=N, C=C, C=S, etc. - Any substance (groups) which absorbs radiation at particular wavelength this may or may not impart colour to the compound. ## Auxochrome - The functional group with non-bonding electrons that does not absorb radiation in near UV region but when attached to a chromophore alters the wavelength & intensity of absorption. - Examples: Benzene λ*max*=255 nm, Phenol λ*max*=270 nm, Aniline λ*max*=280 nm. - The new chromophore that is formed is of have a different value of absorption maximum as well as the extinction coefficient. - Examples: Benzene - 255nm (ɛ*max* - 203) and Aniline - 280nm (E*max*- 1430), so the auxochrome group is - NH. - Other examples: - OH, - OR, -NH, -NHR, -NR,, -SH etc., ## Adsorption and Intensity Shifts 1. **Bathochromic Shift (Red Shift):** When absorption maxima (λ*max*) of a compound shifts to longer wavelength, it is known as bathochromic shift or red shift. 2. **Hypsochromic Shift (Blue Shift):** When absorption maxima (λ*max*) of a compound shifts to shorter wavelength, it is known as hypsochromic shift or blue shift. 3. **Hyperchromic Effect:** When absorption intensity (ε) of a compound is increased, it is known as hyperchromic shift. 4. **Hypochromic Effect:** When absorption intensity (ε) of a compound is decreased, it is known as hypochromic shift. ## Laws Involved 1. **Beer's Law:** When a beam of monochromatic light is passed through a homogeneous absorbing medium, the rate of decrease of intensity of radiation with increase in the concentration (c) of absorbing species is directly proportional to the intensity (I) of the incident light (radiation). $-dI/dc = k I$ and $-dI/I = k dc$. On integration: `-ln I = k c + b` (b= integration constant). When concentration = 0, there is no absorbance. Here, I = I_o. Therefore, substitution into the integrated equation: `-ln I = kx0+b` and `-ln I = b`. Substituting the value of b: `-ln I = k c - ln I<sub>o</sub>`. Rearranging: `ln I<sub>o</sub> - ln I=kc`. Using the log rules: `ln I<sub>o</sub> / ln I = kc` and `I<sub>o</sub> / I = e<sup> kc</sup>` (removing natural logarithm). Rearranging again: `I/I<sub>o</sub> = e<sup>-kc</sup>` (inverse both sides). So `I = I<sub>o</sub> e<sup>-kc</sup>`. 2. **Lambert's Law:** When a beam of monochromatic light is passed through a homogenous absorbing medium, the rate of decrease of intensity of radiation with thickness of absorbing medium is directly proportional to the intensity of the incident light (radiation). `dI/dt = kI`. I= intensity of incident light of wavelength λ & t= thickness of medium. So, `I = I<sub>o</sub> e<sup>-kt</sup>`. 3. **Beer-Lambert's Law:** Combining the above equations: `I = I<sub>o</sub> e<sup>-kct</sup>`. Converting natural logarithm to base 10: `I = I<sub>o</sub> 10<sup>-kct</sup>`. Inverting to isolate I₀/I: `I<sub>o</sub> / I = 10<sup>kct</sup>`. Taking the log on both sides: `log I<sub>o</sub> / I=kct`. Here, transmittance (T) = I/I_o, and Absorbance (A) = log 1/T. So: `A = log I<sub>o</sub>/I`. Using the derivations from Beer's Law and Lambert's Law: `A=kct`. Instead of k we can use ε, the above equation will be as follow: `A=ect`, which is the mathematical equation for Beer's-Lambert's Law. **A=ect** Where: - A = Absorbance - ε = Molecular extinction coefficient - c = Concentration of sample - t = Path length (normally 10mm or 1cm) ɛ can be expressed as follows: ε = E_1%^1cm x Molecular weight / 10 ## Instrumentation - Source of radiation. - Collimating system. - Mono-chromator system. - Sample holder. - Detector. - Amplifier - Read-out devices ## Source of Radiation **Requirements of an ideal source:** - It should be stable and should not allow fluctuations. - It should emit light of continuous spectrum of high and uniform intensity over the entire wavelength region in which it's used. - It should provide incident light of sufficient intensity for the transmitted energy to be detected at the end of optic path. - It should not show fatigue on continued use. **1. Tungsten Halogen Lamp:** - Its construction is similar to a house hold lamp. - The bulb contains a filament of Tungsten fixed in evacuated condition and then filled with inert gas. - The filament can be heated up to 3000 k, beyond this Tungsten starts sublimating. - It is used when polychromatic light is required. **Demerit:** - It emits the major portion of its radiant energy in near IR region of the spectrum. **2. Hydrogen Discharge Lamp:** - In Hydrogen discharge lamp, a pair of electrodes is enclosed in a glass tube (provided with silica or quartz window for UV radiation to pass through) filled with hydrogen gas. - When current is passed through these electrodes maintained at high voltage, discharge of electrons occurs which excites hydrogen molecules which in turn cause emission of UV radiations in near UV region. - They are stable and widely used. **3. Xenon Discharge Lamp:** - It possesses two tungsten electrodes separated by some distance. - These are enclosed in a glass tube (with quartz or fused silica) and xenon gas is filled under pressure. - An intense arc is formed between electrodes by applying high voltage. This is a good source of continuous plus additional intense radiation. Its intensity is higher than the hydrogen discharge lamp. **Demerit:** - The lamp since operates at high voltage becomes very hot during operation and hence needs thermal insulation. **4. Mercury arc Lamp:** - In Mercury arc lamp, mercury vapor is stored under pressure and excitation of mercury atoms is done by electric discharge. **Demerit:** - Not suitable for continuous spectral studies, (because it doesn't give continuous radiations). ## Collimating System - The radiation emitted by the source is collimated (made parallel) using lenses, mirrors and slits. - **Lenses:** - Materials used for the lenses must be transparent to the radiation being used. - Ordinary silicate glass transmits between 350 to 3000nm and is suitable for visible and near IR region. - Quartz or fused silica is used as a material for lenses to work below 300nm. - **Mirrors:** - These are used to reflect, focus or collimate light beams in spectrophotometer. - To minimize the light loss, mirrors are aluminized on their front surfaces. - **Slits:** - Slit is an important device in resolving polychromatic radiation into monochromatic radiation. - To achieve this, entrance slit and exit slit are used. - The width of slit plays an important role in resolution of polychromatic radiation. ## Monochromaters - This device is used to isolate the radiation of the desired wavelength from the wavelength of the continuous spectra. - **The essential elements of monochromaters:** - An entrance slit - Dispersing element - Exit slit - The entrance slit sharply defines the incoming heterochromatic radiation. - The dispersing element disperses heterochromatic radiation into its component wavelength. - The exit slit allows the nominal wavelength together with a band of wavelength on either side of it. - **Following types of monochromatic devices are used:** - Filters - Prisms - Gratings - **Filters:** - Two types of filters are used: - **Absorption filters:** works by selective absorption of unwanted radiation and transmits the radiation which is required. Examples: Glass and Gelatin filters. - **Interference filter:** works on the interference phenomenon, causes rejection of unwanted wavelength by selective reflection. It is constructed by using two parallel glass plates, which are silvered internally and separated by thin film of dielectric material of different (CaF2, SiO, MgF2) refractive index. It has a band pass of 10-15nm with peak transmittance of 40-60%. - **Advantages of Interference filters:** - Provide greater transmittance and narrower band pass (10-15nm) as compare to absorption filter. - Inexpensive - Additional filters can be used to cut off undesired wavelength. - **Advantages of Absorption filters:** - Simple in construction - Cheaper - Selection of the filter is easy - Less accurate - Band pass (bandwidth) is more (±20-30nm) i.e. if we have to measure at 400nm; we get radiation from 370-430nm. Hence less accurate results are obtained. - **Prism:** - made from glass, Quartz or fused silica. - Quartz or fused silica is the choice of material of UV spectrum. - When white light is passed through glass prism, dispersion of polychromatic light in rainbow occurs. - By rotation of the prism, different wavelengths of the spectrum can be made to pass through in exit slit on the sample. - The effective wavelength depends on the dispersive power of prism material and the optical angle of the prism. - **Gratings:** - Most efficient in converting a polychromatic to monochromatic light. - A resolution of +/- 0.1nm could be achieved by using gratings. - Gratings are expensive but commonly used in spectrophotometers. - Two types: Diffraction grating and Transmission gratings. - **Diffraction grating:** - More refined dispersion of light is obtained by means of diffraction gratings. These consist of large number of parallel lines (grooves) about 15000-30000/ inch is ruled on highly polished surface of aluminum. To make the surface reflective, a deposit of aluminum is made on the surface. Diffraction produces reinforcement. - **Transmission grating:** - It is similar to diffraction grating but refraction takes place instead of reflection. Refraction produces reinforcement, this occurs when radiation transmitted through grating reinforces with the partially refracted radiation. - **Advantages of Gratings:** - Grating gives higher and linear dispersions compared to prism monochromaters. - Can be used over wide wavelength ranges. - Gratings can be constructed with materials like aluminium which is resistant to atmospheric moisture. - Provide light of narrow wavelength. - No loss of energy due to absorption. ## Sample Holder or Cuvettes - The cells or cuvettes are used for handling liquid samples. - The cell may either be rectangular or cylindrical in nature. - For study in UV region; the cells are prepared from quartz or fused silica where as fused glass is used for visible region. - The surfaces of absorption cells must be kept clean. No fingerprints should be present on cells. - Cleaning is carried out washing with distilled water or with dilute alcohol, acetone. - The cell or cuvette that contain samples for analysis should fulfil 3 conditions: - They must be uniform in construction, the thickness must be constant and surfaces facing the incident light must be optically flat. - The materials of construction should be inert to solvents. - They must transmit light of the wavelength used.