Absorption Spectroscopy PDF

Terms employed in absorption spectroscopy : When light interacts with matter it can do one of several things, depending on its wavelength and what kind of matter it encounters: it can be transmitted, reflected, refracted, diffracted, adsorbed or scattered.  I◦ = It + Ia + Ir + If + Is...

Terms employed in absorption spectroscopy : When light interacts with matter it can do one of several things, depending on its wavelength and what kind of matter it encounters: it can be transmitted, reflected, refracted, diffracted, adsorbed or scattered.  I◦ = It + Ia + Ir + If + Is It= transmitted light Ia=absorbed light Ir=reflected light If= refracted light Is= scattered light For clear solutions, Is=0 If& Ir can be cancelled by using blank solution Many compounds absorb ultraviolet (UV: 200nm - 400nm) or visible (Vis.: 400 nm - 800 nm) light. The diagram below shows a beam of monochromatic radiation of radiant power I0, directed at a sample solution. Absorption takes place and the beam of radiation leaving the sample has radiant power I. The amount of radiation absorbed may be measured in a number of ways: Transmittance Transmittance : T = I / I0 % Transmittance: %T = 100 T Absorbance: A = log10 (1 / T) = -log10 (T) A = log10 (100 / %T) A = 2 - log10 (%T) The absorption of a solution increases as the transmittance decreases  The relationship between absorbance and transmittance is illustrated in the following diagram: So, if all the light passes through a solution without any absorption, then absorbance is zero, and percent transmittance is 100%. If all the light is absorbed, then percent transmittance is zero, and absorption is infinite. Link the structures to the UV spectra A–D. Answer: A diphenhydramine, B sulfurazole C atenolol D propranolol.  Lambert's Law: When a beam of monochromatic radiation enters an absorbing medium; its intensity decreases exponentially with the increase of thickness [b] of the medium traversed mathematically log I0/It=Kb concentration C held constant K is the proportionality constant , I0 and It are intensity of incident and transmitted radiations. Beer's Law: It relates absorption capacity to the concentration of an absorbing solute. It stated that absorption is proportional to the number of absorbing molecules in the light path log lo/l=K C where K is proportionality constant and C is the concentration while b is held constant. Beer's-Lambert's law: This is a combination of both laws; Log lo/l= abc where (a) is a constant called absorptivity, (b) is the pathlength in cm and (c) is the concentration in grams/Liter. Log lo/l is usually substituted by A (Absorbance), then the equation become; A = abc The value of (a) will clearly depend upon the method of expression of the concentration. If (c) is expressed in moles/liter, and b in centimetres then a is given the symbol ε Epsilon ( l.mol-1cm-1) and is called the molar absorption coefficient or molar absoptivity A=εbc when ε is measured at λmax it is called εmax when (c) is in g/100 ml and b in centimetres. We get A=A(1%,1cm)cb The A1%1cm is valuable for natural products identification when their constituent is unknown (molecular weight is not definitely known). A1%1cm can be converted easily to ε by the equation; Deviations from Beer-Lambert Law: When the results obey Beer-Lambert law, the concentration versus absorbance gives a straight line passing through the origin (calibration curve) as indicated by the solid line in figure below. In some cases, deviation from Beer-Lambert law occur which may be: (1) Real deviation: At high concentrations, due to crowding, molecular interaction and association as well as charge distribution. (2) Instrumental deviation (errors): a) Irregular deviation due to: Unmatched cells, unclean handling and unclean optics. b) Regular deviation: due to: - Errors in wavelength scale. - Slit width control. - Stray light is any radiation of wavelength other than those which are absorbed. Also includes any light reaches the detector without passing through the sample.. c) Other errors: Non-linear response of photo cells. Radio and TV interferences. Unstabilized power supply. (3) Chemical deviations: pH effects. Solvents interaction due to high concentrations. Temperature effects. Dipole interactions Time factor affect oxidation, reduction, or hydrolysis reactions which may be occur.  Colorimetr y The energy of transition determines λ of absorption and if the λmax is in the visible range (400-800 nm), a colour is obtained and its measurement can be made visually by using simple apparatus. Thus colorimetry is the technique of measuring visible radiation. colorimetry is the method for quantitative analysis which depends on measuring the absorption of VIS radiation  It must be noted that; the absorbed colour is different from the actually observed color. The observed colour is complementary color which remained after the wave length absorbed from white light (see the following table and colour wheel). chromogen( reagent): is substance capable of conversion into a pigment or dye.(lacks definite color but may be transformed into a pigment) General Requirement for an ldeal chromogen: It should be Colorless Its excess easily separated from the color product. Has no absorbance at λmax of the coloured product Selective (react with analyte only) Reacts quantitatively with the analyzed substance (stoichiometry) Give one single colored product with specified λmax. Sensitive & gives highly color product. Color development must be rapid Note: The range from 200 - 400 nm can not be detected by the eye and must be measured spectrophotometrically (using spectrophotometers). Advantages of colorimetry: 1- Give more accurate results at low concentrations than the corresponding titrimetric and gravimetric procedures. 2- It is frequently applied under conditions where no satisfactory gravimetric or titrimetric procedure exists, e.g. for biological substances. 3- It is considered a simple and rapid techniqe if compared with titrimetry and gravimetry. Quantitative Measurements of EMR Absorption Two Techniques that can be used: Visual Methods Instrumental methods (A) Standard series method (B)Balancing (Varying depth) method  [I] Visual Methods Are used for measuring the coloured solutions only (A) Standard series method The test solution contained in a test tube is matched with a series of standards similarly prepared, then the concentration of unknown solution is equal to that of the standard solution whose colour is matched with it exactly. Example, is the determination of copper with ammonia by forming blue colour or determination of iron (III) by forming blood red colour with thiocyanate anion.