Mansha Analyt Techniq Mod

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Analytical TechniquesDr. Muhamma Mansha

Spectroscopy

Spectroscopy is the use ofthe absorption, emission, or scattering of electromagnetic

radiation by matterto qualitatively or quantitatively analyze or study the matter or to

study physical processes. The matter can be atoms, molecules, atomic or molecular

ions, or solids. The interaction of radiation with matter can cause redirection of the

radiation and/or transitions between the energy levels of the atoms or molecules.

Beer-Lambert Law: Beer-Lambert law or simply Beers law is the heartof spectrometry. Consider the absorption of monochromatic radiation as in

Figure. Incident radiation of radiant power P0 passes through a solution of an

absorbing species a concentration c and path length b, and the transmitted

radiation has radiant power P.

Transmittance (T), is defined as:

A new term absorbance is defined as :

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The figure shows the case of absorption of light through a sample cell and includes other processes

that decreases the transmittance such as surface reflectance and scattering. For a monochromaticlight absorbance is proportional to path length b, and concentration of absorbing species c.

Instrumentation

General aspects of UV-Vis spectrophotometers are given in the introductory document on UV-Vis

spectroscopy. Single-beam spectrophotometers can utilize a fixed wavelength light source or a

continuous source. The simplest instruments use a single-wavelength light source, such as a light-emitting diode (LED), a sample container, and a photodiode detector. Instruments with a

continuous source have a dispersing element and aperture or slit to select a single wavelength

before the light passes through the sample cell. (see schematic below).

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In either type of single-beam instrument, the instrument is calibrated with a reference cell

containing only solvent to determine the Po value necessary for an absorbance measurement.

Schematic of a wavelength-selectable, single-beam UV-Vis spectrophotometer

Ultraviolet/Visible (UV-Vis) Spectroscopy of Potassium Permanganate

Potassium permanganate is used to kill bacteria in reclaimed water

Use UV-Vis to ensure that the concentration of Potassium Permanganate is at acceptable

limit Potassium permanganate (KMnO4) in solution is purple / violet color meaning maximum

absorption should be at 500 550 nm, which is confirmed from the absorption spectrum ofpotassium permanganate at two different concentrations shown in the figure.

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Absorption Spectrum of Light4Wavelength of

maximumabsorption (nm)

Color Absorbed Color Observed

(complementry)

380 420 Violet Green-Yellow

420 - 440 Violet-Blue Yellow

440 470 Blue Orange

470 500 Blue-Green Red

500 520 Green Purple

520 550 Yellow-Green Violet

550 580 Yellow Violet-Blue

580 620 Orange Blue

620 680 Red Blue-Green

680 - 780 Purple Green

Five known concentrations of KMnO4: 1ppm, 20ppm, 40ppm, 60ppm, 80ppm were

prepared and absorbance readings were taken at 520 nm.

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The absorbance readings are shown in the table.

UV-Vis Absorbance Readings for PotassiumPermanganate at 520 nm

Average A (after

3 runs)

Standard

Deviation (A)

1 ppm 0.015 0.004

20 ppm 0.256 0.001

40 ppm 0.520 0.004

60 ppm 0.753 0.002

80 ppm 1.046 0.001

Unknown #4 0.462 0.001

The calibration curve is in accordance with Beers law from which the concentration of the

unknown solution is determined.

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Atomic Absorption Spectrometry (AAS)

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Potentiometry

Introduction

Potentiometry is the field ofelectroanalytical chemistry in which potential is measured under theconditions of no current flow. The measured potential may then be used to determine the analytical

quantity of interest, generally the concentration of some component of the analyte solution. The

potential that develops in the electrochemical cellis the result of thefree energy change that would

occur if the chemical phenomena were to proceed until the equilibrium condition has beensatisfied.

This concept is typically introduced in quantitative analysis courses in relation to electrochemical

cells that contain an anode and a cathode. For these electrochemical cells, thepotential difference

between the cathode electrode potentialand the anode electrode potential is the potential of theelectrochemical cell.

If the reaction is conducted understandard state conditions, this equation allows the calculation of

the standard cell potential. When the reaction conditions are not standard state, however, one mustutilize theNernst equation to determine the cell potential. In the following equations Ox stands for

oxidized species and Red for its reduced form. For example Cu2+ is oxidized form and Cu is

reduced form.

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pH Meters

pH meter measures the pH of a solution using an ion-selective electrode (ISE) that responds to the

H+ concentration of the solution. The pH electrode produces a voltage that is a function of theconcentration of the H+ concentration, and making measurements with a pH meter is therefore a

form ofpotentiometry.

The pH electrode is attached to electronics which convert the voltage to a pH reading and displays

it on a meter.

The difference in concentration of hydrogen ions on both sides of membrane causes the potential

to develop. There are no half reactions and the Nernst equation can be written as:

E = Eo 0.0592 log {[H+]internal / [H+]external }

Since the internal [H+] is constant, it can be lumped into Eo

E = E* + 0.0592 log [H+]external

E = E* 0.0592 p[H+]

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Instrumentation

A pH meter consists of a H+-selective membrane, an internal reference electrode, an external

reference electrode, and a meter with control electronics and display. Commercial pH electrodes

usually combine both electrodes into one unit that is then attached to the pH meter. The potentialscale is calibrated in pH units with each pH unit equal to 59.19 mV at 25 oC.The pH meter is

adjusted with the calibration knob with the help of a standard buffer.

Picture of a pH meter

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Separation Techniques:Chromatography is a separations method that relies on differences in partitioning behaviorbetween a flowing mobile phase and a stationary phase to separate the the components in a

mixture.

A column (or other support like paper) holds the stationary phase and the mobile phase carries thesample through it. Sample components that partition strongly into the stationary phase spend agreater amount of time in the column and are separated from components that stay predominantly

in the mobile phase and pass through the column faster.

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As the components elute from the column they can be quantified by a detector and/or collected for

further analysis. An analytical instrument can be combined with a separation method for on-lineanalysis. Examples of such "hyphenated techniques" include gas and liquid chromatography with

mass spectrometry (GC-MS and LC-MS).

GC: It is applied to volatile organic compounds. The mobile phase is a gas andthe stationary phase is usually a liquid on a solid support or sometimes a solidadsorbent.

Distribution of analytes between phases

http://teaching.shu.ac.uk/hwb/chemistry/tutorials/chrom/chrom1.htmThe distribution of analytes between phases can often be described quite simply. Ananalyte is in equilibrium between the two phases;

Amobile Astationary

The equilibrium constant, K, is termed thepartition coefficient; defined as themolar concentration of analyte in the stationary phase divided by the molarconcentration of the analyte in the mobile phase.

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K = [A]s/[A]M = cS/cMWhere the bracketed terms are activities of solute A in the two phases. Weshall substitute c to represent molar analytical concentration. Ideally the cs isdirectly proportional to cM over a wide concentration range of solute.

The time between sample injection and an analyte peak reaching a detector at the end of the

column is termed the retention time (tR). Each analyte in a sample will have a different retentiontime. The time taken for the mobile phase to pass through the column is called tM (In the diagram

19.3 it is shown as unretained peak.

A term called the retention factor, k', (also known as capacity factor in old literature) is often used

to describe the migration rate of an analyte through a column. The retention factor for analyte A isdefined as;

k'A = (tR - tM )/ tM

tR and tM are easily obtained from a chromatogram. When an analytes retention factor is less thanone (note that it is never negative) elution is so fast that accurate determination of the retention

time is very difficult. High retention factors (greater than 20) mean that elution takes a very long

time. Ideally, the retention factor for an analyte is between one and five.

We define a quantity called theselectivity factor, , which describes the separation of two species(A and B) on the column;

= k 'B / k 'A

When calculating the selectivity factor, species A elutes faster than species B. The selectivity

factor is always greater than unity. The selectivity factor for two analytes in a column provides a

measure of how well the column will separate the two.

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`The Theoretical Plate Model of ChromatographyThe plate model supposes that the chromatographic column is contains a largenumber of separate layers, called theoretical plates. Separate equilibrations of thesample between the stationary and mobile phase occur in these "plates". The analytemoves down the column by transfer of equilibrated mobile phase from one plate to

the next.

It is important to remember that the plates do not really exist; they areimaginary. The concept helps us understand the processes at work in thecolumn.They also serve as a way of measuring column efficiency, either by statingthe number of theoretical plates in a column, N (the more plates the better), or by

stating the plate height; the Height Equivalent to a Theoretical Plate (the smaller thebetter).If the length of the column is L, then the HETP is

HETP = L / N

The number of theoretical plates that a real column possesses can be found by examining a

chromatographic peak after elution;

where w1/2 is the peak width at half-height.However when peak width is measured at base line, the formula becomes:

As can be seen from this equation, columns behave as if they have different numbersof plates for different solutes in a mixture.

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Liquid Chromatography (LC): It is used to separate analytes in solutionincluding metal ions. The mobile phase is a solvent and the stationary phase isa liquid on a solid support, a solid, or an ion-exchange resin.

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