Prepared by:Karwan O. Ali

Yousif T. Maaroof

Mzgin. M. Ayoob

UV- VISIBLE SPECTROSCOPY

What is spectroscopy?

The interactions of radiation and matter are the subject of the

science called spectroscopy. Spectroscopic analytical methods

are based on measuring the amount of radiation produced or

absorbed by molecular or atomic species of interest

The Electromagnetic Spectrum

Light exhibits wave property during its propagation and energy particle

during its interaction with matter. The double nature of light (waves and

particles) is known as dualism.

Dual nature of light

• Light consist of energy packets, known as photons.

• The energy (E) of photons is proportional to the frequency i.e.

related to c and . It can be expressed by max plank relation:

• E = h ( = C /)

• where h = max plank constant = 6.63 x 10-27 erg., sec.)

• i.e. E or E 1/

For analytical purposes we use the region of I.R, visible and U.V

radiations.

UV radiation region is classified into : far UV from (10nm-200nm) and

near UV from (200nm-380nm

Why Is a Red Solution Red?

• When a molecule interact with radiant energy, the molecule is said to

be excited , because the outer valence electrons undergo transition

from original energy level ground state (E g) to an exited state (E s).

Interaction of a substance with EMR

Excited state Es

Ground state E g

When a molecule in the ground state absorbs

EMR, 3 energy state transitions will take place.

•These types of transition are:

1) Electronic

2) Vibrational

3) Rotational.

• When the molecule absorb Visible and U.V region.

Raising electrons to a higher energy level, raising the Vibration of

molecule, and increasing rotation of the molecule (Electronic transition

energy + Vibrational transition energy + Rotational transition energy)

1) Electronic

• When the molecule absorb I.R region.

Raising the Vibration of molecule and increasing rotation of the

molecule (Vibrational transition energy + Rotational transition energy)

2) Vibrational

• When the molecule absorbs F.I.R and Microwave regions.

Increasing rotation of the molecule (Rotational transition

energy).

3) Rotational

• Absorption measurements based upon ultraviolet and visible radiation find widespread

application for the quantitative determination of a large variety species [1].

• Beer’s Law:

• A = -logT = logP0/P = abc = bc = 2 - log%T

• Where T = transmittance, T = percentage transmittance, P = transmitted power of

radiation, Po = incident power of radiation, A = absorbance, a = absorptivity,

b = path length, c = concentration, = molar absorptivity, extinction coefficient

An Introduction to Ultraviolet/Visible

Molecular Absorption Spectrometry

The Quantitative Picture

Where the absorbance A has no units, since A = log10 P0 / P

Is the molar absorbtivity with units of L mol-1 cm-1

b is the path length of the sample in cm

c is the concentration of the compound in solution, expressed in mol L-1 (or M,

molarity)

The Beer-Lambert Law (Beer’s Law): A = b c

• Monochromatic incident radiation (all molecules absorb light of one )

• Absorbers independent (Absorbing molecules act independently of one another i.e., low c)

• Path length is uniform (all rays travel the same distance in sample)

• No scattering

• Absorbing medium is optically homogeneous

• Incident beam is not large enough to cause saturation

• All rays should be parallel to each other and perpendicular to surface of medium

Assumptions in derivation of Beer’s Law

Deviations from Beer’s Law• Real Limitations

• Beer’s law is successful in describing the absorption behavior of dilute solutions only ; in this sense it is a

limiting law. At high concentrations ( > 0.01M ),the average distance between the species responsible for

absorption is diminished to the point where each affects the charge distribution of its neighbors

This interaction, in turn, can alter the species’ ability to absorb at a given wavelength of radiation thus leading

to a deviation from Beer’s law [2].

A similar effect is sometimes encountered in solutions containing low absorber concentrations and high

concentrations of other species, particularly electrolytes.

Beer’s law is valid at low concentrations,

but breaks down at higher concentrations

For linearity, A < 1

Deviations from Beer's law also arise because ε is dependent upon the

refractive index of the solution. Thus, if concentration changes cause

significant alterations in the refractive index η of a solution, departures from

Beer's law are observed. A correction for this effect can be made by

substitution of the quantity εη /(η2 + 2)2 for ε in the Beer equation, as shown

below [2]:

A= [εηb /(η2 + 2)2 ] bC

In general, this correction is never very large and is rarely significant at

concentrations less than 0.01 M.

• Chemical deviations from Beer’s law are caused by shifts in the position of a chemical or physical

equilibrium involving the absorbing species. A common example of this behavior is found with

acid/base indicators. Deviations arising from chemical factors can only be observed when

concentrations are changed

Chemical Deviations

HIn Ka H+ + In-

Red, =600nm ColorlessPhenolphthalein:

If solution is buffered, then pH is constant and [HIn]

is related to absorbance.

But, if un buffered solution,

equilibrium will shift depending on

total analyte concentration

CHIn

Expected

Actual

• instrument may be caused by fluctuations in the power-supply voltage, an

unstable light source, or a non-linear response of the detector-amplifier system

• Polychromatic Radiation

• All monochromators, regardless of quality and size, have a finite resolving

power and therefore minimum instrumental bandwidth. Polychromatic

radiation (i.e., light of more than one )

Instrumental Factors

Band A shows little deviation, because ε does not change greatly throughout the band. Band

B shows marked deviations because ε undergoes significant changes in this region

The width of the image produced is thus an important measure of the quality of the

performance of a spectrometer. The figure below shows the loss of detail that accompanies the

use of wider slits. It is evident that an increase in slit width brings about a loss of spectral

detail

Effect of Slit Width on Absorbance Measurements

• quantitative measurement of narrow absorption bands demand the use of narrow slits widths. Unfortunately, a

decrease in slit width is accompanied by a second-order power reduction in the radiant energy; at very narrow settings

spectral detail may be lost owing to an increase in the signal-to-noise ratio. In general, it is good practice to narrow

slits no more than is necessary for good resolution for the spectrum at hand

Another effect of slit width is the change of absorbance values that accompany a change in the slit width. The

figure below illustrates this effect. Note that the peak absorbance values increase significantly (by as much as

70% in one instance) as the slit width decreases

The basic components of analytical instruments for absorption spectroscopy,

as well as for emission and fluorescence spectroscopy, are remarkably alike

in function and in general performance requirements whether the

instruments are designed for ultraviolet (UV), visible, or infrared (lR)

radiation. We often call the UV/visible and IR regions of the spectrum the

optical region

Instruments for Optical Spectrometry

1) a stable source of radiant energy

2) a wavelength selector that isolates a limited region of the spectrum for measurement

3) one or more sample containers

4) a radiation detector, which converts radiant energy to a measurable electrical signal

5) a signal processing and readout unit, usually consisting of electronic hardware and, in modern instruments, a computer.

INSTRUMENT COMPONENTS

To be suitable for spectroscopic studies, a source must generate a beam of radiation that is sufficiently powerful

to allow easy detection and measurement.

In addition, its output power should be stable for reasonable periods of time. Typically, for good stability, a well-

regulated power supply must provide electrical power for the source

Spectroscopic Sources

Spectroscopic sources are of two types:

A- Continuum sources, which emit radiation that changes in intensity only

slowly as a function of wavelength.

B- Line sources, which emit a limited number of spectral lines, each of

which spans a very limited wavelength range.

The distinction between these sources is illustrated in Figure bellow. Sources

can also be classified as

continuous sources, which emit radiation continuously with time, or pulsed

sources, which emit radiation in bursts

tungsten lamp of the type used in spectroscopy and its spectrum (b). Intensity of the

tungsten source is usually quite low at wavelengths shorter than about 350 nm. Note that

the intensity reaches a maximum in the near-IR region of the spectrum (<1200 nm in this

case).

Tungsten lamp

• Tungsten/halogen lamp

• Tungsten/halogen lamps, also called quartz/halogen lamps, contain a small amount of iodine

within the quartz envelope that houses the filament. Quartz allows the filament to be operated

at a temperature of about 3500 K, leading to higher intensities and extending the range of the

lamp well into the UV. The lifetime of a tungsten/halogen lamp is more than double that of an

ordinary tungsten lamp, which is limited by sublimation of tungsten from the filament. In the

presence of iodine, the sublimed tungsten reacts to give gaseous WI2 molecules. These

molecules then diffuse back to the hot filament where they decompose, redeposit W atoms on

the filament, and release iodine. Tungsten/halogen lamps are finding ever-increasing use in

spectroscopic instruments because of their extended wavelength range, greater intensity, and

longer life

A deuterium lamp consists of a cylindrical tube containing deuterium at

low pressure, with a quartz window from which the radiation exits

Deuterium lamp

The word laser originally was the upper-case LASER, the acronym from Light

Amplification by Stimulated Emission of Radiation. Lasers have become useful

as sources in certain types of analytical spectroscopy. To understand how a laser

works, consider an assembly of atoms or molecules interacting with an

electromagnetic wave. Laser radiation is highly directional. Spectrally pure,

coherent. and of high intensity. These properties have made possible many

unique research applications that cannot easily be achieved with conventional

sources

Laser Sources

Spectroscopic instruments in the UV and visible regions are usually equipped with one or more

devices to restrict the radiation being measured to a narrow band that is absorbed or emitted by

the analyte. Such devices greatly enhance both the selectivity and the sensitivity of an

instrument. In addition, for absorption measurements, narrow bands of radiation greatly

diminish the chance of Beer's Jaw deviations due to polychromatic radiation. Many

instruments use a monochromator or filter to isolate the desired wavelength band so that only

the band of interest is detected and measured. Others use a spectrograph to spread out, or

disperse, the wavelength so that they can be detected with a multichannel detector

Wavelength Selectors

Monochromators generally have a diffraction grating to disperse the radiation into its component wavelengths, as shown in Figure bellow

By rotating the grating, different

Wavelengths can be made to pass

through an exit slit

Monochromators consist of:

1- entrance slit

2- collimating mirror or lens

3- a prism or grating

5- focal plane

6- exit slit

Older instruments used prisms for this purpose

Monochromators

Older instruments used prisms for this purpose

Radiation from a source enters the monochromator

via a narrow rectangular opening, or slit.

The radiation is then collimated by a concave

mirror, which produces a parallel beam that

strikes the surface of a reflection grating.

Angular dispersion results from diffraction,

which occurs at the reflective surface

Grating Monochromator

The effective band width of the monochromator depends on the size and quality of the

dispersing element, the slit widths, and the focal length of the monochromator. A high-quality

rnonochromator will exhibit an effective band-width of a few tenths of a nanometer or less in

the ultraviolet/visible region. The effective bandwidth of a monochromator that is satisfactory

for most quantitative applications is about 1 to 20 nm. Many monochromators are equipped

with adjustable slits to permit some control over the bandwidth. A narrow slit decreases the

effective bandwidth but also diminishes the power of the emergent beam. Thus, the minimum

practical bandwidth may be limited by the sensitivity of the detector For qualitative analysis,

narrow slits and minimum effective bandwidths are required if a spectrum is made up of

narrow peaks. For quantitative work. however, wider slits permit operation of the detector

system at lower amplification, which in turn provides greater reproducibility of response.

Most gratings in modern monochromators are replica gratings, which are obtained by

making castings of a master grating. The latter consists of a hard, optically flat, polished

surface on which a suitably shaped diamond tool has created a large number of parallel and

closely spaced grooves. A grating for the ultraviolet and visible region will typically contain

300 to 2000 grooves/mm. with 1200 to 1400 being most common. The construction of a

good master grating is tedious, time consuming, and expensive because the grooves must

be identical in size, exactly parallel, and equally spaced over the length of the grating (3 to

10 cm). Replica gratings are formed from a master grating by a liquid resin casting process

that preserves virtually perfectly the optical accuracy of the original master grating on a clear

resin surface. This surface is ordinarily made reflective by a coating of aluminum or, some

times, gold or platinum

Radiation Filters

Filters operate by absorbing all but a restricted band of radiation from a

continuum source. As shown in Figure bellow, two types of filters are used in

spectroscopy; interference filters and absorption filters. Interference filters are

typically used for absorption measurements, and they generally transmit a much

greater fraction of radiation at their nominal wavelengths than do absorption

filters

Interference Filters

Interference filters are used with ultraviolet and visible radiation, as well as with

wavelengths up to about 14 µm in the infrared region. As the name implies, an

interference filter relies on optical interference to provide a relatively narrow

band of radiation. typically 5 to 20 nm in width. As shown in Figure bellow, an

interference filter consists of a very thin layer of a transparent dielectric material

(frequently calcium fluoride or magnesium fluoride) coated on both sides with a

film of metal that is thin enough to transmit approximately half the radiation

striking it and to reflect the other half.

• the radiant power transmitted, fluoresced, or emitted must be detected in some

manner and converted into a measurable quantity. A detector is a device that

indicates the existence of some physical phenomenon. Familiar examples of

detectors include photographic film (for indicating the presence of

electromagnetic or radioactive radiation.

• The human eye is also a detector; it converts visible radiation into an electrical

signal that is passed to the brain via a chain of neurons in the optic nerve and

produces vision.

Detecting and Measuring Radiant Energy

• The term transducer is used to indicate the type of detector that

converts quantities, such as light intensity, pH, mass, and temperature,

into electrical signals that can be subsequently amplified, manipulated,

and finally converted into numbers proportional to the magnitude of

the original quantity.

• A transducer is a type of detector that converts various types of

chemical and physical quantities into electrical signals such as

electrical charge, current, or voltage.

There are two general types of transducers:

• Photons

All photon detectors are based on the interaction of radiation with a reactive

surface either to produce electrons (photoemission) or to promote electrons to

energy states in which they can conduct electricity (photoconduction). Only

UV, visible, and near-IR radiation possess enough energy to cause

photoemission to occur; thus, photoemissive detectors are limited to

wavelengths shorter than about 2 µm (2000 nm).

Types of Transducers

• Thermal detectors (Heat)

detect a temperature change in a material due to photon absorption

• Thermal detectors can be used over a wide range of wavelengths

.Their main disadvantages are slow response time and lower

sensivity relative to other types of detectors.

Widely used types of photon detectors include phototubes, photomultiplier

tubes, silicon photodiodes, and photodiode arrays.

Photon Detectors

Phototubes

The response of a phototube or a photomultiplier tube is based on the photoelectric effect. a

phototube consists of a semicylindrical photocathode and a wire anode sealed inside an

evacuated transparent glass or quartz envelope. The concave surface of the cathode supports

a layer of photoemissive material. such as an alkali metal or a metal oxide. that emits

electrons when irradiated with light of the appropriate energy. When a voltage is applied

across the electrodes, the emitted photoelectrons are attracted to the positively

charged wire anode, In the complete circuit, a photocurrent then results that is easily

amplified and measured. The number of photoelectrons ejected from the photocathode per

unit time is directly proportional to the radiant power of the beam striking the surface. With

an applied voltage of about 90 V or more, all these photoelectrons are collected at the anode

to give a photocurrent that is also proportional to the radiant power of the beam.

Photoelectrons: are electrons that are ejected

from a photosensitive surface by electromagnetic

radiation.

Photocurrent: is the current in an external

circuit that is limited by the rate of ejection of

photoelectrons.

The photomultiplier tube (PMT) is similar in construction to the phototube but is

significantly more sensitive. Its photocathode is similar to that of the phototube, with electrons

being emitted on exposure to radiation. In place of a single wire anode. however, the PMT has a

series of electrodes called dynodes, The electrons emitted from the cathode are accelerated

toward the first dynode. which is maintained 90 to 100 V positive with respect to the cathode.

Each accelerated photoelectron that strikes the dynode surface produces several electrons, called

secondary electrons, that are then accelerated to dynode 2, which is held 90 to 100 V more

positive than dynode l. Again, electron amplification results. By the time this process has been

repeated at each of the dynodes, 105 to 107 electrons have been produced for each incident

photon. This cascade of electrons is finally collected at the anode to provide an average current

that is further amplified electronically and measured.

One of the major advantages of photomultipliers is their automatic internal amplification.

About 106 to 107 electrons are produced at the anode for each photon that strikes the

photocathode of a photomultiplier tube. Photomultiplier tubes are among the most widely

used types of transducers for detecting ultraviolet/visible radiation. With modern electronic

instrumentation, it is possible to detect the electron pulses resulting from the arrival of

individual photons at the photocathode of a PMT. The pulses are counted, and the

accumulated count is a measure of the intensity of the electromagnetic radiation impinging on

the PMT. Photon counting is advantageous when light intensity, or the frequency of arrival of

photons at t\1e photocathode, is low.

Photoconductive transducers consist of a thin film of a semiconductor

material, such as lead sulfide, mercury cadmium telluride (MCT), or

indium antimonide, deposited often on a nonconducting glass surface

and sealed in an evacuated envelope. Absorption of radiation by these

materials promotes nonconducting valence electrons to a higher energy

state, which decreases the electrical resistance of the semiconductor.

Typically, a photoconductor is placed in series with a voltage source and

a load resistor, and the voltage drop across the load resistor serves as a

measure of the radiant power of the beam of radiation. The PbS and InSb

detectors are quite popular in the near-IR region of the spectrum. The

MCT detector is useful in the mid- and far-IR regions when cooled with

liquid N2 to minimize thermal noise.

Photoconductive Cells:

Diode-Array Detectors

Silicon photodiodes have become important recently because 1000 or more can

be fabricated side by side on a single small silicon chip. (The width of individual

diodes is about 0.02 mm). With one or two of the diode-array detectors placed

along the length of the focal plane of a monochromator. All wavelengths can be

monitored simultaneously, thus making high-speed spectroscopy possible. Silicon

photodiode detectors respond extremely rapidly, usually in nanoseconds.

They are more sensitive than a vacuum phototube but considerably less

sensitive than a photomultiplier tube.

Sample containers, which are usually called cells or cuvettes must have windows that are

transparent in the spectral region .Thus as shown in the figure below, quartz or fused silica is

required for the UV region (wavelengths less than 350nm nm) and may be used in the visible

region and out to about 3000 nm (3 µm) in the IR region. Silicate glass is ordinarily used for

the 375 to 2000 nm region because of its low cost compared with quartz. Plastic cells are also

used in the visible region. The most common window material for IR studies is crystalline

sodium chloride, which is soluble in water and in some other solvents.

Sample Containers

The best cells have windows that

are perpendicular to the direction

of the beam in order to minimize

reflection losses. The most

common cell path length for

studies in the UV and visible

regions is 1 cm; matched,

calibrated cells of this size are

available from several

commercial sources. Many other

cells with shorter and longer path

lengths can be purchased.

For reasons of economy, cylindrical cells are sometimes used. Fingerprints, grease, or other

deposits on the walls markedly alter the transmission characteristics of a cell. Thus,

thorough cleaning before and after use is need , and care must be taken to avoid touching

the windows after cleaning is complete. Matched cells should never be dried by heating in

an oven or over a flame because this may cause physical damage or a change in path

length..

Photometers provide simple, relatively inexpensive tools for performing

absorption measurements. Filter photometers are often more convenient and

more rugged and are easier to maintain and use than the more sophisticated

spectrophotometers. Furthermore, photometers characteristically have high

radiant energy throughputs and thus good signal-to-noise ratios even with

relatively simple and inexpensive transducers and circuitry. Photometers have

the advantages of simplicity, ruggedness, and low cost.

Spectrophotometers

offer the considerable advantage that the wavelength can be varied

continuously, thus making it possible to record absorption spectra and is a

scanning instrument (i.e. a spectrophotometer has a monochromator for

separating the individual wavelengths of light). Several dozen models of

spectrophotometers are available commercially. Most spectrophotometers

cover the UV /visible and occasionally the near-infrared region, while

photometers are most often used for the visible region.. Both photometers and

spectrophotometers can be obtained in single- and double-beam varieties.

Single beam spectrometers

Single beam spectrometers are relatively cheap, simple, portable & ideally

suited to quantitative analysis .It is not possible to scan through the entire

spectrum with such an instrument because both the source intensity & the

detector response vary with the wavelength. To record an accurate value of

the absorbance it is necessary to zero the instrument on a reference/blank

before every measurement. Thus, this is essentially a single wavelength

measurement of absorbance.

Voltage fluctuations and changes in light source present a problem

When a heavy load is placed on the electric power system, lights dim and later brighten

*If measurements are being taken on the spectrophotometer at the same time, thereadings will be unreliable

*Aging lamp source may momentarily flicker and cause the readings to be unstable anderrorneous

So, a single-beam instrument requires a stabilized voltage supply to avoiderrors resulting from changes in the beam intensity during the time required tomake the 100% T measurement and determine %T for the analyte.

Disadvantages of single-beam spectrophotometer

The second type of double-beam instrument. Here the beams are separated in time by a

rotating sector mirror that directs the entire beam from the monochromator first through the

reference cell and then through the sample cell. The pulses of radiation are recombined by

another sector mirror, which transmits one pulse and reflects the other to the transducer. As

shown by the insert labeled "front view", the motor-driven sector mirror is made up of pie-

shape segments, half of which are mirrored and half of which are transparent. The mirrored

sections are held in place by blackened metal frames that periodically interrupt the beam and

prevent its reaching the transducer. The double-beam-in-time approach is generally preferred

because of the difficulty in matching the two detectors needed for the double-beam-in-space

design

Double-Beam in time spectrophotometers

Double-Beam in space spectrophotometers

• Many modern photometers and spectrophotometers are based on a double-beam

design. A double-beam-in-space instrument in which two beams are formed in

space by a V-shape mirror called a beamsplitter. One beam passes through the

reference solution to a photodetector, and the second simultaneously traverses

the sample to a second, matched detector. The two outputs are amplified, and

their ratio (or the logarithm of their ratio) is determined electronically or by a

computer and displayed by the readout device.

The determination of an analyte’s concentration based on its absorption of ultraviolet or visible radiation is one of the most

frequently quantitative analytical methods. One reason for its popularity is that many organic and inorganic compounds

have strong absorption bands in the UV/Vis region of the electromagnetic spectrum.

There are many application

• Environmental Applications

• Clinical Applications

• Industrial Analysis

• Forensic Applications

Quantitative Applications

• Methods for the analysis of waters and wastewaters relying on the

absorption of UV/Vis radiation are among some of the most frequently

employed analytical methods.

Environmental Applications

• UV/Vis molecular absorption is one of the most commonly employed

techniques for the analysis of clinical samples, several examples of

which are listed in Table below. The analysis of clinical samples is often

complicated by the complexity of the sample matrix, which may

contribute a significant background absorption at the desired wavelength

.

Clinical Applications

The Application of UV/Vis Molecular Absorption to the

Analysis of Clinical Samples

• UV/Vis molecular absorption is used for the analysis of a diverse array of

industrial samples, including pharmaceuticals, food, paint, glass, and

metals.

• In many cases the methods are Products that have been analyzed in this

fashion include antibiotics, hormones, vitamins, and analgesics.

• One example of the use of UV absorption is in determining the purity of

aspirin tablets.

Industrial Analysis

• UV/Vis molecular absorption is routinely used in the analysis of

narcotics and for drug testing.

• One interesting forensic application is the determination of blood

alcohol using the Breathalyzer test. In this test a 52.5-mL breath

sample is bubbled through an acidified solution of K2Cr2O7. Any

ethanol present in the breath sample is oxidized by the dichromate,

producing acetic acid and Cr3+ as products.

Forensic Applications

• The energy at which the absorption occurs, as well as the intensity of the

absorption, is determined by the chemical environment of the absorbing moiety.

For example, benzene has several ultraviolet absorption bands due to p p*

transitions. The position and intensity of two of these bands, 203.5 nm (e =

7400) and 254 nm (e = 204), are very sensitive to substitution. For benzoic acid,

in which a carboxylic acid group replaces one of the aromatic hydrogens, the

two bands shift to 230 nm (e = 11,600) and 273 nm (e = 970). Several rules

have been developed to aid in correlating UV/Vis absorption bands to chemical

structure.

Qualitative Applications

1-Limited since few resolved peaks

• Unambiguous identification not usually possible.

Why we cannot use UV / visible Spectroscopy in

Qualitative Analysis ?

Loss of fine structure for

acetaldehyde when

transfer to solvent from

gas phase

Also need to consider

absorbance of solvent

2. Solvent can affect position and shape of curve .

• polar solvents broaden out peaks, eliminates fine structure.

•Loss of fine structure

for 1,2,4,5-tetrazine as

solvent polarity

increases

3.Solvent can also absorb in UV-vis spectrum.

• Solvent for the ultraviolet and visible regions

• Molecular absorption, particularly in the UV/Vis range, has been used for a variety of

different characterization studies, including determining the stoichiometry of metal–

ligand complexes and determining equilibrium constants.

• Stoichiometry of a Metal, Ligand Complex

• The stoichiometry for a metal–ligand complexation reaction of the following general

form

• can be determined by one of three methods: the method of continuous variations,

•the mole-ratio method, and the slope-ratio method

Characterization Applications

• also called Job’s method, is the most popular. In this method a series of solutions is prepared such that the total moles of metal and ligand, ntot, in each solution is the same. Thus, if (nM)i and (nL)i are, respectively, the moles of metal and ligand in the i-th solution, Then

• The relative amount of ligand and metal in each solution is expressed as the mole

• fraction of ligand, (XL)i, and the mole fraction of metal, (XM)i,

Method of continuous variations

A plot of A vs volume ratio (volume ratio =

mole fraction) gives maximum absorbance when

there is a stoichiometric amount of the two.

• A procedure for determining the stoichiometry between two reactants by

preparing solutions containing different mole ratios of two reactants . In

the mole-ratio method the moles of one reactant, usually the metal, are

held constant, while the moles of the other reactant are varied. The

absorbance is monitored at a wavelength at which the metal–ligand

complex absorbs. A plot of absorbance as a function of the ligand-to-

metal mole ratio (nL/nM) has two linear branches that intersect at a mole

ratio corresponding to the formula of the complex.

Mole-ratio method

• Figure (a)shows a mole-ratio plot for the formation of a 1:1 complex in which the absorbance is monitored at a wavelength at which only the complex absorbs.

• Figure (b) shows a mole-ratio plot for a 1:2 complex in which the metal, the ligand, and the complex absorb at the selected wavelength.

• A procedure for determining the stoichiometry between two reactants by

measuring the relative change in absorbance under conditions when each reactant

is the limiting reagent . In the slope-ratio method two sets of solutions are

prepared. The first set consists of a constant amount of metal and a variable

amount of ligand, chosen such that the total concentration of metal, CM, is much

greater than the total concentration of ligand, CL. Under these conditions we

may assume that essentially all the ligand is complexed. The concentration of a

metal–ligand complex of the general form MxLy is

Slope-ratio method

• Determine endpoint by following change in absorbance of:

1) reactant (decrease)

2) product (increase)

3) titrant (increase after endpoint)

• Example Titration curves for

• 𝑺 + 𝑻 → 𝑷

• where S = analyte being titrated, T = titrant, P = product

Photometric Titrations

where S = analyte being titrated, T = titrant, P = product

References

1- Douglas A. skoog, Donald M. West, F. james Holler, Stanley R. Crouch. (2013).

Fundamentals of analytical chemistry. 9th ed . Belmont, USA: Mary finch. pp:651-674.

2- Douglas A. Skoog, F. James Holler, Stanley R. Crouch. (2007). Principles of instrumental

analysis. 6th ed. Belmont, USA: David Harris.pp:335-367.

3- Tony O.(2000). Fundamentals of modern Uv-Visible spectroscopy. 1st ed. Germany:

Agilent technologies .pp:36-43.

4- David H.(2000). Modern analytical chemistry. 1st ed . London: James M smith. Pp: 394-

407.