08 - Chapter 1
08 - Chapter 1
08 - Chapter 1
PART A
PART B
1A.1 COMPLEXOMETRY
1A.2 SCOPE OF COMPLEXOMETRIC TITRATIONS
1A.3 ORGANIC REAGENTS IN CHEMICAL ANALYSIS
1A.4 ORGANIC THIO-LIGANDS AS ANALYTICAL REAGENTS
IA.5 MASKING TECHNIQUES
1A.6 TYPES OF MASKING TECHNIQUES
1A.7 DEMASKING TECHNIQUES
1A.8 SELECTION OF MASKING REAGENTS
1A.9 INDUSTRIAL APPLICATIONS OF MASKING TECHNIQUES
1A10. REFERENCES
2
1A.1 COMPLEXOMETRY
The last four decades have witnessed a tremendous change in the field of
analytical chemistry. The analysts have developed an extensive array of instrumental
techniques. These techniques are extremely sensitive and can yield results rapidly to a
high degree of accuracy, but they are expensive. The classical methods of gravimetry
and titrimetry have therefore faced severe competitions from the instrumental
methods of analysis. Gravimetric methods are slow and involve many steps in the
determination. They are now employed occasionally to establish the reliability of new
instrumental methods but titrimetric methods represent a lively and still growing area
in the field of analytical chemistry. This is due to its versatility, coupled with
simplicity and speed. The introduction of EDTA, a new titrant and metallochromic
indicators have given a unique face to the complexometric technique.
In the past few decades, the titrimetry has undergone a rapid change to keep
pace with the modem developments in the industry and research. Among the
titrimetric methods, complexometry enjoys a great deal of interest as an important
tool of analytical chemistry.
Depending upon the donor centers, i.e., the number of points of binding to the
central metal ion, the ligands may be classified into monodentate, bidentate,
3
multidentate etc. Halide ions, water, ammonia molecule etc., comes under the
monodentate ligands, while acetylacetone, ethylenediamine etc., are classical
examples of bidentate ligands. Multidentate ligands contain many donor atoms per
molecule. EDTA is a hexadentate ligand, which has two donor nitrogen atoms and
four donor oxygen atoms. The denticity of multidentate ligands is also dependent on
the steric factors. The complexation of the metal ion with bidentate or polydentate
ligands leads to the ring formation, including the metal ion. This process is called
chelation. The ring formation offers stability to the metal complex. Generally five and
six member rings are very stable. EDTA forms five member rings with the metal
atoms. Hence a stable complex of EDTA with many of the metal ions are formed.
Steric factors also affect the denticity of ligands.
For simplification purpose, EDTA is assigned the formula H4Y and its
disodium salt is therefore, Na2H2Y. It affords the complex forming ion H2Y2' in
aqueous solution, which reacts with many metal ions in 1:1 molar ratio. The reaction
with the metal cations may be represented as
It is clear from the above equation that the dissociation of the complex
depends greatly on the pH of the solution. Lowering the pH will decrease the stability
4
of the metal EDTA complex. If the metal EDTA complex is highly stable the titration
of the metal ion with EDTA may be carried out at lower pH. Hence the selectivity of
EDTA can be increased by the control of pH. For example, nickel(II) can be titrated
in the presence of alkaline earth metal ion at pH 3.5 without any interference from the
latter, whereas interference will occur at pH 10 in ammonia buffer [2]. Similarly a
mixture of Bi(III) and Pb(II) can be successfully titrated by first titrating the Bi(III) at
pH 2 with xylenol orange as indicator and then titrating Pb(II) by raising pH to 5
using hexamine [3]. The selectivity of the complexometric titrations with EDTA can
also be achieved by using suitable metallochromic indicators [4]. The use of various
metallochromic indicators has greatly enhanced its value in titrimetry. Each of these
indicator is effective over a limited pH range.
When a solution containing two cations which complex with EDTA is titrated
without the addition of a complex forming indicator, and if a titration error of 0.1% is
permissible, then the ratio of the stability constants of the EDTA complexes of the
two metals M and N must be such that Km/Kn > 106, if N is not to interfere with the
titration of M. The constants Km and Kn considered in the above expression should be
apparent stability constants of the complexes. If complex forming indicators are used
for a similar titration the ratio Km/Kn should be > 108 [3],
There are very few cases where the ratio of stability constants of the EDTA
complex of the metal ion in a particular mixture is above 106, complication often
arises when one wishes to determine a desired metal complexometrically. However
the addition of masking agents can often achieve the desired effect. Hence the
selectivity of complexometric titrations can be greatly increased by the application of
masking agents. The masking agent is added either to prevent the interference by
unwanted cations or to displace a cation from its EDTA complex, followed by the
back titration of the liberated EDTA. A wide range of complex forming reagent is
available which form, with a particular cations, complexes that are very stable
preferably soluble, colorless, and inactive towards EDTA and the indicators that are
used. The use of masking agents in the complexometric titrations have allowed an
extension of the possibilities of determination of the metal ions especially in the
routine analysis of alloys, ores, concentrates and the other systems containing the
mixture of metals. Difficulties arise in masking the cations that have small complex
5
forming tendencies or that give rise to very intensely colored complexes, and in the
determination of chemically related cations, such as Co-Ni or Zn-Cd, because of the
lack of specific masking agents.
are formed with EDTA, the equivalence point can be readily detected in such
titrations. Another merit of complexometric titrations is the determination of several
metal ions in semi-microgram levels.
6
example, in a mixture of manganese, magnesium and zinc, the sum of all the ions can
be determined by direct EDTA titration first and using fluoride and cyanide ions as
demasking agents, the amount of magnesium and zinc can be determined respectively [3].
Even by changing or readjusting the pH to different values the determination of
various metal ions in a mixture can be done. With methyl thymol blue, lead may be
titrated at a pH of 6.0 without interference by calcium; subsequently calcium is
titrated at pH 12 [3]. For the successive determination of iron, aluminium and titanium,
Li et al. used a reagent 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol [6]. Dipali and Roy
reported a simple complexometric titration with EDTA as titrant, for the determination
of calcium and magnesium in the same solution and at the same pH [7], after masking
magnesium using acetylacetone at pH 10 the amount of calcium is determined,
followed by demasking of magnesium at 90 °C and titrating with EDTA at the same
pH.
Organic reagents have been used for a long time in analytical work involving
inorganic species. They are highly sensitive and selective in their reaction towards
metal ions. They find extensive application in titrimetry. The most important field of
application for organic reagents are found in titrations of metal ions with complexing
agents both as titrant and as indicators in chelometry. The tests for the determination
of substances with organic reagents can be often performed with higher sensitivity
and selectivity. A fair degree of sensitivity and selectivity can be achieved by the
careful control of pH, reagent concentration and by the use of suitable masking
agents.
Many organic reagents form stable chelate with metal atoms or ions. Such
chelates, particularly of ligands containing two or more fused rings are found to be
extraordinarily stable as compared to non-chelate complexes [8]. Organic compounds
possessing a dual character, viz, of having both acidic and basic functional groups are
7
capable of forming neutral chelate with the metal ion. The acidic group usually
contains a replaceable hydrogen ion and the basic group contains a lone pair of
electrons, which takes part in coordination.
Another condition for effective chelation is that the functional group must be
so positioned as to favour the formation of a strain free ring. Five or six member
chelate rings are naturally the most stable chelates.
Further, the tendency of sulfur to take part in hydrogen bonding is less than
that of oxygen. Hence the sulfur containing complex is less soluble in water than the
corresponding complex containing oxygen in the place of sulfur. The solubility of a
ligand in water decreases as the hydrophobic nature is increased.
If the conditions are favourable, the vacant d-orbitals on sulfur, being probably
of low energy are suitable for back-bonding with metals having filled or partially
filled d-orbitals. With the help of HSAB concept one can expect that sulfur forms
stronger complexes with cations of larger size and in lower oxidation states. This
remarkable ability of sulfur atom as a potential donor has contributed a great deal to
the increasing interest in the use of sulfur containing ligand as analytical reagents.
The soft sulfur present in the thio-ligands can form stable complexes with 'soft
metal ions like Pd2+, Ag+, Tl+ etc., as well as borderline metal ions like Cu2+, Pb2+,
Hg etc. These metal ions being strong 'b’ class acceptors have greater affinity for
sulfur than for oxygen or nitrogen. The formation constants of a series of 'S’ and 'N’
containing chelates with various transition metal ions have been determined [9] and
these values indicate that sulfur is a better donor than nitrogen for 'b’ class metal ions.
8
1A.5 MASKING TECHNIQUES
Very few analytical reagents are highly selective in their action. Hence many
reagents which have otherwise useful properties may have restricted analytical
applications because they give similar reaction with many ions. The attainment of
selectivity has long been one of the principal objectives of research in analytical
chemistry. Considerable success has resulted from the development of new reagents,
which are inherently more selective. Employment of masking reagents affords one of
the most promising approaches to the solution of the problem of selectivity.
9
Recent developments in analytical chemistry such as activation and atomic
absorption spectrophotometry are diminishing the scope of conventional chemical
analysis. But the extent of this erosion will depend largely on how much can be
achieved in simplifying and speeding up the existing titrimetric, spectrophotometric
and gravimetric procedures without sacrificing accuracy and reliability.
It is in this field that much can be hoped for, from the simple and elegant
masking technique designed to enhance the selectivity of chemical reactions and in
many cases to achieve rapid analysis of mixtures that would otherwise take quite long
time by the conventional techniques of analysis. Employment of masking techniques
afford one of the most promising approaches to the solution of the problem of
selectivity, because of simplicity with which it may be carried out in practice.
Thiols are often proposed in place of cyanide for masking because they are less
hazardous. Reagents such as 1,2-dimercaptopropanol (BAL) [14], thioglycollic acid [15] and
unithiol (2,3-dimercapto propane sulfonic acid) [16] are being used successfully in
practical applications. The determination of Ca2+ and Mg2+ in presence of Hg2+, Pb2+
'jt
and Zn was done by masking the metal ions using unithiol [17]. Using 0-
aminoethylmercaptan [18] the titrations of alkaline earth metals are possible in the
presence of bivalent heavy metals except Pb2+ and Mn2+. a-Hydroxy acids [19]
are used in the direct titration of Bi3+ with EDTA. Thiosulfate is used as a selective
masking reagent for the determination of copper [20] and palladium [21].
10
1A.6.1 Masking by Precipitation
When the species to be masked reacts with the masking agent to yield a
precipitate, it is known as masking by precipitation. Sulfate was used to precipitate
lead in the determination of Bi3+ [22]. Using xylenol orange and cetyl pyridinium
bromide as mixed indicator, lead was determined at pH 5-6 from lead-EDTA complex
by selectively masking lead with sulfate ions [23, 24]. The precipitation is often
accompanied by coprecipitation of the species to be determined resulting in obscure
end points with low results or less accuracy.
Masking agents with sulfur as donor atom are selective towards many metals.
Copper can be masked using thiourea [27], while cadmium and zinc can be masked
with cysteine [28]. 3-Mercapto propionic acid [29] is used as a masking agent for
lead.
In recent years, EDTA and related compounds, polyamines and certain sulfur
containing reagents [30] have greatly increased masking probabilities for cations.
Majority of the masking agents for inorganic ions involve soluble complex formation.
Almost every ion can be converted into a water-soluble complex.
11
means of eliminating the interference of Fe3+ [31]. However, this type of masking by
oxidation and reduction has limited application since very few of the common
elements can exist in different oxidation states in aqueous solution. In addition to that
some of the possible oxidation states are so unstable that metals cannot be preserved
in those oxidation states. For example, Co3+ is highly unstable in aqueous phase
At times, if need arises, more than one of the above said types of masking
processes may be applied simultaneously or concurrently to advantage. For example,
Hg2+ can be masked towards a number of reagents by boiling its solution with ascorbic
acid thereby reducing it to the metal which precipitates. Thiosulfate ion [20] and thiourea
[32] in a weakly acidic medium reduces Cu2+ to Cu+ and forms a stable complex with
the latter. Iron(III) is often masked by the addition of cyanide ion followed by
reduction with ascorbic acid to form hexacyanoferrate(II) ion. Both these examples
involve simultaneous reduction and complexation.
If the principal reagent reacts at significantly different rates with the various
species of interest, selectivity can sometimes be achieved by taking advantage of the
differences in the kinetics of the reaction. This approach is termed as kinetic masking [31].
The complexation of Cr3+ ion with many complexing agents proceeds slowly
at room temperature and boiling of the solution is often required to ensure completion
of the reaction. Hence a number of metal ions can be titrated with EDTA without
interference from Cr3+ in the cold condition. Fe3+ and Cr3+ in combination can be
determined [3] by the addition of an excess of standard EDTA to the cold mixture at
pH 5-6 followed by back titration of the solution with lead nitrate solution using
xylenol orange as indicator. If the solution is kept cold, chromium does not interfere
at all. The same solution is then acidified to a pH of 1-2 and treated with excess of
EDTA and boiled for 15 minutes when the red violet Cr3+-EDTA complex is
produced. After cooling the solution to room temperature and readjusting the pH to 6,
the excess of EDTA is titrated with lead nitrate and the used up EDTA corresponds to
Cr3+.
12
By cooling the sample solution, the effective kinetic masking can be achieved
sometimes. For example, the displacement of Ni2+ from its EDTA complex by Bi3+ in
ice cold solution at pH 2.0 is very slow, while cations like Cd , Co , Pb , Cu and
Zn2+ are displaced rapidly. Hence the amount of co-ions can be determined at ice cold
solution while the amount of Ni2+ can be determined at elevated temperature by
adding excess of EDTA to the mixture and titrating the solution with Bi3+.
Masking agents especially those acting via soluble complex formation may be
added after the species has reacted with the principal reagent. This approach, which
may be called indirect masking, sometimes allows the determination of several
species with single aliquot of the sample solution.
For example, Al3+ can be determined in the presence of other cations which
interact with EDTA by complexing all the cations including aluminium with known
excess of EDTA under optimum conditions and back titrating the excess EDTA
followed by the addition of fluoride ion to mask Al3+ and release the EDTA from Al-
EDTA complex and determining the EDTA so released by using standard Zn2+ or
Pb2+ solution.
the presence of other cations which interact with EDTA by complexing all the cations
including copper with known excess of EDTA under optimum conditions and back
titrating the excess EDTA followed by the addition of 2-mercaptoethanol [38] or DL-
cysteine [39] to mask Cu2+ and release the EDTA from Cu-EDTA complex and
determining the EDTA so released by using standard Zn2+ or Pb2+ solution.
13
1A.7 DEMASKING TECHNIQUES
Demasking is the term used to describe the process by which a masked species
regains its ability to take part in its normal reactions, in systems in which, the cations
have been masked by complex formation with a suitable ligand. This can be often
achieved by adding a sufficient amount of another reagent, which has a greater
affinity for the ligand concerned, so that initially masked cation is once again
liberated.
The metal ion forming stronger, more stable complex with the masking agent
is added to the system. The masking agent sets free the metal ion under consideration
and preferentially bonds to the metal ion added. For example in ammoniacal
[Ni(CN)4]2' solution, Ni2+ is demasked by Ag+ ions, since the [Ag(CN)2]' complex
formed is more stable than [Ni(CN>4]2' complex [41,42]
The displaced nickel ions can be determined by the titration with EDTA using
murexide indicator.
Bismuth can be titrated with EDTA in the presence of Hg2+, if Hg2+ is masked
by adding slight excess of potassium thiocyanate to form soluble [Hg(SCN)4]2' [40].
After the titration Hg is demasked by adding slight excess of silver nitrate and the
Hg2+ is titrated with EDTA.
14
1 A.7.2 Conversion of Masking Agent into Non-reacting Species
The reagent, which preferentially reacts with the masking agent, is added to
the system. The reagent sets free the masking agent and combines with it to form non
reacting species. For example in a mixture containing Zn2+ and Cd2+ selectivity can be
achieved by masking Zn2+ in an alkaline medium with potassium cyanide and then
Chloral hydrate [44] or acetone can also be used in the place of formaldehyde.
1A.7.3 Adjustment of pH
The nature of the reagent used to lower the pH of the solution may also be
important. Dilute acetic acid is unsuitable for demasking the complexes of W, Mo, Ce
and the rare earths with EDTA because colloidal precipitates are obtained and
appreciable reduction occurs.
In some cases heat, chemical reaction or both can destroy the masking agents.
Masking by thiocyanate ion can be demasked by the oxidation of the ligands with the
alkaline hydrogen peroxide and heating. The excess of hydrogen peroxide can be
15
catalytically decomposed by adding a small amount of Fe3+. EDTA can be destroyed
in acid solutions by permanganate or other strong oxidizing agents
Ligands, which form volatile compounds by heating with the mineral acids or
form precipitates with some reagents can be removed from the reaction mixture.
Prolonged boiling with strong mineral acids can be used to remove stable but volatile
masking agents such as hydrogen fluoride, hydrogen chloride, hydrogen bromide,
hydrogen iodide. On the other hand silver iodide, bromide, and chloride are
selectively and sequentially precipitated, in that order from a concentrated ammonia
solution. When the ammonia is allowed to escape progressively the three halides can
be separate by filtration over suitable pH range.
When a metal ion can exist in more than one oxidation state there will be
usually differences in stability constants of corresponding complexes with a given
ligand. A change in the oxidation state of a metal ion may so decrease the stability of
a complex that the metal ion may be no longer masked, e.g. Thiosulfate ion forms
very stable complex with copper(I) in slightly acidic solution. If the solution is made
alkaline copper(I)-thiosulfate is readily oxidized to copper(II)-thiosulfate where the
metal is only weakly complexed.
Masking agents by suppressing the effects of interfering species will make the
reaction much more selective or even in some cases specific under chosen conditions.
Masking technique depends largely on complex formation, and the study of factors
governing the stability of metal complexes plays a major role in the selection of
masking agents for new application.
In any particular system many different factors are involved in determining the
effectiveness with which an ion or neutral molecule needs to be masked if it is not to
cause interference, e.g. Relatively weak complex formation may be sufficient to
prevent precipitation or to reduce an already low concentration of a foreign ion below
the level at which it causes detectable interference, but if the same ion is present at
16
very high concentrations much more effective masking may be necessary.
Considerations such as these make it difficult to lay down firm rules for masking
agents. A reagent that is suitable under one set of conditions may be quite inadequate
at different pH and at different metal ion concentration. In general however, stability
constants of the complexes must be sufficiently high, the reaction must be rapid and
the precipitation must be avoided wherever possible.
The best ligands to use as masking agents would be those which form strong
colorless complexes with the ions to be masked, but relatively weak complexes with
other cations that are present. Thus copper(II) is reduced to copper(I) by thiourea with
the formation of complexes that are more stable than those of calcium, where as the
calcium complexes with EDTA are stronger than those of copper(I). Hence calcium is
titrated preferentially with EDTA in the presence of copper.
17
bind most strongly when thiolate ions are involved but there is much weaker bonding
through thioether and -S-S- groups.
There is however, three factors that are directly responsible for the great range
that can be achieved in the stabilities of the metal complexes formed by ligands
containing these groups
One of the most useful concepts for a qualitative discussion of the intrinsic
factors that govern the strengths of the bonds formed between metal ions and
eomplexing agent is that of “hard” and “soft” acids and bases developed by
Pearson[45]. “Hardness” is a characteristic of bonds that have a high ionic character
where as “softness” is related more to covalent bond formation.
Lewis [46] defined a ‘base’ as a species which can donate a pair of electrons
to form a coordinate bond and an ‘acid’ as a species which can accept a pair of
electrons. In terms of these definitions all ligands are Lewis bases and all metal ions
are Lewis acids.
This approach has led to the generalization that hard acids form stronger bonds
with hard bases, where as soft acids prefer to coordinate to soft bases. Such
18
complexes are usually much more stable than related ones formed by a hard acid with
a soft base or soft acid with a hard base.
In general the stability of complexes of a hard metal ion with a hard base
increases with the charge on the metal ion, so that Al3+>Mg2+>Na+. With the soft
metal ions and soft bases the opposite is usually true i.e. Ag+> Cd2+>Au3+>Sn4+
Hard metal ions coordinate best to the lightest atom in a family of elements in
the periodic table. Soft acids coordinate best to one of the heavier atoms of the same
family, possibly because these atoms have vacant d-orbitals which may be used for
the ^-bonding of some of the d electrons of the metal ions that are soft acids.
Many of the masking agents mask all related cations rather than individual
species in isolation. By considering all the above points, the important requirements
that determine the suitability of a reagent as a masking agents can be summarized as:-
a. The kinds of atoms through which masking agent bond the metal ions.
b. The stability constant of the complex formed by the masking agent with
metal ion should have a high value.
c. The complex formed after masking should not have a polar group or a net
electronic charge and color.
d. The masking agent should be much more soluble in water than the metal
complex so that the former is not co-precipitated and excess of masking
agent should not affect on the method of analysis.
The most effective cations, which catalyze chemical reactions, are usually
those of transition metals such as iron, copper and manganese. These may be present
in raw materials used in industry and also be introduced by corrosion, physical or
mechanical wear out and breaking down of non-ferrous alloys and surface coatings.
Traces of heavy metal ions catalyze aerial oxidation of fats, oils, proteins and
many other organic compounds, probably by a free radical peroxide chain reaction
mechanism. Such auto-oxidation reactions led to undesirable effects like rancidity,
turbidity, disagreeable taste and odor, in natural products or manufactured items like
food stuffs and beverages. The auto-oxidation of sunflower or other unsaturated oils is
19
explained by a two-step mechanism involving the formation of R-O-O-H followed by
decomposition. The second step is metal catalyzed, so that complex forming agents
such as EDTA or tartrate ion can inhibit the reaction. The oxidative destruction
includes the loss of important constituent like ascorbic acid.
EDTA has been tried extensively in the preservation of fresh fish. Iron,
vanadium and copper are the most active among the cations which enhance rancidity
in blended fish muscles. By using EDTA, dephosforylation of inosine-5-phosphate
present in the fish muscle is inhibited and although there is little effect on bacterial
growth and hence the storage life of the haddock is increased.
Because of the chelating tendency of most of the dyes, they will form
complexes, if metal ion impurities are present. This will result in some undesirable
properties like change in color, variation in shade, tint, hue etc., between successive
batches of dyeing. So masking of various trace metal ion impurities become necessary
to ensure consistency and uniformity in the shades, tints etc.
The interference of ferric ion and other metal ions during the manufacture of
paper causes discoloration of unbleached pulp and paper. It also reduces the strength
of the paper because of localized oxidation during bleaching. These unwanted effects
can be prevented by the application of EDTA as masking agent. A detergent with
metal-sequestering properties that has been suggested for removing bloodstains
comprises a mixture of an organic acid such as dodecylbenezene- sulfonic acid with
mono-, bis-, tris- and tetra (hydroxypropyl) ethylenediamines [46].
20
1A.10. REFERENCES
21
26. J. S. Fritz, M. J. Richard and S. K. Karaker, Anal. Chem., 30 (1958) 1347.
27. J. Kinnumen and B. Wennwestrand, Chem. Anal., 46 (1957) 34.
28. W. Bemdt and J. Sara, Talanta, 8 (1961) 653.
29. S. Hara, Busekikagaku, 10 (1961) 633.
30. H. R. A. Gadiyar, R. V. Gadag and M. R. Gajendragad, Talanta, 29 (1982) 941.
31. A.N. Reily and A. J. Bam ad, “Handbook ofAnalytical Chemistry”, L. Meites
(Edn.), Me Graw Hill, New York (1963).
32. O. B. Budesky and L. Sinova, Talanta, 9 (1962) 769.
33. B. Mathew, B. Narayana, B. M. Rao, C. H. R. Nambiar and B. Ramachandra,
Mikrochim. Acta, 122 (1996) 295.
34. B. M. Rao and B. Narayana, Tr. J. Chem., 17 (1993) 138.
35. B. M. Rao and B. Narayana, Ann. Chim (Rome), 85 (1995) 105.
36. C.H. R. Nambiar, B. Narayana, B. M. Rao and B. Mathew, Analyst, 120 (1995)
1843.
37. A. Joseph, B. Mathew and B. Narayana, J. Indian Chem. Soc., 73 (1996) 633.
38. B. M. Rao, B. Narayana and K. Subramanya Bhat, Mikrochim. Acta, 112 (1994)
109.
39. B. Narayana and M. R. Gajendragad, Analyst, 117 (1992) 203.
40. L. Barcza and E. Koros, Chem. Anal., 48 (1959) 94
41. S.J. Gedansky and L Gordon, Anal. Chem., 29 (1957) 566.
42. H. Flaschka and F.Huditz, Z. AnaLChem, 137 (1952) 104,172.
43. H. Flaschka, Z. AnaLChem, 138 (1952) 332.
44. R. Pribil, Chem. Listy., 47 (1953) 1173.
45. R.G. Pearson, J. Am. Chem. Soc., 85 (1963) 3533.
46. G.N. Lewis, “Valency and the Structure ofAtoms and Molecules”, Chemical
Catalog Co., New York, (1923)141.
22
PART B
1B.1 SPECTROPHOTOMETRY
1B.2 CALIBRATION CURVE
1B.3 RINGBOM’S PLOT
1B.4 CHOICE OF WAVELENGTH
1B.5 SENSITIVITY OF SPECTROPHOTOMETRIC METHOD
1B.6 PRECISION AND ACCURACY
IB.7 COLOR DEVELOPEMENT
IB.8 CHOICE OF SOLVENT
1B.9 LIMITATIONS
IB. 10 APPLICATIONS
1B.11 PRESENT INVESTIGATION
IB. 12 REFERENCES
23
1B.1 SPECTROPHOTOMETRY
The act of identifying materials based on their color was probably one of the
earliest examples of qualitative molecular absorption spectrophotometry. Also, the
first recognition that color intensity can be the indicator of concentration was
probably the earliest application of employing molecular absorption spectroscopy for
quantitative estimation. The first measurements were made by using the human eye as
the detector and undispersed sunlight or artificial light as the light source. Later it was
found that the accuracy and the precision could be improved by isolating specific
frequencies of light using optical filters. Further improvement of the measurement
came with the use of prism and grating monochromators for wavelength isolation.
Photoelectric detectors were soon developed, but were quickly replaced with
phototubes and photomultiplier tubes. The development of solid state
microelectronics has now made available a wide range of detector type which are
coupled with the computers, provide highly sophisticated readout electronic systems.
24
molecules containing one or more chromophoric groups. The color of a molecule may
be intensified by substituents called auxochromic groups, which displace the
absorption maxima towards longer wavelength (bathochromic shift). The color
determining factors in many molecules is the introduction of conjugated double bonds
by means of electron donor and / or electron acceptor groups. The quantitative
applicability of the absorption method is based on the fact that the number of photons
absorbed is directly proportional to the number or concentration of atoms, ions or
molecules [2].
IB .3 RINGBOM’S PLOT
Ayres [3] pointed out that a straight line obtained in Beer’s law curve does not
show directly the concentration range within which accurate determination of the
colored species is possible. The optimum range for highest precision is determined by
plotting percentage transmittance against log of the metal ion concentration as was
suggested by Ringbom [4]. The optimum concentration range corresponding to a
nearly constant and high rate of change in transmittance with concentration is
indicated by a virtually linear portion of the Ringbom’s plot.
25
the maximum of e. When the transmittance of the solution increases continuously
over the wavelength range covered by the light filter, Beer’s law will not be obeyed.
the metal ligand complex. The awareness of the sensitivity is very important in
spectrophotometric determination of trace metals. The numerical expression [6-8] is the
molar absorptivity (e) at the wavelength of maximum absorbance of the colored
species.
A
Molar absorptivity (e) = — -------- (1)
c1
It is generally stated [10] that the molar absorptivity will not exceed
approximately 105.
26
detection limit is to use indirect determinations, where a stoichiometric gain in the
number of chromophores may result or the newly formed chromophore may have a
higher molar absorptivity. Reaction rate methods can sometimes have lower detection
limits than do conventional spectrophotometric measurements.
27
CV = s>,0°
(2)
X
DL= ------------(3)
S
10 cr
QL = (4)
S
28
1B.6.3 Comparison of the Results
The comparison of the values obtained from a set of results with either (a) the
true value or (b) other sets of data makes it possible to determine whether the
analytical procedure has been accurate and / or precise, or if it is superior to another
method.
There are two common methods for comparing results: (a) Student’s t-test and
(b) the variance ratio test (F-test) [17,18].
degrees of freedom, whilst the sum 2 (x-x) is considered to have n-1 degrees of
freedom, as for any defined value of x only n-1 values can be freely assigned, the
nth being automatically defined from the other values.
This is a test used for small samples; its purpose is to compare the mean from
a sample with some standard values and to express some level of confidence in the
significance of the comparison. It is also used to test the difference between the means
---------------- (5)
It is then related to a set of t-tables [17] in which the probability of the t-value
falling within certain limits is expressed, either as a percentage or as a function of
unity, relative to the number of degrees of freedom.
29
(b) The Variance Ratio Test (F-test)
This is used to compare the precisions of two sets of data of two different
analytical methods or the results from two different laboratories. It is calculated from
the following equation [17,18]:
(6)
The larger value of s is always used as the numerator so that the value of F is
always greater than unity. The value obtained for F is then checked for its
significance against values in the F - table calculated from an F - distribution [17]
corresponding to the numbers of degrees of freedom for the two sets of data.
There are two common methods for comparing results: (a) t-test and (b) the
variance test (F-test).
(a) t - Test
This method is also used to compare the values of the mean and precision of
the test method with those of the reference method [17, 18]. The value of ‘t’ when
X X
comparing two sample means 1 and 2 is given by the expression:
t= (7)
Where Sp is the pool standard deviation, is calculated from the two samples standard
deviations SI and S2 as follows :
(8)
30
(b) Variance Ratio Test (F - Test)
There are only a few elements, which give sufficiently intense absorption by
themselves and are spectrophotometrically measurable. Majority of the substances are
generally determined indirectly in a variety of ways, such as
31
IB.7.2.1 Specificity of the Color Reactions
Very few reactions are specific for a particular substance, but may give colors
for a small group of related substances only. Because of this selective character of
many colorimetric reactions, it is important to control the operational procedure so
that the color is specific for the component being determined. This may be achieved
by isolating the substance by the normal methods of inorganic analysis. But these
separation methods are often tedious and time consuming. Further there is every
possibility of appreciable loss of the analyte during these separations.
32
enough for precise measurement to be made. Stability of the color is influenced by
experimental conditions like temperature, pH etc. The solution must be free from
precipitate if comparison is to be made with a clear standard. Turbidity scatters as
well as absorbs the light.
For inorganic compounds, water normally meets these requirements, but for
majority of organic compounds, it is necessary to use an organic solvent. All solvents
show absorption at some point in the ultraviolet region and care must be taken to
choose a solvent for a particular determination which does not absorb in the requisite
wavelength region. Any impurities present in the solvents may affect the absorption at
certain wavelength and it is therefore, essential to employ materials of the highest
purity.
1B.9 LIMITATIONS
33
imperfections, or simply from scattered light due to dust particles on optical surface.
Because one has usually selected a wavelength at which the compound of interest
absorbs most strongly, the stray light falling on the sample is of wavelengths at which
the compound does not absorb strongly. Thus the stray light errors will result in a
negative bias for absorbance readings which can be represented in the equation
Where p is the fraction of all the light coming from the wavelength isolation
device, which is stray light, and Tobs and 7^ are the observed and true transmittances,
respectively. Normally the absolute amount of stray light tends to be relatively
constant with respect to the wavelength. But the fraction of stray light is highly
wavelength dependent because the amount of energy of the selected wavelength
depends on the source intensity at that wavelength. Thus, stray light errors are most
predominant at long and short wavelengths and when high absorbance is measured.
Errors also occur when a distilled water blank is used instead of a true blank
for 100 % transmittance or baseline reading. Even though there are no known
absorbing species in distilled water as well as in the blank reagent solution, the
difference in the refractive indices between the sample solution and the reference
solution must be kept reasonably close or reflective loses at the cell windows may not
be the same. Even when the incident light is highly collimated and falls on the cell
34
window at normal incidence, a small fraction of the light is reflected back at each
interface where there is a refractive index difference, at the two air-window interfaces,
and the two window-solution interfaces. Because the sample and the reference cells
are of the same composition, reflections from the air-window interfaces are
compensated for. However, reflections from the solution-window interfaces may be
different if the refractive indices of the sample and the blank are not nearly the same.
1B.10 APPLICATIONS
35
Many compilations of methodology for a variety of analytes and sample types
that are regularly updated are available [19-21]. Other general sources for
spectrophotometric analysis are commonly consulted and found helpful [22-24].
Methods specific for metals [25], and nonmetals [26] should be consulted when
dealing with these analytes. Standard methods specific to certain industries and areas
of study are very useful sources when specific sample types are being considered,
such as water and waste water [27] and pharmaceuticals [28].
The work described in chapter-2 presents two new releasing agents such as
thioglycollic acid and 2,2'-bipyridyl used for the complexometric determination of
nickel and thallium respectively. Chapters 3-10 deal with the spectrophotometric
determination of chromium, arsenic, thallium, vanadium, nitrate, iodate and periodate,
selenium and endosulfan. Chapter-3 describes spectrophotometric determination of
chromium using thionin, azure B, variamine blue and saccharin as chromogenic
reagents. Chapter-4 deals with the spectrophotometric determination of arsenic in
various environmental samples using variamine blue and azure B as reagents.
Chapter-5 presents spectrophotometric determination of thallium using thionin, azure
B and leuco xylene cyanol FF as analytical reagents. Chapter-6 includes
spectrophotometric determination of vanadium in synthetic mixtures and alloy
samples using variamine blue and thionin as reagents. Chapter-7 describes
spectrophotometric determination of nitrite in microgram levels based on the
diazocoupling reaction between p-nitroaniline with malanonitrile, ethoxyethylene-
malonicester and ethyleyanoaeetate. Chapter-8 deals with spectrophotometric
determination of iodate in table salts samples and periodate in river water samples
using thionin, azure B and variamine blue as analytical reagents. Chapter-9 describes
spectrophotometric determination of selenium in vegetables, water and soil samples
based on the oxidative-diazocoupling reaction between phenylhydrazine-p-sulfonic
acid with ethylacetoacetate and acetylacetone. Chapter-10 deals with the
spectrophotometric determination of endosulfan in water and soil samples using azure
B and variamine blue as reagents.
36
The structures of the reagents used are given below.
2. 2,2'-Bipyridyl
°2n—^ ^—nh2
3. p-Nitroaniline
0X
0z
0z
4. Malanonitrile
CM
5. Ethoxyethylene H5C2OOC
h5c2o—^
malonicester
COOC2H5
6. nc-ch2-cooc2h5
Ethylcyanoacetate
7.
Thionin
ch3
L
8.
Azure B CH3
J& tx
JYH
9.
Variamine Blue
37
SOgNa
y^s°3H
hn
JL J y
A
NH
H3(T
J *
ch3 ch3
3 ^CH3
O
12. Acetylacetone
h2ct
A XH,
13. Ethylacetoacetate
<A,„
38
1B.12 REFERENCES
39
23. L. C. Thomas and G. J. Chamberlin, Colorimetric Analytical Methods, 9th Edn.,
Tintometer press, Salisbury, England, (1980).
24 Z. Marczenko, Spectrophotometric Determination of Elements, Halsted, New
York, (1975).
25. F. D. Snell, Photometric and Fluorimtric Methods ofAnalysis, part 1 & 2, Wiley,
New York, (1978).
26. D. F. Boltz and J. A. Howell, Colorimetric Determination of Nonmetals, 2nd
Edn., Wiley, New York, (1978).
27. American Public Health Association, Standard Methods for the Examination of
Water and Wastewater, 17th Edn., American Public Health Association,
Wasington, D.C, (1992).
28. United States Pharmacopoeial Convention, United States Pharmacopoeia, 23rd
Rev., New York, (1995).
40