studies of analytical systems involving onium and basic ... · fe(cn)6=, mno4 , reo4- ions into...
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Studies of analytical systems involving onium and basic dye cations andStudies of analytical systems involving onium and basic dye cations andsolvent extraction methodssolvent extraction methods
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Soleymanloo, Shokoufe. 2019. “Studies of Analytical Systems Involving Onium and Basic Dye Cations andSolvent Extraction Methods”. figshare. https://hdl.handle.net/2134/35812.
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AUTHOR
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VOL NO. CLASS MARK
FOR REFERENCE NLV
---------------------------------------------------------------------- -
STUDIES OF ANALYTICAL SYSTUlS INVOLVING ONiill1
AND BASIC DYE CATIONS AND SOLVENT EXTRACTION HErHODS.
by
SHOKOUFE SOLEYI-IANLOO
A Thesis submitted in fulfilment of the requirement
for the award of Doctor of Philosophy of the
Loughborough University of Technology.
Supervisors: DR. A.G. FOGG and DR. D. THORBURN BURNS
@ by SHOKOUFE SOLEYI·lANLOO
1
I
I '
\-· 21 . Lou:;_~oor :,}(FJ'' -.- -;~\ •J;?t·~,i i·i' ·
of f~:C'o1:1C;;:):;':f -t:.~~-,~()"
l·-·--~--":;0~m
1 ~::.· O~lo9-;;/ol No.
I certify that this work has not been submitted
to this or any other institution for consideration
of a degree.
. .
SYNOPSIS
Studies of analytical systems involving onium and basic dye cations and solvent extraction methods.
The original intention of this project was to study selected onium extractions in continuation of previous work carried out in this laboratory.
The extraction of Cr(VI) with onium salts into organic solvents had been studied by previous workers who had shown that Cr(VI) could just as readily be extracted from acid solution without the addition of an onium salt. A study has been made of the effect of various onium compounds, and different organic solvents on this extraction. The effect of pH, as well as interfering ions ~ms studied. In the present study, in spite of the colour stability of the.complex and the coefficient of variation being 0.4%, a low apparent molar absorptivity was obtained.
This extraction method has also been used in the development of a new Atomic Absorption Spectrometric method of determining chromium. This method has been applied satisfactorily to several British Chemical Standard Steels. The procedure has the advantages of using an air-acetylene flame, and of being free from interference from iron.
The formation of a colloidal solution of a basic dye salt has been applied to the determination of inorganic phosphate in biological systems. The reaction between 12-molybdophosphate and Crystal Violet in the presence of polyvinyl alcohol was found to give excellent results. Other materials commonly found in plasmas do not interfere. It is considered that this procedure could readily be automated and could be superior to the existing automatic molybdate method.
Previous workers have studied phenylfluorone as an analytical reagent. In the present work, it was found that the use of phenylfluorone in. dimethylformamide instead of water, improved the reproducibility of the method. The sensitivity of the colour reaction between tin(IV) and phenylfluorone has been increased by addition of cetyltrimethylammonium bromide. Beer's law is obeyed from 0.2-0.8j1g/ml of tin. The final method is simple and is suitable for the determination of low concentrations of tin(IV). Its main disadvantage is that the control of pH is particularly critical.
· Diphenyleneiodonium bisulphate was prepared by published methods and its analytical properties were studied. Extraction
Fe(CN)6=, Mno4 , Reo4- ions into different organic solvents
was examined. A disadvan!~ge of this onium .salt was its extremely low solubility (1.06 x 10 M) which limited the sensitivity.
Finally a liquid state ion selective electrode was developed incorporating the periodate salt of an onium compound. This electrode was applied to the determination of periodates. Iodates and other common anions did rtot interfere. The only disadvantage was found to be the short life time of the electrode.
ACKNO\VLEDGEMENT.
I wish to thank my supervisors Dr. A. G. Fogg and
Professor D. Thorburn Burns for their stimulating guidance,
invaluable suggestions and encouragement throughout my .
studies. Also my thanks are due to the members of the
technical staff who have always been very co-operative and
helpful.
I ~10uld like to thank Hr. A.A. Al-Sibaai for his advice
on ion-selective electrodes.
I would also like to thank my many friends particularly
Dr. C. Thirkettle for advice and company during my stay at
Loughborough and Dr. S.E. Affifi for part of proof reading my
thesis.
My thanks are also due to the University of Tabriz (Iran)
for study leave.
Finally, but by no means least, I would like to thank
my husband for his patience, help and encouragement.
I
DEDICATION
This Thesis is respectfully dedicated
to my brother
Dr. Soleymanloo
for without his encouragement over the
years none of this could ever have been
achieved.
CONTENTS
SYNOPSIS
ACKNO\'iLEDGEHENTS
DEDICATION
CHAPTER I
GENERAL INTRODUCTION
1.
2.
3;
Principles of solvent extraction
1.1. Distribution ratio or extraction coefficient
1.2. Fundamental principles.
a) Distribution law.
b) Process of extraction.
1.3. Extraction system.
1.4. Theoretical considerations.
1.5. Practical considerations.
Onium compounds.
Basic dyes.
CHAPTER II
Extraction and spe.ctro hotome.tric determination of chromium VI with Tri-n-Butylbenzylphosphonium chloride.
1.
2.
Introduction.
Experimental.
2.1. Re.age.nts.
PAGE NO.
1
1
3
8
15
20
22
28
36
4o 4o
2.2. Procedure. 41
2.3. Effect of different organic solvents. 42
2.4. Effect of different onium compounds on the. 45
solvent extraction of dichromate (cr2o
72-)
into chloroform.
2.5. Conformity to Beer's Law. 45
2.6. Composition of the. complex. 46
2. 7. Stability of the complex with time.. 47
2.8. Reproducibility of results for a fixed 47 volume of standard potassium dichromate solution (H/200) at 448nm.
2.9. Holar absorptivity.
2.10. Study of interferences. 47
47
4.
2.11. Reducing potassium permanganate in the presence of dichromate using sodium azide.
Preparation method of diphenyleneiodonium bisulphate.
Conclusion and discussion.
PAGE NO.
50
51
53
CHAPrER III. Determination of chromium in steel by Atomic Absorption spectrophotometry using an air-acetylene flame.
1.
2.
Introduction
Procedure.
54
69
2.1. Instrumental alignment, settings and 69 the effect of burner height and fuel composition on the determination of chromium.
2.2. Preparation of the sample. 70
2.3. Reagents. 71
2.4. Experimental. 73
2.4.1. The effect of observation height on Cr(VI) 73 in aqueous and MIBK.
2.4.2. The effect of $lame composition on the determination of Cr(VI) in organic phase (MIBK).
74
2.4.3. The effect of variation of slit width. 74.
2.4.4. The effect of flame composition on the 75 interference of iron in the determination of Cr(VI) extracted into MIBK.
2.4.5.· The effect of increasing concentration 75 of iron(III) on the absorption signal of 20 g per ml and chromium(VI) extracted into !HBK.
2.4.6. Effect of flame composition on the 77 interference of iron in the presence of a melting out agent in the determination of chromium(VI).
2.4.7. Study the effect of the additHm of 80.
2.4.8.
different acids on the extraction of Cr(VI) from aqueous media into l1IBK.
Study of the effect of hydrogen ion concentration on the extraction of Cr(VI) into MIBK. Effect of Ce(IV~ ions on the absorption of Cr(VI).
81
82
PAGE NO.
2.4.10. Calibration curve for Cr(VI) extracted 82 into HIBK.
2.4.11. Methods of separating iron(III) from 83 Cr(VI).
2.5.. Analysis of steel samples. 86
2.5.1. Determination of chromium in steels. 88.·
2.5.2. Final procedure. 88
3- Conclusion and discussion. 90
CHAPl'ER IV
The Determination of Inorganic Phosphate in Biological Systems.
1.
2.
3.
General Introduction.
Direct measurement of inorganic phosphate.
Reduction of molybdophosphate complex.
Complex formation between molybdophosphate and basic dyes.
Experimental.
93 93 95
99
105
2.1. Reagents. 105
2.2. An experimental assessment of the 106 determination of phosphate with Halachi te Green according to the method of Itaya and Ui.
2.3. Application of Itaya and Ui's method to 107 different dyes.
2.4. Application of solvent extraction to Itaya 109 and Ui's method using different dyes.
2.5. Investigation of the determination. of the 110 phosphate with Crystal Violet.
2.5.1. Reagents 112
2.5.2. Basic procedure. 113
2.5.3 •. Study of the effect of temperature on 113 the formation of molybdophosphate-Crystal Violet complex.
2.5.4. Standard curve and absorption spectrum. 114
2.5.5. Comparison of the present method with 115 Altmann et al's method.
2.5.6. Possible application of the present 116 method on determination of phosphate in biological systems.
2.5.7. Checking the method using an unknown 117 sample.
Conclusion and discussion. 119
PAGE NO CHAPTER V
Study of the Possibility of Improving the Phenylfluorone Method for the Determination of Tin.
1.
2.
3.
Introduction.
1.1. Phenylfluorone.
1.2. Effect of surfactants on reagent-metal complex system.
1.3. Some studies in DHF - water system.
Experimental.
2.1.
2.2.
2.3.
2.4.
2.5.
2.6.
Ethanol-water system.
DMF system.
Reagents.
Procedure.
Stability of the tin(IV)-phenylfluorone complex with time.
The precision of the determination.
Conclusion and discussion.
CHAPTER VI
The Development of an Ion-Selective Electrode Responsive to Periodate.
1.
2.
3.
Introduction.
Experimental.
2.1. Possible precipitation and extraction of salts of different• onium ions with periodate into a-dichlorobenzene.
2.2. The electrode assembly.
2.3. Preparation of the membrane and the use of the electrode.
2.4. Evaluation of the electrode.
2.5. The effect of length of time of soaking in organic solvent.
2.6. Stability of the electrode response.
2.7. Electrode response.
2.8. Study of the interfering ions.
2.9. Application of TCEPI-periodate electrode in potentiometric titrations.
Conclusion and discussion
121
121
123
124.
125
125
127
127
128
128
129
129
131.
135
135
135
137
138
139
139
139
141
141
142
PAGE NO.
CHAPTER VII
Final Conclusion and discussion 148
REFERENCES
CHAPTER I
GENERAL INTRODUCTION
The ion-association-extraction technique is widely used
in both qualitative and quantitative analytical procedures. A
large cation is used to extract the anion to be determined from
water into an organic phase as an ion pair. Two types of
reagents are in common use. These are the onium compounds and
the basic dyes. In the following discussion the principles of
solvent extraction are introduced first, followed by a general
discussion of the use of onium compounds and basic dyes in
analytical chemistry.
1. Principles of solvent extraction.
Solvent or liquid-liquid extraction is based on the
principle that a solute distributes itself in a certain ratio
between two immiscible solvents, one of which is usually water
and the other an organic solvent such as benzene, chloroform,
carbontetrachloride or methylisobutylketone (HIBK). In certain
cases the solute can be more or less completely transferred into
the organic phase. The distribution coefficient will depend
upon the activities of the solute in the two solvents. The
effectiveness of extraction can be defined as follows:
1.1. Distribution ratio or extraction coefficient.
The distribution of a solute between two immiscible
solvents is given by the distribution ratio, or extraction
coefficient, D, as follo\~s:
1
D·== Total metal concentration in the organic phase Total metal concentration in the aqueous phase
where the subscripts o and w refer to the organic and aqueous
phases respectively.
i) Degree of extraction (E)
For practical purposes for analytical chemists E is
more-useful than D.
100 { A] V 0 0
(AJ V +(A) V 0 0 w w
100 D
where V represents solvent volume, { A)0
and {A] w total metal
concentration in organic and aqueous phases respectively, D,
distribution ratio.
ii) Separation factor ( j3 )
A term must be introduced to describe the effectiveness
of separation of two solutes. The following equation relates
the separation factor to the individual distribution ratios:
{A)/[A)w B 0
= B i B w = B w
where [A J and {B) represent the respective solutes. Complete
separations can be achieved quickly and easily when one of the
distribution ratios is _very small and the other comparatively large.
2
----------------------------------------------- - -
1.2. Fundamental principles
a) Distribution law
In the simplest solvent' extraction systems the distribution
of solute between the aqueous and organic phases is constant in
accordance with the classical Nernst( 1)distribution law. At
equilibrium, at a particular temperature, the ratio of the solute
concentrations in the two phases is always constant provided that
there are no chemical interactions between the solute and either
of the solvents. In the simple case the distribution coefficient
(~) is given by:
~ =
The two main cases of deviation from the distribution law result
from variations in the activity coefficient and chemical interaction
between solute and solvent. The first cause does not normally
produce large deviations but the second cause can sometimes have a
marked effect on the solute distribution. The chemical interaction
effect can be illustrated by considering the distribution of acetic
acid between benzene and water. ,
The distribution of acetic acid itself may be represented by:
~=
In the aqueous phase acetic acid dissociates:
3
f CH3cooH)0
fCH3COOHJw
f H+) [ CH3coo-)
(CH§OOHJ w
(1)
(2)
I
In benzene there is dimer formation:
The overall distribution of acetic acid is given by:
I CH3COOH I 0
I CH3COOH I w-
[ CH3cOOH] 0
+
(3)
(4)
Incorporating the (1), (2), (3) equilibrium expressions into the
equation (4):
D = Kr, ~ + 2KJ CH3COOH ] J 1 + KA/[ H+]
(5)
Thus the distribution of acetic acid varies as a function of pH
and acetic acid concentration.
b) Process of extraction.
As shown above the acetic acid extraction was seen to
involve three chemical aspects:
i) Chemical interactions in the aqueous phase.
ii) Distribution of an extractable species.
iii) Chemical interactions in the organic phase.
i) Chemical Interactions in the Aqueous Phase.
A major distinction between organic and inorganic extractions lies in
the extent to which the formation of an uncharged extractable species
depends on chemical interaction in the aqueous phase. Most organic
4
I , __ :
compounds are uncharged, although those containing acidic or basic
functional groups can undergo proton-transfer reactions that result
in charged species, such as RCOO-. Such species have obviously
different solubility characteristics in the aqueous and organic
phases, and their formation must be accounted for in calculating
D. Hence many organic and biochemical extractions can be carried
out by means of pH control. Metal salts are generally soluble
in aqueous media, not only because of the high dielectric constant
of water, which readily permits dissociation of ionic species,
but also, more important because the basic character of water
results in the solvation of metal ions which gives these ions a
solvent sheath that reduces electrostatic interaction and makes
them more "solvent-like". The role of the complex-forming metal
extraction agents is essentially to supplant the coordinated water ·
from around the metal ion to give a species that is more likely
to be compatible with organic solvents.
ii) Distribution of Extractable Species.
Although the ratio of solubilities of a solute in each of two
solvents may not be critically related to KD' there is some
correlation between the distribution and the relative solubility
in the two phases. In solutions where specific chemical forces are
not active, the classical principle of "like dissolves like" is of
great help in predicting relative solubility and extractibility. This
principle may be expressed in modern terms as Hildebrand's Theory(2)
of regular solutions, from which the solubility is seen to increase
as values of the solubility parameter, ur .. of the solute and
(2) solvent approach each other • The solubility parameter, defined
as the square root of the heat of vaporization per millilitre, is a
s
measure of cohesive energy denaity.Comparison of solubility
parameters should be of maximum assistance in dealing with those
organic extraction systems in which specific chemical or associative
f . t' (3) orces are 1nopera 1ve • In systems where hydrogen bonding
may be present, particularly those involving an aqueous phase,
the solubility parameter is inadequate in predicting solubility.
This might be expected as this concept is, strictly, only
applicable to regular solutions.
iii) Chemical Interactions in the Organic Phase.
Chemical interaction of the extracted species in the organic phase
·would lower its concentration in that phase, and, hence, improve
extractibility. For example, with carboxylic acid extractions,
the use of an organic solvent in which the acid dimerizes, results
in a higher D value than would be obtained using an organic
phase in which dimerization cannot occur. Ion-association
complexes which are dipoles often form higher aggregates in
organic solvents at higher concentrations. Where polymerization
occurs, the dissociation constant will vary with the concentration
of the extracted material.
Most metal salts are ioni~ compounds which dissolve readily
in water. The high dielectric constant of water facilitating
dissociation into the oppositely charged ions. At the same time
the salts are usually insoluble in organic solvents because these
have low dielectric constants. Metal ions dissolved in water are
usually solvated, i.e. they are coordinated 11ith one or more
molecules of water; for example the iron(III)ion is Fe(H2oj~+. In
all metal extraction systems some or all of these water molecules
must be removed in order to obtain the element in a suitable form
for extraction into an organic solvent,this form must also be uncharged.
6
The formation of no overall charge is a prerequisite for the
extraction of metal ions into organic solvents. Such species may
be formed by metal-containing ions through coordination with basic
ligands which replace the'water molecules originally in the
solvation shell. The charge on the ion is neutralized either by
the ligand, or some other component of the system.
(4) . One new procedure that has been suggested by Murata to
speed up extractions is the formation of a homogeneous phase by
raising the temperature of the system. On cooling the two phase
system is reestablished and equilibrium is quickly attained. This
method is based on the higher solubility of organi~ solvent in water
at higher temperatures. The complex is formed as soon as a homogeneous
solution is obtained. This mechanism of equilibration by achieving
an homogeneous state is significantly different from the common
mechanical shaking method. In the homogeneous method, molecules
of the organic solvent enter freely into the aqueous solution and
consequently, the water structure of the aqueous medium and the
environment of the solute species are altered extensively by the
intervention of the organic solvent molecules.
Murata and Ikeda(5)have devised a liquid-liquid extraction •
method of this type for the determination of molybdenum(VI). The
method is based on the property that a relatively hydrophilic
organic solvent, propylene carbonate (4-methyl-1,3-dioxane-2-one)
which forms a homogeneous phase with water at )70°C but is immiscible
with water at room temperature. Molybdenum(VI) in dilute hydrochloric
acid medium at pH 2.0 to 3.0 is extracted by shaking the solution
0 with an equal volume of solvent, and the mixture heated above 70 C;
extraction is achieved when the organic layer separates at room
7
temperature. (4)
Murata, Yokoyama and Ikeda also used the
homogeneous liquid-liquid extraction method for the extraction of
iron(III)thenoyltrifluoroacetonate. Iron(III) was extracted
rapidly and completely. The efficiency arises from using
propylene carbonate solvent and operating at the higher extraction
temperature. The higher temperature yields the following results:
the constitution of the mixed solvent (water and propylene carbonate),
the easy mass transfer and interaction between iron(III) and
thenoyltrifluoroacetonate(TTA) because of the appearance of a single
phase, and the completion of the successive chelation of iron(III)
with TTA.
1.?) Extraction Systems
Solvent extraction provides a rapid and convenient
approach to the separation and isolation of metal ions from complex
mixtures. Extensive bibliographies tabulated over 5000 reported
procedures that utilize this method. Systems have been classified
both by element and by complexing agent. The monograph of ' (6) . .
Stary comprehens1vely covers the topic and reviews the material
published before 1964, providing a massive amount of information
on the optimum conditions for me~al chelate extraction. The last
of the encyclopaedical and very comprehensive biennial revie\1sof
Freiser(?l, on the application of solvent extraction in
analytical chemistry covers the pertinent material up to the
.beginning of 1966.
For many years a review on extraction procedures was
included in the biennial review in Analytical Chemistry, but the
last one was in 1968(B)and next one is expected in 1976.
The progress of analytical chemistry during the period
1910-1970 is reviewed by Brooks and Smythe (9). The topics
considered are: the volume of the relevant literature, the countries
in which the work was done, the language in which the paper was
written, the literature of analytical chemistry, broad trends in
the subject, methods used, and the analytical chemistry of
individual elements. Some tentative conclusions also are made
about future short-term trends. Katz(10)has also reviewed the
publications of solvent extraction methods in Analytical Chemistry
from 1968 to 1971. A review by Bailes and Winward(11 )considers
process developments, extractors, liquid-liquid extraction and.
process design. A review with 198 references has been given by
St ' (12) ary • In this review the theory of solvent extraction and
its applications to the separation of alkali-metals., alkali-earth
metals, transition metals, transuranic elements, halogens, chal-·
-cogens and Groups III A and IV A is discussed. Current uses of
liquid-liquid extraction in analytical chemistry has also been
(13) revie~ed by Irving
In considering the chemistry of extraction processes,
several classification schemes are possible and a number of them
have been given detailed consideration. Some schemes have
classified the procedures according to the type of reaction
governing the transfer of the inorganic species from the aqueous
·to the organic phase; others have classified extraction systems
according to the extractant or to the solvent type used.
Obvi~usly, none of these classifications can be very exact. In most
extraction systems, more than one reaction takes place simultaneously.
Ionic dissociation and molecular aggregation frequently complicate
extraction equilibria. In addition, some extractants belong to
9
more than one class, depending on experimental conditions.
The extraction sytems have been distinguished by
Kertes( 1G)into four main systems as follows:
a) Distribution of Simple Uolecules,
b) Extraction by Compound Formation, the extractants being
chelating agents, carboxylic and alkyl-aryl sulphonic acids, and
acidic phosphorus esters. In the majority of chelating extractive
systems it seems to be a rule that whenever the coordination
number of the metal is double its ionic charge, the chelate formed
satisfies the coordination requirements of the metal and the metal
is readily extractable into both polar and non-polar solvents.
Extractions by carboxylic and sulphonic acids have been useful in
the separation of metals having similar chemical properties, by a
simple adjustment of the pH of the aqueous phase. Carboxylic
acids, or their solutions in an organic solvent, extract a large
number of metals(14•15). Due to the relatively high solubility of
carboxylic and acidic sulphonic compounds and their metal salts
in·aqueous solutions and their usually low extractive power, they
have not been very popular during the last few years.
c) Extraction by Solvation; extraction of acids by
solvating the hydrogen ion and extraction of salts by solvating
the metal cation. The extractants are either carbon-bonded or
phosphorus-bonded oxygen bearing extractants. The oxygen in these
compounds may be replaced by any other donor atom. All oxygen-
bearing organic solvents are considered to extract electrically
neutral species by virtue of solvation. Despite the similar
extraction reaction, it is reasonable to distinguish between carbon-
10
------------------------------------------
I
I
bonded and phosphorus-bonded oxygen bearing solvents. It is the
strong~y polar character of the latter which is responsible for
their different extractive efficiency. One of the most striking
differences between the two types lies in the specific role
ascribed to water. In organophosphorus ester systems, water
is often eliminated from the organic phase metal complex, whereas
in ethers and ketones water is a necessary part of the complex,
usually acting as a bond between the solvating molecules and the
inorganic salt. Another dominant difference is the extent of
solvation of the inorganic species. Owing to the high extractive
power of phosphorus extractants, solvation by a few solvent molecules
is sufficient to transfer the metal salt from the aqueous
solution.
d) Extraction by Ion Pair Formation; the extractants
being bulky ionic extractants of the polyphenyl metal-base type, •
the polyalkyl-ammonium type and the salts of high molecular weight
aliphatic amines.
The overall heterog'eneous extraction equilibrium involving
the salts of polyphenyl bases usually conforms to simple mass action
equations, except for systems inyolving qua-ternary ammonium
compounds. This difference is due to the tendency of ammonium
compounds towards molecular association complexes. In the case
of polyphenyl bases, slight structural modifications, such as the
.substitution of the normal alkyl chain in the triphenyl n-propyl-
phosphonium ion to give the isomeric triphenyl iso-propylphosphonium
ion, may bring about marked differences in the extractability of
ion pairs (19). Increasing the alkyl chain from methyl to heptyl
produces a steady increase in the metal extractability but
11
increasing the side chain beyond 7 carbon atoms does not further
( 19) enhance the extractability •
Ammonium compounds having at least one, but preferably
more, long-chain alkyl radicals in the molecule have proved to be
useful practical extractants. Extraction can be controlled by
the choice of reagent structure. Symmetrical cations are usually
better extractants than unsymmetrical ones with an equal number
of carbon atoms. Substitution of an aromatic group for an aliphatic
one, frequently enhances the extractive power. Short alkylchain
qua~ternary ammonium salts exhibit surface active properties
and form miscells. The critical miscell concentration depends on
the structure of the cation, the nature of the anion, the presence
of electrolytes in the solution, and the temperature.
. (17) Fre1ser has classified the inorganic extraction systems
into two broad categories of neutral extractable complexes; those
that involve chemical bonding or coordination, and those that form
by essentially electrostatic forces or ion-association.
The neutral coordination complexes could be classified
as follows:
a) Simple(Monodentate)Coordination Complexes, these are
formed by the combination of cationic metal ions, such as Hg(II),
Ge(IV), and As(III), with anionic monodentate ligands (e.g. halide
.anions), giving neutral complexes that are extractable in hydrocarbon
(e.g. c6H6) and chlorinated hydrocarbon (e.g. CHC13
) solvents.
b) Heteropoly Acids, in these complexes, the central ion
is itself complexed rather than monoatomic, e.g. phosphomolybdic
acid, H3Po4.12Mo03. Heteropoly acids are highly solvated by
12
hydrogen bonding; therefore their extraction requires the use
of oxygenated solvents.
c) Chelate(Polydentate Coordination) Complexes, these are
formed by bonding of the metal ion. by ligands which can occupy at
least two coordination sites, resulting in a cyclic compound.
When the charges of metal ion and the ligands match, as with Fe(III)
and three 8-hydroxyquinolinate ions, a neutral chelate results,
which is often much more soluble in organic solvents than in
aqueous media and is therefore of great interest in extraction
procedures.
The complexes that form by electrostatic forces or
ion-associations could be described by the following scheme:
d) Simple Ion-Association Complexes, large and poorly
hydrated ions tend to associate to form neutral compounds that are
soluble in organic solvents, particularly when one of the ions
has organic character. Thus tetraalkylammonium ions (R4N+) will
form benzene, amylalcohol, and chloroform soluble salts with large
inorganic anions, such as perchlorate, thiocyanate, and such organic
anions as tetraphenyl borate and ~lkylphenolates.
In addition to the types of extraction systems mentioned
above, there are. several other important types in which the
formation of the extractable complex involves a combination of
these factors:
a) Mixed Simple Coordination and Ion-Pair Systems:
Quite a number of metal cations form negatively charged
complexes with monodentate anionic ligands, such as the halides,
thiocyanate, and oxyanions. These complexes when paired with suitable
13
cations are extractable in organic solvents. One such cation
formed in acidic solution is the hydrated hydronium ion,[H(H2o) ] 3 o+
whose requirement of further hydrogen bonding for stabilisation
necessitates the use of oxygen-containing solvents. Thus extraction
of Fe(III) from 611 HCl as (H9o4 +, FeC14 -), is quantitative when
ethers, alcohols, ketones, or esters are employed, but negligible
if hydrocarbons or chlorinated hydrocarbons are used. A more
stable type of cation, such as a tetrasubstituted ammonium,
phosphonium, or arsonium cation permits ion-pair extraction of
2- -FeC14-, Znc14 and Mno4 into hydrocarbon solvents. In this
type, cations of triphenylmethane dyes not only are suitable for
such extractions but also provide the basis for colourimetric
determination of the extracted metal ion.
b) Mixed Chelation and Ion-Pairing Systems.
If a neutral chelating agent such as o-phenanthroline
reacts with a metal ion, the resulting chelate is positively
2+) charged (e.g. Fe phen.3
• Such large cations readily pair with
suitable anions, such as Clo4- to give extractable species.
Analogously, if an anionic chelating agent forms a negatively
charged chelate, pairing with a suitable cation such as (c4
H9
)4N+
can bring about extraction. For example ethylenediaminetetraacetic
acid (EDTA) chelate can be extracted using high molecular-weight
quaternary ammonium ions.
c) Mixed Ligand Chelates
Metal ions whose coordination number is more than twice
their electrical charge react with bidentate ligands to form
chelates that are termed coordinatively unsaturated. In such
chelates, for example, !1g(oxinate)2 , the coordination sites not
14
occupied by chelating agents are filled by water, which results
in a rather poorly extractable complex. Improved extraction
results when these water molecules are replaced by organic ligands
such as alcohols, ketones or amines. Extraction of Th(IV)
with TTA (thenoyltrifluoroacetone) is greatly enhanced in the
presence of tributyl phosphate(TBP) because of the formation of
the mixed ligand chelate, Th(TTA)4 .TBP.
1.4) Theoretical Considerations
a) Solvent Extraction;
The following discussion concerns the distribution of one or
more substances between two immiscible liquid phases in equilibrium
at the same temperature and pressure.
The first quantitative studies of solvent extraction were
carried out by Berthelot and Jungfleisch( 1S), .who verified experimentally
in 1872 that 11 les quantites dissoutes par un rapport constant".
In 1891, Nernst(1)investigated.the subject more thoroughly and
stated the "partition isotherm" in the following form: "A solute
will distribute itself between two essentially immiscible solvents
in such a way that the ratio of the concentrations of the solute
in the two phases after equ~ilibration has been achieved at a
particular temperature is a constant, provided the solute has the
.same molecular weight in each phase".
For the chemical species A, distributing between solvents
1 and 2, equilibrium is established, at constant temperature and
pressure,
A in each
when the partial molar
( 10) phase are equal:
15
free energies (' of the speCies
(1)
substituting /' = fo + RTlna (2)
in which~ is the standard partial molar free energy, .a is the
activity, R is the gas constant and T the absolute temperature.
Equation (2) may be applied to both solvents, thus at equilibrium:
~oA, 1 + RT ln aA, 1 = ~o A,Z + RT lnaA, 2 (3)
or (4)
which reduces to
expfcto e. A,1
or
-foA,2)/RT} = (A)2/(A)1 = f2(A)2/f1
f/f2 exp {( t 0 A,1 - (oA,2/RTJ - liJ>
in which KD is the distribution coefficient, and f1
and f2
are ·
the activity coefficients for the species A.
For practical purposes the analytical chemist described the
(6)
efficiency of extraction procedures in terms of percent extracted,
% E. This term can be defined as the percentage of the species
A initially present in phase 1 1 which extracted into phase 2. This
quantity is related to the distribution coefficient.
in which v1 and v2 are the volumes of the initial and the
extracting phases, respectively.
16
(?)
Two types of application can be distinguished in solvent
extraction techniques. Most common is the use ·of liquid-liquid
extraction to separate the material of interest. The other major
application is the cleanup of the reaction products to remove
unwanted impurities. The first type is of major interest in
analytical chemistry. The effectiveness of a separation is usually
expressed in terms of the separation factor j3 which is related
to the individual distribution coefficient:
~= (A);/(B)
2 (A)/(B)1 =
(A);!(A)1
(B)/(B)1
= (8)
in which (A) and (B) represent the activities of the species A
and B, respectively. In situations in which one of the distribution
coefficients is very small and the other is rather large, clean
separations can be achieved quickly and easily.
In those situations in which the separation factor is sufficiently
large, but the smaller of the tt~o distribution coefficients is great
enough to allow significant amounts of the interfering material to be
extracted, it is necessary to resort to various techniques to
suppress the extraction of the updesired component. These techniques
include stripping, backwashing, using masking agents, and using
salting-out agents.
b) Ion Pair Formation
Ion pairs are symmetrical electrolytes and they have no net
charge, although they should have a dipole moment. They will
therefore make no contribution to the electrical conductivity.
Their thermodynamic effects will be those of removing a certain
number of ions from solution and replacing them by half the number
17
I
of dipolar 'molecules'.
In ion pair formations the production of an extractable uncharged
species occurs by physical forces of attraction in contrast to
chemical bonding as in the formation of coordination complexes.
Both types, however, behave in accordance with law of mass .. action.
Hence for the reaction A+ plus B- to form the ion pair A+B- we
have:
K = fA+B-)
(A+J[ B-)
The main feature which characterises an ion pair is the electrostatic
nature of the interaction so that there is no ionic specificity 1
apart from a size effect and no fixed geometrical arrangement or
coordination.
According to the Bjerrum Theory(3) 1 the average effects of
ion-pair formation on the basis that all oppositely charged ions
within a certain distance of one another are 'associated'' into
ion-pairs. In reality a momentarily fast-moving ion might come
within the critical distance of another ion and pass by without
forming a pair. According to this theory, the value of the ion
pair formation constant, K, depends on the dielectric constant of
·the solvent,£, the temperature, T, and the size and charge of the
ions.
~----~----------------------------------~-----
X"'
K 4JTN
= 1000
b Q(b) = exp.x. x ·dx J -4
b =
Q(b.)
2kT = potential energy of the ion pair.
a = mean diameter of the ions.
t= critical distance for association (ions must approach within
this distance) e.g. 3-572 for 1:1 electrolytes.
Solvents of lower dielectric constant favour ion pair formation
since the critical distance is larger and can exceed the ionic
radii. The effect of temperature on K depends on the variation of
£_with T. \1ith most solvents of high dielectric constant 'i:...T
falls with T and hence association increases.
High electrolyte concentrations help in these ways:
1) mass action effect (if the electrolyte possesses suitable
associating ions)•
2) reduces [., •
3) reduces water activity and hence replacement of hydration
sheath occurs.
Any interactions in addition to the coulombic forces serve to
stabilize the ion pair.
To reduce the behaviour of ion-association extraction systems
to mathematical expressions is more difficult than in the case with
chelate systems.
19
1.5) Practical Consideration
The reason for the popularity of solvent extraction lies
in the speed, ease, and convenience of the technique. The
separations are clean, because the relatively small interfacial
area between the two liquid phases avoids any effects analogous
to the undesirable coprecipitation phenomena that plague most
precipitation separations. This technique can be used for
purposes of separation, preparation, purification, enrichment,
and analysis, on all scales of working, from microanalysis to
production processes. In most cases of its use in analytical
chemistry a simple separating funnel is all the apparatus required.
The extraction step usually requires only a few minutes to carry
out, and the procedures are applicable to both tra-::e and macro
analysis. A further important advantage of the method lies in
the convenience of subsequent analyses of the extracted species.
Thus, if the extracted species is coloured, as in the case with
many chelates, the spectrophotometric method can be employed.
Alternatively, the solution may be aspirated for atomic absorption
or flame spectroscopic analysis. If radiotracers are used,
radioactive count~ng techniques can be employed. Another
objective measure of the continued popularity of this technique
in analytical chemistry lies in the wealth of literature continuing
to appear on the subject.
When employing the solvent extraction technique, one of the
most important considerations is the selection of a suitable
organic solvent. Apart from the fact already mentioned that it
must be virtually immiscible with water, the solubility of the
complex in the solvent must be high if a good separation is to be
obtained. To prevent changes in volumes, the phases should be presaturated
20
with each other before use, and t1~0 phases must separate quickly
and without emulsion formation.
Violent shaking of the two phases in the extraction vessel
is not necessary and must be avoided. This action is likely
to produce an emulsion. Sometimes the sample itself will serve
as an emulsifying agent. To break an emulsion, filtration,
centrifugation, adding neutral salts to the aqueous layer, adding
an additional solvent, changing the pH slightly, or altering
the volume ratio may be useful.
From the safety point of view the flammability and toxicity
of the organic solvent will play a part in the final choice.
Solvent extraction separations are dependent mainly on two things
for their useful operation:
a) The distribution ratio of the species between the organic
and aqueous phases,
b) The pH and salt concentration of the aqueous phase. !1uch
of the selectivity which is achieved in solvent extraction
is dependent upon adequate control of the pH of the
solution.
The application of solvent extraction to the analysis of
inorganic materials may be divided into two types:
i) Those in which interfering elements are removed, leaving
the element or elements to be determined in the aqueous
phase,
ii) Those in which the element or elements to be determined are
extracted into the organic phase.
21
2) Onium Compounds
The group of ionic compounds kn01m as the "onium" salts has
a number of common structural characteristics. (19) The cation
has a single positive charge and consists of a single atom
surrounded by a number of organic groups or hydrogen atoms. The
central atom may be an element from group rv·, V, Vi, or VII,
while the organic group may be alkyl or aryl, or a combination of
these. The number of attached groups is determined by the tendency
of the central atom to attain an inert gas'structure. Thus the
elements of Group V are surrounded by four groups, those of Group
VI by three and those of Group VII by two. !1any onium salts are
of interest as analytical reagents. Their most important property
from this point of view is their ability to form sparingly soluble
ion-association products; many of these are extractable, into
organic solvents.
When the ion-association products are extractable, a spectra-
photometric method of determination is often possible, whereas if
the product is precipitated quantitatively a gravimetric finish
may be used. The reactions which give these ion-association
products may be classified conv~niently as follows:
a) The reaction with oxyanions, ·for example perrhenate, dichromate,
permanganate, arid perchlorate.
b) 1lne reaction with halogen complexes, those of mercury(II},
tin(IV), and cadmium and zinc.
c) The reaction with thiocyanate complexes, those of iron(III)
and cobalt(II).
d) The reactions with other anions, which include those formed
from nitre compounds, e.g. hexanitrodiphenylamine, anionic
22
I
!
complexes of metals formed with reagents like potassium dithio-
oxalate, and those of certain heteropoly acids, e.g. molybdophosphoric
acid.
The preparation and general properties of onium compounds
. (20) have been rev1ewed by Heal. The reaction and the analytical
applications of tetraphenylarsonium, triphenylmethylarsonium,
tetraphenylphosphonium, tetraphenylstibonium, triphenyltin,
triphenylsulphonium, selenium, and tellurium cations have been
reviewed by Bowd, Thorburn Burns and Fogg. These authors report
a large number of analytical methods of which the gravimetric and
spectrophotometric procedures are of particular interest in the
present study. Tetraphenylarsonium chloride seems to have been
studied in the greatest detail. It forms quantitatively, a white,
crystalline, water insoluble precipitate with perrhenate and thus
provides a basis for a gravimetric method for the determination
. (21 22) of perrhenate 10n. '
A colorimetric method for the estimation of rhenium in rhenium
ores also was first reported by Tribalat (23). Rhenium is oxidized
to perrhenate, extracted quantitatively with chloroform as the
tetraphenylarsonium perrhenate, ~onverted to the yellow
thiocyanatorhenate(VI) and then determined spectrophotometrically.
Manganese( 24)has been determined in calcium carbonate by first
oxidising Hn(II) to permanganate with metaperiodate and then
extracting the tetraphenylarsonium permanganate with chloroform;
Murphy and Affsprung( 25)have shown that gold may be determined
spectrophotometrically by forming the chloroform soluble precipitate
of tetraphenylarsonium chloroaurate(III).
23
- -· -- -- ------------- --
. (26) As far as the platinum group of metals is concerned, osm1um
can be extracted as the hexachloro-osmate(IV) with the tetraphenylarsonium
ion into chloroform, and can be determined spectrophotometrically,
(27) while studies of the solubility of tetraphenylarsonium chloro-osmate
gave the conditions for its quantitative precipitation with
tetraphenylarsonium chloride.
The formation of thiocyanate complexes, their reaction with
tetraphenylarsonium chloride, follo1;ed by their extraction into a
suitable organic solvent, form the basis of a large number of
(28) . successful metal determinations. Thus cobalt may be est1mated
spectrophotometrically by extracting t)le tetrathiocyanatocobaltate(II)
complex into chloroform. This method has been extended to the
determination of cobalt in steels. Affsprung and Murphy(29)have
developed a spectrophotometric method for determining tungsten in
steels and alloys, by extracting (Cc6H5\As+J (1-I(OH)2(SCN\-)
into chloroform. The absorbance of the extract is measured at
460nm. The method is selective, under the conditions described, for
tungsten after niobium has been masked with fluoride.
An improved method over the procedure publishad(29)is
recommended by Fogg, Harriott and. Burns. (30) In this method
quinol is added to the chloroform used for extraction to prevent
the formation of the red oxidation product, and titanium(III)
chloride is used to complete the reduction of the tungsten(VI).
An alternative procedure is described for the calorimetric
determination of molybdenum in steel by p'ogg et al. (31 ) In this
method molybdenum~I) is reduced to molybdenum(V) with ascorbic
acid and titanium(III) before reacting with thiocyanate and
24
extracting it with tetraphenylarsonium chloride into chloroform
that contains quinol. The procedure is reported to be sensitive
-1 -1 4 ) (t- = 17400 1 mol cm at 70nm • max in this method up to forty
fold excess of tungsten over molybdenum did not interfere.
Tetraphenylphosphonium chloride has been suggested by Tribalat(3Z)
as a means of separating and detecting traces of rhenium in neutral
ores, as the chloroform soluble perrhenate. Neeb(33)has determined
iridium and osmium absorptiometrically by the solvent extraction
of their hexachloro. complexes with tetraphenylphosphonium chloride.
Analytical procedures involving the use of onium compounds
appear not only to compare favourably with other methods but in some
instances have distinct advantages. For example when tungsten( 29)
is estimated by extracting its thiocyanate complex as a tetraphenyl-
arsonium salt instead of as the free acid, not only is the colour
of the extract much more stable, but the
is less critical. Also when iron(34)is
pH of the aqueous phase
extracted as the
triphenylmethylarsoniumhexathiocyanatoferrate(III)complex, similar
advantages are obtained.
The presence of the triphe~ylmethylarsonium ion confers stability
on the coloured anion. The method for manganese(35)based on the
extraction of the permanganate ion with tetraphenylarsonium chloride,
is highly sensitive and is reported to be more accurate, and more
reliable than any other permanganate absorptiometric methods.
Although the reactions of the tetraphenylarsonium and
tetraphenylphosphonium ions are very similar, the tetraphenylarsonium
salts are reported(3G)to be more soluble. There appears to be no
particular analytical advantage to be gained by using the
25
tetraphenylarsonium and tetraphenylphosphonium salts. (19). Further
the costs of both tetraphenylarsonium and tetraphenylphosphonium
chlorides are similar, although the tetraphenylphosphonium chloride
is synthesized more easily.
In recent years an increasing interest has been shown in
extraction systems involving quaternary ammonium, phosphonium,
arsonium and stibonium salts. These investigations have been
largely concerned with the use of quaternary cations to extract
anionic metal complexes.
The extraction of quaternary phosphonium and arsonium salts
from aqueous solution into the organic solvents, chloroform,
dichloromethane, 1,2-dichloroethane and 2,2'-dichlorodiethylether
over a range of cation and anion concentrations has been reported
by Gibaon and Weatherburn(3?). They have concluded that, under
the conditions studied the quaternary salts exist as ion pairs or
as dissociated ions in the organic phase. The extraction of
triphenylmethylarsonium chloride from hydrochloric acid solution
was anomalous; the distribution ratio reaches a maximum at about 2M
hydrochloric acid and decreases rapidly with increasing acid
concentration. This behaviour is attributed to the formation of ion
pairs of the quaternary salt in the aqueous phase.
The analytical chemistry of the iodonium salts has also been
the subject of a study of Bowd and Thorburn Burns. (38-44) One of ,
the aspects of these salts which they considered,was the alleged
similarity of the diphenyliodonium and thallium(!) cations. <45). As
the initial investigation into this comparison revealed that the
similarity was rather superficial, Bowd and Burns decided to engage
26
in a much wider investigation, which has proved of considerable
interest. One important result from this work was that the
diphenyliodonium salts were shown to be prepared conveniently in
the laboratory by the method proposed by Beringer et a1.<46 l
provided that particular care is taken in the preparation and
purity of the reagents and in the careful control of the reaction
temperature,< 44>. Higgens< 47)studied the certain aspects of the
analytical chemistry of diphenyliodonium ·chloride, and separated
the diphenyliodonium ion in the presence of several other "onium"
ions by chromatography and by electrophoresis. The possible uses
of diphenyliodonium chloride as an· analytical reagent was also
investigated. Spectrophotometric determination of cobalt by
formation and extraction of diphenyliodonium tetrathiocyanato
cobaltate(II) in the presence of high concentration of iron was
developed.
Triphenylmethylarsonium chloride has been studied by Gibson
and White<48- 49&ho have reported a series of titrations with
extractive end-points, in addition to calorimetric applications.
The titrations are interesting in that they obviate the need for
redox indicators to detect the end-point of certain reactions and
allow the titration of highly coioured and turbid solutions.
The triphenylmethylarsonium cation forms highly coloured chloroform
soluble compounds with permanganate( 4S)and dichromate (49). These
extractions have been used as a basis for the extractive detection
of the end-points in titrations involving the use of these t1~o
oxidizing agents.
A.J. Bowd<44 lalso re-examined these titrations using diphenyl-
iodonium chloride as the extractive reagent. Only partial success was
achieved, because high but reproducible blank values were obtained
compared with those obtained with the arsonium salts. In the present
27
work the possible use of diphenyleneiodonium bisulphate as
alternative to the other "onium" salts is justified for·two
reasons:
a) diphenyleneiodonium bisulphate can be prepared readily
and cheaply in the laboratory and in a high stage of purity,
b) t'here was the possibility that diphenyleneiodonium
bisulphate and several different anions have been investigated
with a view to developing quantitative procedures •. The reactions
between this onium compound and a number of oxyanions proved to
be disappointing.
3) Basic Dyes
The history of the dyeing industry is very old. The only
natural basic dye at that time was berberine, a yellow dye derived
from the barbery shrub.(50)
In 1771 Woulfe prepared picric acid by the action of nitric
acid on indigo, and showed that it dyed silk in bright yellow
shades. Laurent in 1842 converted phenol into picric acid. It
was not until 1856 that the synthetic dye industry got under way
with the discovery of Mauve by Perkin. This was the first
synthetic dye to be manufactured in any quantity. Many Rhodamines
and Thiazines and Triphenylmethane dyes were discovered during the
period of 1870 to 1890 along with many other basic dyes. (51 ~ Dyes
may be categorised in a number of different ways: by their origin,
their dyeing properties or by their chemical structure. The group·
of dyes known loosely as the basic dyes were the earliest known
organic synthetic dyes. They are characterised by exceptional
brilliance, high tinctorial strength but low fastness to ·light.
----- -------------------------------
In the analytical field, dyes are used in a number of ways:
as indicators, as titrimetric reagents, for spot tests, as
extractants and chromogenic reagents.in solvent extraction
procedure and a few miscellaneous cases. Many dyes find applications
as pH indicators (52) They are used either singly or as mixtures,
which exhibit an easily detectable colour change over a given pH
range. For example the dye Methyl Red and Phenolphthalein are
well known indicators for acid-base titrations. A review of
the literature has shown that many workers have found basic dyes
to be useful in determining large anions, particularly those of
the P-block elements, as ion-association complexes. A classification
and study of the basic dyes has been given by Willcox (53)··
According to this classification, basic dyes were assigned to
the following nine groups named: Diphenylmethane dyes,
Triarylmethane, Xanthone, Acridine, Azine, Oxazine, Thiazine,
Antipyrine and th~ newer cationic dyes.
According to Blyum and Pavlova,< 54) the reagents most widely
used in analytical chemistry are based on two groups of basic dyes:
a) Triphenylmethane dyes (structure a)
b) Xanthone-rhodamine dyes'(structure b)
+
·x
8
29
------------------------------
a b
Dye A B Dye A B
1 N(cH3
)2 H 1 N(C2H5)2 COOl!
2 N(CH3
)2 NHCH3
2 N(C2H5)2 COOC2H5
-
3 N(CH3
)2
N(CH3
)2 3 N(C2H5)2 COOc4u9
4 N(C2H5)2 H 4 NHC2H5
cooc2H5
a b
1) Malachite Green 1) Rhodamine S
2) Methyl Violet 2) The ethylester of Rhodamine 3 (Rhodamine 3B)
3) Crystal Violet 3) The butyl ester of Rhodamine S
4) Brilliant Green 4) Rhodamine 6 J
The dyes of these groups differ from each other only in the nature
of the substituents on the benzene rings, so they all will react
with the same anions. However. they are not entirely of equal value
as analytical reagents. An important difference in the dyes of
the second group is their ability to fluoresce, this can be attributed •
to the greater rigidity of their molecular structure arising
from the additional oxygen bridge between the benzene rings.
The extractive photometric methods for the determination of
·elements in the form of ionic associates with basic dyes was reviewed
by Blyum and Oparina,(55) and the use of Xanthene, Triphenylmethane
and other basic dye cations as reagents for the determination of
anions is reviewed by Fogg, Burgess, and Thorburn Burns (5G),
30
------------------------------~----------------------
The basic dyes can, under suitable conditions, form extractable
compounds with a large number of elements. In most cases however,
selective extraction of the element to be determined can be
achieved relatively easily by careful selection of appropriate
solution conditions. Thus formation of an anionic complex of an
element which is reactive with respect to the dye, and hence which
is capable of producing a useful compound containing the dye cation
and the complex anion of the element to be determined, is only
possible if the solution contains an appropriate anionic ligand.
The extractabilities of the salts of cationic dyes vary with
the pH of the solution. Each compound can be extracted only within
a definite range of pH, whose limits depend on the nature of cation
and of the complex anion. The greatest selectivity is achieved by
extracting the element to be determined from a solution containing
the maximum permissible concentration of hydrogen ion.
The purity of the extractant is a necessary condition for
stability of the extract colour. The presence of traces of fatty
contaminations such as grease from the tap of a separatory funnel
can lead to rapid destruction of the coloured compound, particularly .
if the element extracted is easily reduced to a lower valency state,
for example compounds of Tl(III), Sb(V) and others (54).
A necessary condition for formation of a compound between the
.dye cation and the complex anion of the element to be determined, is
the presence of at least a small amount of the requisite form of
the dye in the solution. This requirement determines the pH
range over which any particular dye can be used for analysis. The
concentration of singly charged dye cations decrease with
31
increasing concentrations of hydrogen ion. The upper limit of
hydrogen ion concentration,at which a given dye can still. react with
a given anion, increases with the increasing stability of the
compound between the cation and the complex anion of the element
to be determined.
On formation of the ion association complex behreen the dye
cation and the complex anion, the interactions of the participating
cation and anion with the solvent (water) dipoles become much
weaker, and this leads to transfer of the ion association complex
to the solid phase and means that it can easily be extracted by a
(54) solvent of low polarity
The first ion association re.action of a basic dye with an
anion was reported by Eegril<e in 1927 (5?). Although the mechanism
of the reaction was not indicated, he described a sensitive test
for antimony in which antimony(III) was oxidised with nitrite to
antimony(V) and Rhodamine B was added. A trace of antimony gave
a characteristic violet or blue violet colour, 50j1g of antimony
could be detected in the presence of 12,400 times as much tin.
Tungsten(VI) at the 50j1g level gave a similar reaction to antimony.
Although the basic dyes are generally very sensitive analytical
reagents, they are not always very selective. Some improvement in
selectivity can be achieved by careful choice of solution
conditions (the type of buffer or acid used) in addition to the
choice of dye. In general, an increase in the molecular weight of
the dye cation will increase the solubility of the ion-associate
in the organic phase, but decrease the selectivity.
32
Triphenylmethane dyes reported by Burgess(5S)to be more
selective than the xanthene dyes, His main field of study was
the triphenylmethane dyes, which resulted in the development of
several improved methods for the determination of such ions as
perchlorate, perrhenate, antimony and thallium.
In some cases, selectivity within one group of the periodic
table may be achieved by varying the oxidation states of the ions.
For example gallium, thallium, and indium only react with Rhodamine
dyes in the(III)oxidation state, but because of the varying
stability of the oxidation state within the group, selectivity can
be achieved by the use of a reducing agent. Thus gallium, which
is stable as gallium(III) but not as gallium(I), can be determined
in the presence of thallium(III), by reducing the thallium to the
+I oxidation state by the addition of titanium(III) chloride.
Willcox( 53)investigated four types of dye, with respect to
their analytical uses: The Rhodamines, Methylene Blue and its
related compounds, Brilliant Green and Crystal Violet. An extensive
study also has been performed on Methylene Blue and its 1,9
Dimethyl derivatives, Taylor 1 s Blue, and a method for the .
determination of perchlorate using Taylor's Blue is proposed.
The Butylester of Rhodamine B was prepared, its analytical
properties were studied. A method for the determination of
chromium using Butyl Rhodamine B was suggested.
Recent work on the analytical
Brilliant Green< 59)showed that the
uses of basic dyes particularly
results obtained in the
determination of gold, varied considerably depending on which
commercial sample of Brilliant Green was used. Impure samples of
33
Brilliant Green gave lower absorption readings, but by using pure
batches, good results were obtained. In this method the determination
of perrhenate-Brilliant Green ion-association complex extracted
into toluene was unaffected by reagent purity. Very little work
has been done on the purification of basic dyestuffs. Burgess (58)
has investigated the purification of basic ~e stuffs and found
that, re-crystallisation was not satisfactory, as it was very
wasteful of dyestuffs and the liquors formed were difficult to
filter. A simple and effective method for dyestuff purification
is given by the same author. The method is a continuous hot
recrystallisation process using a solvent which saturates at a low
concentration of dyestuff. This method was applied successfully
to a number of dyes, including Safranine o, Rhodamine B, Victoria
Blue 4 R, and Sevron Red GL.
The use of basic dyes in making ion-selective electrode has
been examined by several authors. Entwistle and Hayes ( 60~ 1•ere
among the first to make successful use of basic dye salts in
electrode technology with the introduction of a Methylene Blue
uranyltribenzoate electrode for the determination of uranium.
Because of the selective formation of basic dye association complexes
with particular anions and their complete extraction in hydrophobic
solvents, there is a strong indication that they may be of great
potential for use in selective-ion electrodes. A Brilliant Green
tetrathiocyanatozincate(II) ion-selective electrode has been used
for the determination of zinc by Fogg, Duzinkewycz and Pathan (61 l.
In this work, liquid state, heterogeneous silicone· rubber and carbon
paste electrodes based on Brilliant Green tetrathiocyanatozincate(II)
were prepared and studied. They reported that the most satisfactory
34
electrode was the liquid state electrode, consisting of a
10-3M solution of Brilliant Green tetrathiocyanatozincate(II)
in o-dichlorobenzene supported on lightly cross-linked natural
rubber. Brilliant Green has also been used as liquid-state
(62) ion-selective electrodes for the determination of perchlorate ,
and tetrafluoroborate(G3)based on their water-insoluble Brilliant
Green salts. Liquid-state electrodes based on the hexachloro-
antimonate(V) and tetrachlorothallate(III) salts of Sevron Red L,
Sevron Red GLand Flavinduline 0 are given(64lfor determination of
antimony and thallium.
35
. I
CHAPTER II
EXTRACTION AND SPECTROPHOTOHETRIC DETERHINATION OF CHROHIUM(VI) WITH TRI-N-BUTYLBENZYLPHOSPHONIUM
CHLORIDE
2.1.Introduction
The direct calorimetric determination of chromate or
dichromate cannot be compared in sensitivity with the well-known
diphenylcarbazide reaction, but the latter suffers serious
interference from iron(III), molybdenum(VI), mercury(II),
vanadium(V) and large concentrations of highly coloured cations.
In addition the colour is not stable and fades very quickly (65).
Concentration of the chromate into a small volume of a suitable
solvent should compensate for this main disadvantage of lower
sensitivity. Bernhardt (66) extracted the chromium - .1 ,5-
diphenylcarbohydrazide complex with 1-hexanol or cyclohexanol,
but all the metals that interfered with the usual aqueous colour
development remained troublesome. Dean and (67) Beverly suggested
a rapid, selective, and accurate calorimetric determination of
chromium based upon the solvent extraction of Chromium(VI) from •
aqueous 1Hhydrochloric acid with 4-methyl-2-pentanone and the
development of the magenta colour of the chromium-1,5-diphenyl-
carbohydrazide complex in the extract •
. The absorbance of the colour was measured 15 minutes after
mixing at 540nm. The optimum concentration range extended from
1.0 to 10.0/'g of chromium in 6ml. of solution. Very large amounts
of iron and moderate amounts of vanadium offered no interference.
36
This method is particularly applicable to cast iron and steel
samples whose chromium content is low.
The solvent extraction of the ion-pairs of c.hromium(VI) an-
ions with organic cations has been described by several authors.
It was found( 68 )that the triphenylsulphonium cation was extracted
Cr(VI) 2-as Cr2o7
by chloroform from aqueous solution.
Tetraphenylphosphonium compounds were suggested for determination
of Chromium(VI) as Cr2o7
2- by Book and Jainz (69); chloroform was
used as an extractant and at below pH 6 the chromate showed more
than 9~/o extraction. As Zeigler(70)reported, triphenylselenium
salts have properties analogous to those of oxonium and sulphonium
( 2- [ J salts. Chromium as cr2o7
) can be extracted as (c6H5)~J 2cr2o7 into dichloromethane from aqueous solution at pH 1 to 2 and can then
be determined photometrically at 362 or 445nm.
Solvent extraction of chromium(VI) by various amines from acid
solutions where obviously ammonium cations took part in.the
(71) extraction process has been described by Shevchuk and Simonova
2-These authors reported that Cr2o7
was almost completely extracted
from aqueous solutions, 0.15N to 3N with respect to sulphuric acid,
by solutions of tridecylamine, dtdecylamine, dihexylamine and
tribenzylamine in organic solvents. Complete extraction \1as
achieved with 0.1M tribenzylamine in chloroform, dichloroethane,
cyclohexanol. or tributylphosphate.
A solvent extraction method for the radiochemical determination
of chromium has been described by Maeck, Kussy and Rein (73) . This
method is based on extraction of the chromium(VI)tetrabutylammonium
ion-association complex into methylisobutylketone. Oxidation of
37
-- - - - - - --------------
chromium(III) to chromium(VI) with divalent silver prior to
extraction was investigated, the oxidation was rapid but because
of the formation of silver chromate and silver chloride in subsequent
steps, the reagent was unsatisfactory. Hexavalent chromium is
conveniently obtained by fuming chromium(III) with a mixture of
sulphuric and perchloric acids.
Although chromium(VI) may be extracted directly into
methylisobuty1k.etone from 1M or higher concentrations of hydrochloric
acid, a quaternary ammonium salt is added to ensure complete
extraction over a wide range of acidity. 2-The divalent Cro4 . ion
does not form an ion-association complex with the quaternary
ammonium ion. Thus contact with sodium hydroxide solution completely
strips the chromium from the organic phase. It was found that
chromium could not be completely removed from methylisobutylketone
solution in the absence of the quaternary ammonium ion, this was
considered to be due to some type of solution mechanism.
A method is suggested by Chalmers and Dick(7~)for the systematic
separation and determination of 12 metals by a combination of
solvent extraction and spectrophotometric methods.fn this
procedure Cr(III) was oxidised t~ Cr(VI) with potassium persulphate
and then Cr(VI) was extracted into ethyl acetate at pH(2. The
absorbance was measured at 580nm against a blank.
The formation of blue perchromic acid represents one of the
most selective and fairly sensitive tests for the identification
of chromium. The coloured product decomposes rapidly (74~ but
if the blue oxidation product obtained by acidification of a chromate
solution with sulphuric acid and treatment with hydrogen peroxide
can be extracted into tri-n-butyl phosphate (27~ in benzene), the
. I
colour remains essentially stable for at least 48 hours. This ·
extraction method could be used for the detection of chromate at
a concentration as low as 5 x 10-5M. In this method the chromium
can be estimated either by back extraction into the aqueous phase
followed by volumetric determination or by direct measurement
of the intensity of the blue product in the organic phase. This
method was also reported to be free from interferences by large
quantities of Cu, Ni, Go, Cr, Fe, V, and small quantities of
Mo and Ti (?5-?6)
The highly selective extraction of chromate from slightly
acidic solution (0.1 - 0.2t~ sulphuric acid) with a chloroform
solution of trioctylamine (Alamine 336-s) or trioctylmethylammonium
chloride (Aliquat 336-S) is described by Adam and Pribil (??).
In this method vanadium(V) 1 uranium(VI) and molybdenum(VI) are
extracted but the addition of 10ml of saturated sodium chloride
solution prevents the extraction of vanadium and uranium and to a
lesser extent that of molybdenum. Small amounts are coextracted
and give a positive interference. Molybdenum can easily be removed
from the extract by washing ~ti th dilute ammonium oxalate solution.
Arsenic, antimony and selenium do not interfere: .. if the acidity is •
properly adjusted before extraction. Manganese(VII) is easily
extracted into trioctylmethylammonium chloride but the colour
fades rapidly. On extraction with trioctylamine, manganese(VII)
forms MnO(OH)2 • Manganese(VII) can be reduced selectively with
oxalic acid and alcohol before extraction. Tungsten(VI) interferes
but is normally removed, along ;rith niobium and tantalum, during
dissolution of the sample. rhis method is reported to be very simple
and selective enough for the determination of chromium in alloys,
39
slags, minerals, etc. The final determination of chromium is based
on the measurement of the absorbance of the extract at 445-450nm.
In the method suggested by Higgens ( 47 l; dichromate ion v1as
extracted from aqueous solution with tetrachloroethane as the
diphenyliodonium dichromate ion-association complex. Hydrolysis
of the dichromate was prevented by addition of acetic acid.
In the present work the solvent extraction technique has
been used for determination of dichromate ion (Cr2o7
2-) using
tri-n-bu;!:ylbenzylphosphonium chloride. It was thought, that
the extraction technique might form the basis of a spectrophotometric
method. The following work is concerned with investigations into
the method.
2.2.Experimental
In carrying out the investigation, particular regard was
given to the reproducibility of results, the conformity to Beer's
law, molar absorptivity, and the stability of the chromium(VI)-tri-
n-butylbenzylphosphonium system. The effect of using different
onium compounds as well as organic solvents was studied. Job's
method showed the composition of,the complex to be 1:2 ratio of
the chromiuml'tri-n-butylbenzylphosphonium chloride (TBBPC).
2.2.1. Reagents:
The initial investigation followed the procedure outlined
b H. (47)
y 1ggens
a) Standard potassium dichromate solution (M/200), M/100 with
40
respect to hydrochloric acid, was prepared by dissolving 0.7354g
of K2cr
2o
7 in M/100 HCl and diluting the solution with HCl (M/100)
to volume in a 500ml volumetric flask.
b) Tri-n-butylbenzylphosphonium chloride M/50 was prepared
by dissolving 0.6578 g of reagent in distilled water and diluting
to volume in a 100ml volumetric flask.
2.2.2. Procedure
To measured volumes of standard dichromate solution in a 100ml
separating funnel was added 6ml of TBBPC solution. Chloroform
(10ml) was now added, and after extraction the lower solvent layer
was filtered through a Whatman No. 1 filter paper into a 25ml
volumetric flask containing glacial acetic acid (2ml). This
extraction procedure was repeated twice more, using 8ml and· 5ml of
chloroform respectively and the solution in the volumetric flask
was made up to volume and thoroughly mixed. The absorbances of
the extracts were measured at 448nm in 1 cm silica cells against
pure chloroform. The calibration curve was rectilinear as shown
by the results in Table r.
TABLE I
Calibration results for a known volume of standard potassium dichromate solution(M/200)
Volumes of standard Cr2o
72-(ml) Absorbance
0 0.018 1 0.073 2 0.13b 3 0.223 4 0.305 5 0.364
41
The number of extractions required was checked as follows:
To 5ml aliquots of standard dichromate solution in a
separating funnel, was added 6ml of TBBPC solution and 10ml
of chloroform. The solution was mixed for 30sec. The organic
layer was filtered into a 25ml volumetric flask containing 2ml of
glacial acetic acid, and then the solution in the flask was made
up to volume with chloroform. The same procedure was applied to
the same sample and from the following results, was found that
even two extractions were sufficient.
TABLE II
Number of extractions reguired
Extraction Absorbance Hi
First 0.362
Second 0.017
Third o.oo
Total absorbance was found to be 0
2.2.3.Effect of different organic solvents
Preliminary studies were carried out usi
of H/200 potassium dichromate and H/50 TBBPC
solutions were shaken with equal volumes of v
solvents. The extraction of chromium(VI) was
by the transfer of the orange colour from the
phase.
42
xing Time
30 sec
" "
.379·
ng measured volumes
solutions. These
arious organic
qualitatively noted
aqueous to the organic
A series of quantitative extraction studies were carried
out on the most promising systems by treating a fixed amount of
chromium(VI) (5ml) with a measured volume of the TBBPC and
diluting to a convenient volume. These aqueous solutions were
extracted with equal volumes of different organic solvents.
The results are shown in Table III.
TABLE III
Effect of different organic solvents on the solvent 2-extraction of Cr2o
7 ion with Tri-n-butylbenzyl~
phosphonium chloride
Organic Solvent Absorbance Time (min) 1) 0-dichlorobenzene 0.482 0
0.470 5 0.451 10 0.438 15
2) 1,2 dichloroethane 0.376 0 0.376 5 0.376 10
3) dichloromethane 0.379 0 0.379 5
4) tetrachloroethane 0.359 0 5) chloroform 0.381 0
0.381 5 0.381 10
As shown in Table III, in the case where 0-dichlorobenzene was used,
higher absorbance values were obtained but the absorbance decreased
significantly with time. All the other solvents tested gave lower
absorbances compared with those obtained with chloroform, so
chloroform has been used throughout this work.
43
L_ _______________________ -
----------------------
TABLE IV
solvent
Equivalent to Abbrev- Absorbance Onium Compound M/50(w/v) iation .
1) (Methyl)-triphenylphosphonium 0.71% MTPB extraction was bromide not complete
2) Triphenyl-n-butylphosphonium 0.73% TBPB 0.347 bromide
3) Tri-n-butylmethylphosphonium o. 68",6 TB HPI easy_oxidation iodide
. of I to 12
4) Triphenylmethylphosphonium o. 70}{, MPC 0.352 chloride
5) Tetrabutylphosphonium 0.58% TBPC 0.362 chloride
6) 3,5-di-t-butyl, 4 hydroxy- 1.09;6 BHTPB • the complex benzyltriphenylphosphonium formed is more bromide soluble in
water than in chloroform
7) Tri-n-butylbenzylphosphonium o. 65",0 TBBPC 0.380 chloride
. • 8) Tris(3-chlorophenyl)methyl 1.01% TCMPI easy_oxidation ph~sphonium iodide of I to 1
2
• 9) Diphenyleneiodonium bisulphate DPIBS • 0.120 (a)
Compounds marked • were insufficiently soluble in water to produce 0.02M solutions and were therefore used as saturated solutions.
(a) The preparation method will be given in part 2.3.
44
L__ _______________________________ -- - - ----·---
-----------------------------------------
2. 2.4. :'rhe .effect
extraction
of different onium compounds 2-. of Cr
2o
7 ~nto chloroform.
on the solvent
Attempts were now made to form the ion-assocation complexes
between several other onium compounds and the dichroma.te ion and
to extract them with chloroform. Studies were carried out in
aqueous solutions using M/200 potassium dichromate and M/50 of
solutions of different onium compounds. The results are shown in
Table IV.
As is shown in Table IV, the highest absorbance v1as obtained
using TBBPC as reagent. So in the present work this reagent has
been used.
2.2.5. Conformity with Beer's Law
The same procedure was carried out as described in paragraph 2.2.2.
The absorbances of the extracts were determined at 448nm. The
blank value was found to be 0.018 1 mol-1cm-1 , and the absorbances
obtained were corrected accordingly. The linear plot obtained is
shown in Figure I, showing that the system obeys Beer's law with
0-10-3 M dichromate in the sample solution.
TABLE V
Conformity with Beer's Law
Volume 2-of Cr2o7
M/200 Absorbance
ml .
1 0.055
2 0.120
3 0.205
4 0.287
5 0.346
45
-------------
c u c
Fig.1
8t:?C?r's law plot for tht:? extraction of
tri_ n _ butytbenzyt phosphoni urn- dichromate?
into chloroform
0.5
0.4
~ 0 0.3 VI .0 <(
0.2
0.1
1 2 3 4 5 ml cro2-CM/200l 2 7
-------------------------------------------------------------------------------
2.2.6.Composition of the complex
Amounts between 0 and_10ml of M/200 potassium dichromate
were transferred to a 100ml separating funnel and s uitable amounts
of M/200 of tri-n-butylbenzylphosphonium chloride solution were
added so that the total volume of two solutions was ah1ays 10ml.
Each mixture was extracted with 10, 8 and 5ml of eh loroform and
the extracts combined and made up to volume. The a bsorbances of
the combined extracts were determined. The results are shown in
Table VI, and Figure (II).
TABLE VI
Composition of the complex.
VC O 2- (ml) VTBBPC(ml) Absorban r2 7
ce
1 9 0.072
2 8 0.152
3 7 0.216
4 6 0.226
5 5 0.198
6 4 0.160
7 3 0.136
8 2 0.087
9 1 0.042
From the results imum absorbance obtained, it is clear that the max
is obtained when 1 :2 volumes of potassium dichromat e and tri-n-
butylbenzylphosphonium chloride are mixed and extra cted into
chloroform. According to Job's method of continue us variations,
this corresponds to the formation of .1:2 complex:
46
0.3
Q1 u c 0
.0 0.2 <... 0 Ill
.0 <(
0.1
Fig.2
Job's plot for the extraction of TBBP=c~ o?
into chloroform
I 0
1 9
/ G
3 7
/........_
0 .
5
5
0
7 3
0
\
2.2.7. Stability of the complex with time
r.rime: 0 5 10 15 20 30
Absorbance: 0.333 0.336 0.386 0.379 0.334 0.380
From the results given, the complex has been shown to be stable
for up to 30 minutes.
2.2.8. Reproducibility of results for a fixed volume of standard potassium dichromate solution (M/200) at 448nm.
Volumes taken were 5ml for each determination and the
absorbances obtained were: 0.379, 0.381, 0.381, 0.334, 0.382, 0.381,
fl-d. No. of readings average value standard 99"/o confidence
deviation limit
10 0.3313 1.475 1.517
2.2.9. Molar absorptivity
The average molar absorptivity calculated from the sets •
of results given for the reproducibility test in the latter
paragraph was found to be 381 1 mol-1cm-1 •
2.2.10. Study of interferences
Suitable amounts of sodium salts of anions (except
in the case of ferrocyanide and ferricyanide where potassium salts
were added) that might interfere in the determination of dichromate
ions were added to solutions containing 5ml of M/200 ~· chromium
47
per 25ml final volumes. Absorbances were measured as described
earlier (2.2.2.).
It was found that I ions are easily oxidised, because the
reduction potential of the couple I 2/2I- is equal to 0.54 volt,
and the reduction potential of Cr(VI)/Cr(III) is·1.33. So this
oxidation-reduction takes place, especially in the presence of
acidic media which facilitate the reduction; and due to this
oxidation-reduction, very high absorbances were obtained.
Potassium ferrocyanide and potassium ferricyanide in the presence
of TBBPC, were extracted into chloroform, and erratic results·
were obtained. Phosphate ion, gives a complex that is more
soluble in water than in chloroform, so very low results were
obtained. In the presence of permanganate, ferrocyanide,
ferricyanide and iodide positive interference was observed; in
the presence of phosphate, thiosulphate and sulphite, the
interference was negative. The procedure was carried out as
mentioned in 2.2.2. and the results are given in Table VIII.
TABLE VIII
Study of the interferences of anions •
.Anion added Absorbance
- 0.37g . 2 B407 0.375
2-Moo4 . 0.380
C6H507 3- 0.373
2-As04 0.378
so 2-4 0.377
- 0.376 BF4 -Bro
3 0.377
48
Anion added Absorbance
BO 3-3 0.377
NO- . 3 0-379
AsO 3-3 0.371
Cl- 0.369
-Br 0.374
-Reo4 0.379 2-
C406H4 0.383
(CH2 - COOH)2
- 0.381
C204 2-
0.382
Sio3
2- 0.382
-Clo4 0.37&l
The following anions do not interfere below the concentrations
shown:-
!04 - less than 10-3M
WO 2-4 less than 1.32 X 10-3M
SCN - less than 2.4 x .10-~
N02 - less ,than 10-3M
The method which has been developed for the determination of
dichromate is an illustration of the potential use of the reagent.
As neither chloride, nor fluoride anions, nor to a certain
extent molybdate and tungstate (up to 1.32 x 10-3M), interfered,
it was thought that the method might be suitable for the determination
of chromium in certain steel samples, alloys and ores. Therefore
the interferences of vanadium(V), manganese(VII) and iron(III)
49
was studied. In particular the possible methods for the
determination of chromium(VI) in the presence of manganese(VII)
was studied and particular attention was given to the reduction . 2-
of Hno4- in presence of Cr 2o7
•
2.2.11. Reducing permanganate (Hno4-) in the presence of
2-dichromate (cr2o7
) using sodium azide.
The method carried out is the similar pr ocedure given
by Blundy ( 121 ).
Procedure: Pipette out 5ml H/200 Cr2
o7
2 into a conical
flask, add 2ml of 2.5M sulphuric acid and different volumes
of KMno4.H/200. Bring the volume to 15ml with water. The
Erlenmeyer flask was immersed in boiling water for 25 minutes,
removed and cooled to less than 10°C. To the solution was
added 2% (w/v) sodium azide (addition must be done under fume
cupboard) dropwise with swirling until the colour of potassium
permanganate is discharged. The solution is transferred into
a 100ml separating funnel, add 6ml of TBBPC·and extract filter
with 10, 8, 3ml of chloroform. Filter into a 25ml volumetric
flask containing 2ml of glacial acetic acid. The absorbance
was measured in 1 cm silica cells against chloroform. The
results are as follows:
Molarity of solutions in final solution absorbance
10-3M 2-cr2o7
. 0.382
-3 2-10 H Cr2o7
plus 6 X 10-5M Hno4 - 0.335
10-3H cr2o7
2- plus 3 X 10-5H Mno4 - 0.386
10-3H Cr2o7 2- plus 8 X 10-5H Hno4
- 0.392
From the results, no interference from permanganate on dichromate
2- -was observed if the ratio of Cr2o7
:Mno4 was 16.6. At higher
ratios interference was obtained.
The effect of different onium compounds Table IV as ,1ell
as different organic solvents Table III on the separation of
- 2-Mn04 from Cr2o7 was studied, but none of these attempts gave
satisfactory results. The method of Richardson(7S)was also tried.
In this method, manganese(II) was oxidized to permanganate using
potassium metaperiodate, which afterwards >las extracted into
chloroform as tetraphenylarsonium permanganate and measurements
were made directly at 532nm against chloroform. In the present
method Mno4- was used directly, but due to the reaction between
tetraphenylarsonium and Cr 0 2- (?9)as well as Mno4-, erratic
2 7
results .were obtained.
The interference effects of iron(III) and vanadium(V) were
also studied and from the results obtained, vanadium(V) was seen
to give positive interference, but iron(III) up to 4 x 10-4
molar was without effect.
2.3. Preparation !1ethod of Diphenyleneiodonium bisulphate
2. 3.1. Preparation Method of Peracetic acid, oxidation and cyclization
·All preparations and reactions with organic peracids should
(80) be carried out behind a safety shield
Aliquots (25ml) of hydrogen peroxide 25-30/o was added slO\dy
from a dropping funnel to 100ml of acetic anhydride >lhile the
temperature of the reaction mixture >las maintained O.:t 0°C by cooling
51
in an ice bath. The reaction mixture was kept at 0°C until the
solution became homogeneous, then was left overnight at room
temperature.
A solution of 2-iodobiphenyl·was made by dissolving 20g of
this iodo compound in 40ml acetic anhydride. 2-lodobiphenyl
solution was added to 100ml of peracetic acid solution at 0°C
in a 500ml three necked round bottomed flask equipped with a
thermometer and a motor driven stirrer, and left to stand
overnight at room temperature. The reaction mixture which now
contained the iodoso compound was cooled in an ice-water bath
and to the cold stirred solution 20ml of concentrated sulphuric
acid was added dropwise (mixture stirred for four hours at 0°C
(81) and then allowed to stand overnight) at room temperature •
A standard solution of NaHS03 was added to this slurry to destroy
the excess of peracetic acid remained in the solution. The
addition of NaHso3 was stopped when the starch-iodide paper did
not change colour. The solid biphe.nylene 2-2' iodonium bisulphate
was collected and washed with water and subsequently with benzene.
The sample was dissolved in hot concentrated formic acid and left
to crystallize, then filtered off, washed .with formic acid and left
dried in vacuum desiccator. Melting point: 248 - 250°C; yield 80'~.
From the results obtained from infra red spectroscopic determination
it was shown that even we could not obtain I peaks, but because
of the starting material that contain:ed I in 2 position, it was
obvious that after cyclization and oxidization procedure, diphenylene-
iodonium bisulphate was prepared. The results obtained were checked
by Lee's(81
)method for completion of the ring closure, and also
2-the infrared spectrum of this compound sho;red a peak due to so4
•. ·
52
2.1t. Conclusion
The investigation into the formation and extraction of
tri-n-butylbenzylphosphonium-dichromate revealed that the
dichromate in the complex was sufficiently stable ih the chloroform
with respect to time. In a further study conditions were found
which enabled the dichromate ion to be extracted from acid
2-solutions as the TBBP-Cr2o7 ion-association complex.
A spectrophotometric procedure was developed for the
determination of dichromate ion in which some glacial acetic acid
was added to the extract to prevent the conversion of dichromate
to chromate. The linear plot obtained sh011ed that the system
obeyed Beer's law. In this method reproducible results were
obtained as well as low standard deviation. Compared to the molar
absorptivity values given by Katz ( 193)and Vogel {1 25·)(398 1 ml-\m - 1
and 371 1 mol-1cm-1 respectively) for the spectrophotometric
determination of dichromate, the value obtained in present procedure
(381 1 mol-1cm-1) was not an improvement on the existing methods.
For this reason attempts were made to adapt the solvent extraction
procedure in an atomic absorption spectrophotometric method.
53
_j
CHAPTER 3
DETERMINATION OF CHROMIUM IN STEEL BY ATOMIC ABSORPTION SPECTROPHOTOMETRY USING AN AIR-ACETYLENE FLA}!E
3.1. Introduction
The use of atomic absorption for analysis was first
suggested by Walsh(B2) in 1955 1 and since then this technique has
become one of major importance to the analyst.
Atomic absorption spectroscopy is concerned with the
measurement of radiant energy absorbed by atoms of various elements.
Each element in the atomic state absorbs only radiation of
characteristic and well-defined wavelengths. In the process of
absorption the atom becomes excited, and this is the reverse
physical process to that involved in flame photometry •. Although
interference of one metal with another, or interference of excess
acid upon a metal's absorption, does occasionally occur, atomic
absorption spectroscopy is in general remarkably free from such
effects. Perhaps the most inconvenient and difficult interference
to overcome among the determinations commonly demanded is that
of iron upon chromium. This interference is particularly •
important in the determination of chromium in steels, ores, and
minerals, since a large ratio of iron to chromium is normally
present in such samples.
In atomic absorption spectrophotometry, sensitivity
for chromium in the air-acetylene flame is critically dependent
upon the flame stoichiometry. Fuel-rich flames produce highest
sensitivity, but interference is observed from Fe, Ni, and other
elements in these flames. These effects are minimized in a
54
leaner flame and it may therefore be necessary to sacrifice
sensitivity to gain freedom from interferences. Interferences
in the rich flame, however, may be suppressed by the addition
of ammonium chloride(83l, ammonium bifluoride(84lto sample and
standard solutions. A number of procedures have been suggested
for the determination of chromium in steel by atomic absorption
spectrophotometry using the air-acetylene flame, and either the
addition of a releasing (83-85-86) . . (87) agent or matrJ.x matchJ.ng
or both are used to overcome interference effects. Ammonium
chloride appears to function as a protective agent and the
addition of relatively high concentrations of ammonium chloride
to sample solutions has been successful in minimising the
interferences of iron in the determination of chromium.
Ammonium chloride has also been used in the determination
faeces (88 ) using an air-acetylene flame. of chromium in
The sample of faeces was dried and ashed and the Gr2o3
was
oxidized to Cr2o72- with HClo4• ·An aliquot (1ml) of solution was
diluted to 100ml with 2% aqueous NH4Cl, and aspirated into the
burner. In this method a rectilinear response was obtained with
up to 6j'g of chromium per ml. Giammarise(83)proposed the use of
strontium chloride or ammonium chloride in the solution sprayed.
This overcame the interference of 1000/B per ml of iron at the·
3-~g per ml. level of chromium. (85) Barnes observed that both
aluminium and ammonium chloride decrease the depressant effect of
iron on the absorption of chromium at 357.9nm. Ammonium chloride
(~la w/v) appeared to be particularly effective in this respect.
Flame conditions in this method were strictly controlled, and
an analytical procedure was suggested for the determination of
55
chromium in low alloy steels, and in certain types of iron. Iron
was not added to the chromium standards and the method appears
to be the simplest and most rapid developed to date. This reagent
. (86) at a concentrat1on of 25g/l has been used by Gomez et al
to overcome the interference from iron. After dissolving the
steel sample in an acid mixture (75ml HC104 , 55ml HN03
and 20ml
HCl) the solution was evaporated until dense fumes appeared, and
then the residue was dissolved in 50ml of hot distilled water
and diluted to 100ml. After addition of 2.~~ammonium chloride
solution, the chromium content of the solution was measured by
atomic absorption spectrophotometry. In this method a low
temperature flame was used. A composite scheme for the analysis
of steels by atomic absorption spectrophotometry using an air
acetylene flame was given by Nall, Brumhead, and Whitham. <89)
In this method the steel samples were dissolved in hydrochloric
acid, and oxidized by nitric acid. The solution was then evaporated
to dryness and redissolved in hydrochloric acid. Iron was added
to the standard solution. Ammonium chloride was used to overcome
interferences from iron using an air-acetylene flame. Kinson et
al(90)determined chr~mium by using an air-acetylene flame. The
effect of different solvent acid~ was examined. In this method
iron caused a major reduction in absorption sensitivity but there
was no significant reduction in absorption sensitivities when
hydrochloric,phosphoric, sulphuric or phosphoric-sulphuric acid
media were used. A mixture of suulphuric and phosphoric acids
was selected because of its advantage in retaining tungsten and
hydrolyzable elements in solution, and then the solution was
oxidized with nitric acid. The most useful flame type was
found to be a slightly rich mixture(10.4 1 of air per minute
and 2.4 1 of acetylene per minute at S.T.P. with the absorption
path 8mm above the base of the burner flame).
In some cases interferences can be overcome by using
interference suppressors. Hurlbut et al( 91 )studied a wide
range of interference suppressors and absorption enhancers for
chromium analysis. They studied the effect of the following
suppressing agents: sodium sulphate, sodium sulphite, potassium
sulphate, potassium persulphate and ammonium chloride. The
recommended suppressing agent for interference in the atomic
absorption spectrophotometric determination of chromium(VI) by
1000 ppm of Ba, Ti, Mo, Ni, Mn, W, Fe, Co, Cu, Ag, Cd, Hg, Al and
Ce was found to be sodium sulphate. However in this method
sodium sulphate did not suppress interference caused by 1000 ppm
Mg or Ca. Sodium sulphate was also reported to be an effective
suppressing agent in chromium determinations in industrial
waste water and sediment using an air-acetylene flame.(92)
Although this reagent suppressed interference from Fe, Ni and Co,
Mg interference had to be suppressed by addition. of ci trate. Elrod
end Ezel( 93)reported a determination of chromium in chromium-
alumine catalysts, aluminium metals and ores, in which samples
are decomposed with 50% H2so4 or H2Soz/HNOjHCl. •
The addition
of 1% potassium persulphate (K2s2o8 ) was recommended to permit
accurate determination in the presence of large quantities of Al,
Fe and Ti, eventhough aqueous chromium standards were used as
reference solutions. Vogliotti( 94)used solutions of 0-20g/l
potassium pyrosulphate (K2s 2o7) to control the interferences from
0.02 - 2ppm Al, Co, Cu, Fe and Ni in the determination of 5ppm
of chromium. Chromium in environmental samples such as water
and soil was determined by oxidizing chromium to Cr(VI) >Ii th
57
KMn04 and then extracting with MIBK containing 0.3% of
trioctylamine. The organic layer was sprayed into an air-
(95) acetylene flame. In this method large amounts of metals such
as Fe, Ni, Cu, Al, Zn, Pb and Mn did not interfere.
Roos(96), Roos and Price(9?) studied the mechanism of
interference and releasing action in atomic absorption
spectrophotometry. According to them, the enhancement by
releasing agents might occur because of the following possible
effects:
1) The possibility that the reagents added affect one or
more properties (e.g. temperature) of the flame. The effect of
the presence of large amounts of ammonium chloride or sodium
sulphate on flame properties were tested by fitting a second
nebuliser to the cloud chamber of the instrument. A solution
of chromium plus iron was sprayed simultaneously, but separately,
with a solution of sodium sulphate or ammonium chloride. Since
no enhancement of the chromium signal was observed, it was concluded
that any changes in the flame properties brought about by the
presence of the inorganic salts did not noticeably alter the
interference of iron in the determination of chromium.
2) The possibility that the reagents affect the subdivision
of the sample material after its introduction into the flame.
Addition of ammonium chloride, alkali chlorides and sulphates,
for example, have been shown to enhance chromium absorption in
the presence of iron. These substances are all relatively volatile
and evaporate or dissociate below 1000°C. Their action maY be
to bring about the volatilisation of all or part of the matrix,
58
with concomitant production of finely subdivided sample material
which can compensate for the low.volatility of the iron in the
sample.
3) The possibility that the releasing agent causes fractionation
of the sample material. The facilitation of the distillation of
impurities by the addition of a substance with a relatively low
boiling point (the carrier) was first demonstrated by Scribner
and Mullin(9S) and then investigated by Samsonova snd Strzyzewska.<99)
Carriers are considered to function by transferring impurities
to the vapour phase. If the distilling ammonium chloride vapour
becomes partially saturated with chromium and iron, enhancement
of both elements absorbances would be expected. However, since
the concentration of iron in the particle clotlet far exceeded
the concentration of chromium, a much higher proportion of the
chromium actually present in the particles would distill with
the ammonium chloride. Ottaway( 100) reported that 8-hydroxyquinoline
was a more effective releasing agent for metallic interferences
than were ammonium chloride, sodium sulphate or potassium
pyrosulphate. The disadvantage of this method was that the careful
control of fuel-flow was essential. The optimum flame was found
to be fuel-lean, because under fuel-rich flame conditions iron(III)
still interfered in this method. In the present work similar
results were obtained using 8-hydroxyquinoline as a releasing agent,
but it seemed that this reagent was very pH dependent.
Although for the detemination of low levels of chromium
in steels the air-acetylene flame offers the advantages of greater
sensitivity and reduced noise compared to the nitrous oxide-
acetylene flame, some workers have used this nitrous oxide-acetylene
59
( 101) . flame and reported good results. Compared Wkth the amount of
published work on the air-acetylene flame, comparatively little
work has been reported in which nitrous oxide-acetylene flame was
used for the determination of chromium in steel. The introduction
f t . .d t 1 1 . 1 . ( 102) . 6 o he nktrous ox1 e-ace y ene f ame by Wa l1s 1n 19 5
ushered in a new era in. atomic absorption spectrophotometric
determinations. Not only did this new flame allow the extension
of atomic absorption techniques to elements which could not
previously be determined, or which had a very low sensitivity in
the air-acetylene flame, but it was also found to completely
eliminate many interferences which had been very troublesome. It
seems an oversight that little work has been carried out using the
nitrous oxide-acetylene flame for the determination of chromium
in the presence of iron. Marks and Welcher( 103)in a study of
several elements including chromium in the nitrous oxide-acetylene
flame. observed an interference effect by iron on chromium. The
interference varied from suppression to enhancement depending
upon the excess of iron added. The addition of 5gfg per ml of
iron to 10fg per ml of chromium resulted in a suppression of 6%.
Upon increasing the iron from 200 to 5000~g per ml, the interference
manifested itself as an enhancem&nt varying from 1 - 9%. However,
it was stated that most interferences could be eliminated by proper
selection of flame conditions. . ( 101) Thomerso'!and Pr1ce . recently
proposed a method using the nitrous-oxide-acetylene flame without
a releasing agent. This produced acceptable results for a wide
range of steels (containing between 0.11 and 25.6% Cr) and
illustrates the greater freedom from interference in this flame.
It was·:necessary; however, to add iron to the standards to
compensate for the iron in the sample solutions. It is important
60
to use chromium metal and not potassium dichromate to make up
chromium standards, as potassium reduces ionization of chromium
in the nitrous oxide-acetylene flame in the standards only, thus
causing low results to be obtained for the samples. Although
there is a considerable depression due to iron in an air-acetylene
flameJin nitrous oxide-acetylene, iron enhances the absorption
of chromium, and this enhancement varies with the iron concentration.
Therefore, it is necessary to incorporate iron in the calibration
solutions and also add iron to solutions if dilutions are necessary,
in order to maintain a constant concentration of iron in all the
samples and calibration solutions. In this method, most of the
interferences are overcome by using perchloric acid as the solvent.
Tungsten still interfered in this method. The nitrous oxide
(104) acetylene flame was also used by Husler. • In this method
the alloy steels were dissolved in nitric acid and hydrofluoric
acid. Potassium was also added to all standards.
The papers just discussed are empirical procedures. l1ore
however, have been attempted. Yanagisawa theoretical approaches,
et al( 105)reported that chromium absorption was affected by various
cations for various flame conditions, and most serious interferences •
with chromium absorption have been found with fuel-rich flames.
They classified the interfering elements as follows:
a) Those which show an enhancing effect (Cu, Ba, Al, Mg, Ca),
i.e. those elements having higher heats of formation. The
enhancement may occur because these elements when present in large
amounts, compete with chromium in the formation of .volatile oxides,
so that production of chromium atoms is enhanced.
61
b) Those which show depressing effect (Na, K, Sr, Zn, Sn).
The formation of mixed oxides appears to be a likely source of
interferences from Na, K, and Zn.
c) Those which show an enhancing effect for Cr(VI), but
a depressing effect for Cr(III). Iron belonging to this category,
depress the absorption of Cr(III) and enhance the absorption of
Cr(VI). The depressing effect increased with increasing iron
concentrations, and may be derived from the formation of non-volatile
compounds with chromium such as chromite. Nickel also showed a
depressing effect on Cr(III) absorption/but no .notable enhancing
effect on Cr(VI) absorption. Sastri et al(106)suggested that
many inter-element interferences in atomic absorption spectra-
photometry may be due to the formation of mixed oxides. The
mechanism given is fairly general, and may not apply to the specific
interferences of iron.
The "organic solvent effect" of increased sample transport
to the flame may be usefully exploited by the solvent extraction
of the analyte element into a water-immiscible solvent. This
technique may also accomplish pre-concentration of the analyte
element (by extraction from a large volume of aqueous solution
into a small volume of organic solvent) when its concentration
in the original sample solution is too low to permit its
determination by atomic absorption spectrophotometry. However,
when large amounts of matrix element or other species which give
rise to chemical or physical interference are present in the aqueous
sample, a degree of selectivity in the solvent extraction procedure
may be useful for the elimination of these effects.
62
The extraction of an element from an aqueous solution into
an organic solvent, which was then sprayed into the flame was
first described by Dean and Lady (107>. Since then a number of
papers have reported the application of this technique to the
determination of various elements. The concentration of trace
metals present is too low to determine directly in aqueous samples
and so the organic solvents have been used to increase the
sensitivity.
(108) .. In 1957, Bryan and Dean after dissolving the samples
in acid, converted chromium into dichromate by oxidation with
potassium peroxydisulphate in the presence of silver ion.
The dichromate was then extracted from aqueous solution, molar
with respect to hydrochloric acid, with methylisobutylketone (MIBK),
by a single extraction. The organic phase was aspirated into an
oxyacetylene flame and the characteristic line emission of
chromium was measured. This method can be used in the presence
of large amounts of Fe(III), 1;hich is the only element that may
be extracted with chromium. In fact the introduction of the
organic solvent in place of water increases the emission intensity
of chromium fiftyfold when compared to aqueous solutions of equal
concentration. This method was applied to all types of steels
and cast-iron samples, aluminium alloys, monel metal, and clays.
According to Allan (109),in the determination of chromium
by atomic absorption spectrophotometry, when certain organic
solvents are added to aqueous solutions to extract an element,(zinc~
iron, manganese and copper) small increase in analytical sensitivity
was obtained. This increase in sensitivity obtained in their work
by spraying an organic solution, must be due entirely to an increase
63
in the amount of solution reaching the flame and to a temperature
effect. Of all the organic solvents tested, only esters, and
ketones were found to behave satisfactorily in the flame. With
these solvents the flame was steady, combustion complete, and no
absorption due to the solvent ~1as observed. Allan also studied
the mechanism of the absorption. According to him, the magnitude
of the absorption shown by an element sprayed into a flame depends
on:
a) . The concentration of atoms in the flame gases,
b) The length of the light path through the flame,
c) The width of the absorption line, which at constant
pressure is proportional to the square root of the
temperature.
The concentration of atoms in the flame could have been increased
in the follo,~ing ways:
1) by an increase in the amount of solution reaching the flame,
2) by lowering the flame temperature (giving smaller volumes
of flame gas) ,
3) by an increase in the r~te of vaporization of the metal
compound,
4) by an.increase in the extent of the dissociation of the
metal compound into atoms.
d o t p 0 (110) o t" f h o ;• 1 t" Accor Lng o rLce ,asp1ra 10n o c rom1um 1ron so u 1ons
will produce relatively large solid particles, which after
reduction by the flame gases consist of chromium (boiling point
2480°C) in a matrix of iron (boiling point 3000°C). These are
not completely vaporized and the atomization efficiency of the
64
chromium is low. This incomplete volatilization implies that,
at the temperature of the flame, the droplets produced by the
nebulizer have given rise to solid particles which, because of
their high vaporization temperature, their speed through the flame,
or both, are not completely converted to a vapour. The degree of
atomization is therefore lower then would be expected.
Coker et al( 111 )have studied the mechanism of atom formation
in flames. They have reported that in hydrocarbon flames the
ground state.population of atoms and the mechanism by which they
are formed are dependent both on the stoichiometry of the flame
and on the nature of the solution used. When other metallic
elements are also present in the same solution, interelement
interferences may result and these will probably also depend on
both the above.factors.
(112 113 114) It has been suggested ' ' that the role of the flame
as a source of metal atoms in analytical atomic spectroscopy is a
function of the chemical reducing properties rather than the
temperature, and so the flame cannot be regarded simply as a thermal
dissociation medium.
An attempt also has been made by Morris(11 5)to identify the
nature of the solid material formed when iron and chromium are
sprayed together into an air-acetylene flame by collecting the
particles formed in the flame and subjecting the sample obtained
to X-ray diffraction analysis, but the Tesults obtained were
inconclusive.
65
Fukamachi(116)extracted chromium(III) diethyldithiocarbamate
into MIBK. Chromium(VI) reacts with diethyldithiocarbamate(DEDTC)
within a few minutes at room temperature and at pH 4.5-7 (acetate
buffer) to give a Cr(III)-(DEDTC) complex, 1~hich is extracted into
MIBK. The optimum pH for extraction is 5.4. This reaction
takes only a few minutes at room temperature. For the determination
of total chromium, Cr(III) should first be oxidized to Cr(VI),
because complex formation between the Cr(III) salts present
initially with diethyldithiocarbamate is much slower, and further
selectivity will be obtained due to the fact that many elements
react in only one oxidation state. Thus Cr(III) has to be oxidized
to the Cr(VI) state. ·The extract was then aspirated into a
fuel-lean air-acetylene flame at 35'7.9nm. Chromium(VI) ((lppM ·)
was determined without interference from Cr(III), (present in
amounts less than the amount of Cr(VI)], Fe(5mg), Ag, Mg, Ca, Mn,
Ni, Cu, Zn,Cd, Hg, Pb, Al, CN-, or Po43-i but CO(II) caused
negative errors. A selective extraction procedure for different
oxidation states of Cr(III) and Cr(VI) in waste water is given
by Yanagisawa, Suzuki and Takeuchi (117).. Chromium(III) and
Cr(VI) traces were extracted into MIBK. at pH 6 or 4 respectively
after complexation of Cr(VI) with sodium diethyldithiocarbamate
or Cr(III) with 8-hydroxyquinoline or thenoyltrifluoro acetate,
prior to atomfc absorption spectrophotometric determination. In
this method both air-acetylene and nitrous oxide-acetylene flames
can be used.
Kono( 118)oxidized chromium in hot dilute sulphuric acid
solution with potassium permanganate. The excess of potassium
permanganate was decomposed with potassium bromide. The Cr(VI)
solution 0.5N and 2N with respect to potassium bromide and sulphuric
66
acid, respectively was extracted into MIBK, and determined by
atomic absorption spectrophotometry in a fuel-rich air-acetylene
flame. Interference from iron was prevented by the addition of
Na4P2o7
before extraction. The extraction increased the sensitivity
to O.O~g chromium per ml. at 1% absorption. (119) Feldman and Purdy
studied the optimum conditions and developed a sensitive method
for the determination of chromium by atomic absorption spectroscopy.
They showed that MIBK quantitatively extracted chromium(VI) even
at very low levels. The ketone extracts chromium that is present
in the hexavalent oxidation state, and if chromium(III) is present,
it must be oxidized to chromium(VI) prior to extraction. They
used potassium permanganate as an oxidant because the completion
of the oxidation is apparent when the pink colour persists. This
is an advantage of the atomic absorption technique over the
calorimetric technique. In the latter, it is necessary to destroy
any oxidant (without reducing the chromium) since the excess
oxidant will react with the reagent to be added for calorimetric
determination of chromium. But in the present case, extraction
of the chromium can be carried out in the presence of excess
permanganate without any adverse effects. This procedure also
has considerable potential advantage in increasing the sensitivity
of the determination, and is therefore of importance in the
determination of small concentrations of chromium. In 1967
Feldman et al( 1ZO) applied this method to the determination of
chromium in biological materials. In this method, blood, plasma,
.urine, and diet samples were wet ashed with nitric acid, sulphuric
acid, perchloric acid (3:1:1), and Cr(III) was then oxidized to
Cr(VI) with potassium permanganate and the Cr(VI) was extracted at
5°0 from dilute hydrochloric acid medium into MIBK. The organic
solution was then sprayed into the fuel-rich air-hydrogen flame.
In the present work Ce(IV) was used as an oxidant. This was
recommended by Blundy( 121 ) who in developing an extractive
colorimetric procedure for the determination of chromium in 1958,
made a study of the effectiveness of several oxidants for Cr(III)
and concluded that of those studied, ammonium hexanitratocerate
in hot acid solution gave complete oxidation and good reproducibility.
The atomic absorption spectrophotometry developed in this work is
based on :this :oxidation method. This work was undertaken to
investigate an extractive method for the suppression of interference
in the atomic absorption spectrophotometric determination of chromium
by use of sodium fluoride using air-acetylene flame. This study
is considered of importance, since the determination of chromium,
particularly in steel by this technique is an especially attractive
technique in terms of ease and speed of application, as well as
sensitivity. This is especially so if the interference by iron
can be eliminated, so that steel samples can be simply dissolved
and after extraction, sprayed into the flame.
The use and effectiveness of the previously.suggested
interference suppressants has been investigated, and sodium fluoride
was found to be as effective suppressant for iron. A simple and
effective method for the determination of chromium in steels has
been developed.
62
3.2. Procedure
3.2.1. Instrumental alignment, settings and the effect of burner height and fuel composition on the
determination of chromium
Before investigation of the interference·of iron on
chromium, it was necessary to find the conditions for optimum
sensitivity, with a simple aqueous solution.
Firstly, it is essential to ensure that the lamp beam
is correctly positioned along the optical axis of the instrument.
With the meter response switch in the "fast" position, as it was
during all peaking-up operations, the chromium lamp was gently
rotated in its clamp until a maximum meter deflection was obtained.
A piece of graph paper was placed on the photomultiplier lens,
to check the lining up of the lamp and photomultiplier by observing
the lamp image. Adjustment was made by altering the lamp clamp
position as necessary. The graph paper was then placed on the
burner slit, and the burner was aligned exactly \iith the lamp
image, by use of the burner rotation and traverse controls. This
final setting should also correspond with a maximum meter deflection.
V/hen this had been done, the flame was lit and the burner allowed
to warm up for at least fifteen minutes.
The monochromator wavelength was set at the value required.
This setting is not precise, and it was necessary to peak up on
the line by careful manual adjustment. When the precise line
wavelength was reached, the meter was at a maximum deflection, and
as the wavelength was changed beyond this setting in either direction,
this deflection decreased. By crossing the maximum deflection
using increasingly smaller movements of the control, the precise
69
wavelength was set. With practice, this peaking-up operation is
quite rapid.
The effect of changing each operating parameter on the
sensitivity was then systematically checked. With fuel composition,
lamp current and slit width set at the manufacturer.•s recommended
values for the determination, absorbances for a solution of 20fg
per ml Chromium(VI) were observed at a range of burner heights.
At the same time·, the absorbances at a range of burner heights
for the aqueous chromium(VI) solution as well as chromium(VI)
extracted into MIBK were tried.
Preparation of the sample
The·preliminaries in the preparation of the sample should be
kept to a minimum to avoid the risk of contamination with
extraneous sources of the element to be determined. The importance
of this is obvious if it is remembered that flame photometry is
a very sensitivie method of analysis, and that it is customary
to work with concentrations in the parts per million range. The
method of decomposition of the starting material should be the
simplest possible consistent with the need to keep the concentrations . . of certain species below a critical level, dilution of the sample
to the working concentration range should be done in the fewest
number of steps and be made with really pure water (usually de-
ionized water). Care should be taken to avoid loss by adsorption
on glass ware, and contamination by extraction from glassware.
The reagent used should be as Bure as possible and a reagent blank
should be run with the analysis. If organic solvents are used in
preparing the sample, their effect on the atomization rate, the
70
Fig.3
Calibration curve for chromium (VI)
extracted into MIBK
1. 0
• 0.8
01 u c: .B '- 0.6
/ 0 Vl
.0 <(
0.4
0.2
4 8 12 16 20 Cr(Vtl (fg/ml)
flame temperature, and the emissivity of the elecnent to be determined
must be taken into account. The sample solution should be free from
dust particles which could cause local variationS in the atomization
rate and, if incombustible, become incandescent in the flame and
emit transient continuous spectra. If preliminary separation
must be made to remove interfering elements, and an extraction
method is used, the influence of the reagents and the solvent on ·
the subsequent flame reactions must be considered. The organic
solvent chosen should not be appreciably soluble in water and also
must be directly combustible in the flame without the production
of a smoky flame or toxic products. Aromatic solvents such as
benzene or toluene generally give rise to the production of a smoky
flame which·may affect the analytical precision, while chlorinated
solvents such as chloroform and carbontetrachloride give rise to
(109) unpleasant and toxic combustion products. Allan has reported
that esters and ketones are generally satisfactory for use with
the air-acetylene flame, since they produce stable flames, are
completely burnt and do not contribute to the background
absorption of the flame.
Methylisobutylketone has been employed extensively for
solvent extraction followed by atomic absorption spectrophotometry.
In the present work also this solvent has been used.
3.2.3. Reagents
1) Stock solution of chromium(VI), (1000/ g per ml)
2.8282 g of A.R. potassium dichromate was dissolved in
distilled (de-ionized) water and made up to volume in a 1000ml
graduated flask, and the solution was stored in a polythene bottle.
71
2) ~larking solution of Chromium(VI) (20J'g per ml)
·An aiiquot (10ml) of stock solution was diluted to 500ml in
a graduated flask.
and made up weekly.
This solution was stored in a polythene bottle
3) Iron(III) .solution, (20,000)< g per ml Fe3+)
5.810 g of Fec13
was dissolved in water containing fe1~ drops
of concentrated hydrochloric acid. The solution ~<as diluted to
volume in a 100ml graduated flask, and 1~as stored in a polythene
bottle.
4) Ammonium hexanitranoeerate,. (0.02N)
10.965g of analytical reagent grade ammonium hexanitra.tocerate
~~as dissolved in water. The solution was made N with respect
to sulphuric acid when diluted to 1000ml in a graduated flask.
5) Sulphuric acid, (12.5% V/V)
6) Hydrochloric acid, (8 molar)
7) Hethylisobutylketone,
Equal volumes of methylisobutylketone and molar hydrochloric
acid were mixed thoroughly to saturate the organic layer with
hydrochloric acid (M). The layers were allowed to separate,
the aqueous layer run off and the MIBK layer passed through a
~atman No. 1 filter paper into a clean dry bottle.
8) Sodium fluoride, (20,ooo;te per ml)
11.5 g of NaF was dissolved in distilled water and made up
to 250ml in a eraduated flask,and the solution stored in a polythene
bottle.
72
9) Working solution of Chromium(III)
Aliquots of Chromium(VI) working solution were reduced by
passing so2
through the solution, and expelling the excess so2
by heating.
3•2 .4 Experimental
1. The effect of observation height on Chromium(VI) in
an aqueous and MIBK
The effect of flame height was tried in both aqueous and
organic phases because the flame possesses definite zones differing
in temperature, and reducing and oxidizing conditions, therefore
the number of atoms in a sample >~hich are in the ground state will
be different in the various zones. A maximum is thus expected at
a certain height in the flame. So 20Jig Cr(VI) per ml in aqueous
solution as well as fixed volumes of the same Cr(VI) solution
extracted into the same volume of MIBK to obtain the same concentration,
were sprayed into the flame. The acetylene flo>T >TaS 3cm, and air
flo>T was 12cm., and the results are given in Table IX. and Fig. V.
TABLE IX
The effect of variation of observation height
Burner height Absorbance of 20/"g per ml chromium(VI) mm . Aqueous Organ~c
1 0.58 0.58 2 0.2 0.23 3 0.09 0.33 4 0.05 0.'+3 '5 0.03 0.53 6 0.02 0.48 7 0.02 0.53
7.5 0 0.55 6 0.30 V.7'J
73
1.0
f:! c 0 .0 0.8 .... 0 C/1 .0 <(
0.6
o.4
0.2
Fig.4
Effect of flame compos-ition on the interference of
iron in determination of 20fg perm! Cr(VI)
2
a l Chromium (VI l ( 20f g per ml) extracted
into MIBK
b) Chromium( VI) (20tg permll plus 200001'9
3 .4
'
perm! Fe( IT! I
5 Acetylene flow rate (I per min.)
2 •. The effect of flame composition on the determination of Chromium(VI) in an organic phase (MIBK)
With all other parameters, including burner height,
fixed, the fuel composition was varied over a wide range, and
the absorbance at each fuel mixture noted (Table X). After
each solution, distilled ~1ater and HIBK were respectively sprayed
into the flame and the zero reset as necessary.
TABLE X
Effect of flame composition on the determination of Cr(VI) in MIBK
Acetylene flow rate Absorbance of 2Sfh per ml Cr(VI) (cm)
2 0.07 2.5 0.19
3 0.44
3-5 0.64 L
4 0.81 V.L.
4.5 o.8o v.L.
5 0.75 V.L.
L: luminous
V.L: very luminous
3. The Effect of variation of slit width
The effects of varying the slit width on the absorbance
of 20Jg per ml Cr(VI) in MIBK, when all other operating
conditions were at their optimum values were measured. The
results are given in Table XI.
74
•
TABLE XI
Effect of variation of slit width
Slit width mm o.oo8 0.009 0.010 0.011 0.012 0.015
Absorbance of 20 /' g per 0.49 0.49 0.50 0.48 0.43 0.43 ml Cr(VI) in MIBK
The settingsfor optimum sensitivity are given in Table XII
TABLE XII
Optimum instrumental conditions
Wavelength 357.9nm
Lamp current 10 mA
Slit width 0.010mm
Burner height ?.5:nm
Acetylene flow 3 cm • Air flow 12 cm •
conversions to actual flow rate were based mainly on maker's recommendation.
The ·effect of flame composition on the interference of iron in the determination of chromium (VI) extracted into HIBK
( 121) Blundy's method was carr,ied out by adding a thousand
fold excess of iron(III) solution (20.000/'g per ml) and the results
obtained are shown in Table XIII·and Tabl~V.
Effect of increasin concentration of iron(III) on the absorption signal of
MIBK
The effect of varying amounts of iron(III) upon Chromium(VI)
was investigated. Optimum instrument conditions were used and the
results are given in Table XIV.
75
08
0.7
01 u c 0.6 tl .0 .... 0 Cl) 0.5 .0 <(
0.4
Fig.5
Effect of observation height on absorption signal
of 20 f!9 per ml Chromium (VI I
a) aqueous solution
b) extracted into MIBI<
\ a
2 4 6 8 10 Observation height (mm}
TABLE XIII
Effect of flame composition on the interference of iron on chromium:·
Acet;2:lene flow rate Absorbance
Cr(VI) Cr(VI)plus Fe(III)
2 0.07 0.04
2.5 0.19 0.10
3 0.44 · 0.22 N
3-5 0.64 0.27 N
4 0.81 V.L. 0.28 N
4.5 . 0.8 V.L. 0-3 NoeL
5 0.75 v.L. 0.33 N ,..L
V.L.: very luminous
N noisy
TABLE XIV
Effect of increasing concentration of iron(III) on 20 t' g per ml chromium(vr)
Solution extracted into Absorbance MIBK •
Chromium(VI) 0.31 - 0.32
Chromium(VI) plus 2000~ per ml iron(III) 0.31
Chromium(VI) plus 6ooo)"g per ml iron(III) 0.31
Chromium(VI) plus 100oo)g per ml iron(III) 0.31
Chromium( VI) plus 14000fg per ml iron(III) 0.29
Chromium(VI) plus 200oofg per ml iron(III) 0.23
6. Effect of flame composition on the interference of iron in the presence of a salting-out agent in determination of
chromium(VI)
The role of salting-out agents is essentially in the binding
of water molecules, as a result of which, less free water remains
to dissolve the salt undergoing extraction. Despite this
apparently unspecific action in the extraction of inorganic substances,
the choice of salts for salting out is limited.
A salt suitable for this purpose must be very readily soluble
in water, and as little soluble as possible in the organtc solvent
used for extraction. It must also not enter into reaction either
with the salt being extracted or with any salts present that
are not being extracted, since the new compounds so formed would
behave differently towards extraction than the original ones.
A suitable salt, in addition, must not introduce difficulties
in the subsequent treatment and must be readily available in a
(122) pure state.
For a study of the effect of flame composition on the
interference of iron and the effect of salting-out agents in the
determination of chromium(VI~20rlil aliquots of 20/g per ml of
chromium(VI) were taken and ltml of 8M hydrochloric acid and 2g of
salting-out agents, and 2ml of iron(III) (20,00gfS per ml) were
added respectively. The volumes were diluted to 32ml with water
to make the solution molar with respect to hydrochloric acid.
The solution was then extracted into 20ml methylisobutylketone
previously saturated with molar hydrochloric acid. Then the
solutions were sprayed into the flame and the effect of altering
the acetylene flow was studied. The results are shown in Table XV
and Fig. VI.
77
TABLE XV
Effect of flame composition on the interference of iron in presence of salting-out agent in the determination of chromium(VI)
Jibsornances
Acetylene flow Cr(VI) Cr(VI)plus Fe(III) Cr(VI)plus Fe(III) Cr(VI)plus Fe(III) rate plus NH4Cl plus NaCl plus Na2so4
2 0.07 0.05 0.09 0.14 .
2.5 0.19 0.16 0.21 0.25 .
3 0.44 0.31 0.35 o.44
3-5 0.65 L 0.4 0.45 0.54
4 0.81 V.L. ,.......o.4 V.N. 0.42 0.52
4.5 0.8 V.L. ;_,o.4 V.N. 0.42 ,...., 0.42 L<>0 N
5 0.75 V.L. ,....-0.4 V.N. 0.39 r-' 0.38 L oaN .
L = luminous V;L: very luminous N: noisy V.N.: very noisy
Cr(VI)plus Fe(III)
0.04
0.1
0.22 V.N.
,..__. 0.27 N
,..... 0.28 N
rv0.28 N.-L
0.33 N c:.aL
g 0 .0
0.8
0.7
0.6
5 0.5 Vl
.0 <(
0.4
0.3
0.2
0.1
Compari.son of the efficicmcy ofNHCI, NaCI, Na SO as sup·p· res~ctr\tfor Fe(IJI) in the determination . 4 2 4 . .
of Cr(VI) I
I
I
I
0~
0"" a
2 3 4 5. Acetylene flow rate ( llm1n)
a)Chromium(20tg permll extracted : into MIBK
b)Chromium( 201' gper m!) plus iron(
The optimum acetylene flow rate was found to be 3 1/minute.
The most effective salting-out agent was found to be sodium sulphate
because in these conditions no depression due to presence of iron(III)
upon Cr(VI) was observed.
The effect of 8-hydroxyquinoline(oxine) reported by Ottaway
( 100) . and Pradhan was studLed here using the optimum instrumental
conditions; the results are summarised in Table XV~.
TABLE XVI
Eff t f 8 h d 1. ec 0 -1ycroxyquLno J.ne a t d.ff ~ er en t aCL L l.es on chromium(VI) in Eresence of iron(III)
Absorbances
Acidity of the solution Cr(VI) Cr(VI)plus Cr(VI)plus with respect to hydroch.: Fe(III) Fe(III) plus loric acid oxine
o.8 11 0.86 0.48 0.22
1 M 0.88 0.48 0.45
1.5 H 0.88 0.52 0.6
Apart from obtaining lower absorbances in the presence of
8-hydroxyquinoline, the effect of this reagent also seemed to be
very hydrogen ion dependent. In fact by changing the molarity of
the solution from 1.5 to 0.8 the absorbance decreased by about
79
?. Study of the effect of the addition of different acids on the extraction of Chromium(VI)from an
aqueous media into MIBK
Since many chromium analysis are performed in the presence
of acids or in some cases in the presence of bases, it was
desirable to determine the effect of acids upon the chromium
absorption. Therefore the effect of the addition of hydrochloric,
nitric and sulphuric acids upon the extraction of ,chromium(VI)
into MIBK was studied using Blundy;s(121 )method. In this method
the recommended acid concentration was molar with respect to
hydrochloric acid. The other acids were also used at molar
concentration.
In the case of nitric and sulphuric acids no absorbance due
to the extraction of chromium(VI) into MIBK was obtained, but
by the addition of hydrochloric acid into both solutions,
extractions were complete. In fact the presence of chloride was
found to be necessary for the extraction of chromium(VI).
The study of the effect of chloride ion on the extraction
of chromium(VI) into MIBK was made by Katz, McNaab and Hazel ( 123~
The effect of chloride was studied by extracting 3N sulphuric
acid solutions containing a fixed amount of chromium(VI) and
varying amounts of potassium chloride with equal volumes of MIBK
and analysing aliquots of each phase for chromium(VI). They showed
that the extraction of Cr(VI) into MIBK was dependent on the first
power of chloride ion concentration.
8. Stud of the effect of h dro en ion concentration on the extraction of chromium VI) into MIBK
To investigate the effect of varying the acidity of the
solution sprayed into the flame, to a series of aqueous solutions
(20ml) containing 2~g per ml was added varying amounts of 10
molar hydrochloric acid solution. The solutions were made up to
50ml with distilled water, and then were extracted into 20ml
of MIBK.
Using the optimum instrumental conditions, organic phases
were sprayed in order of increasing hydrogen ion concentration.
Distilled water and organic solvent were sprayed between each
solution. The results are shown in Table XVII.
TABLE XVII
The effect of acidity on the extraction of Chromium(VI) into MIBK
Acidity of aqueous Cr(VI)(20j<g per ml) solution with respect to HCl prior to
extraction
0.8 molar
1.0 molar
2.0 molar
3.0 molar
4.0 molar
Absorbance
0.88
0.9
0.9
0.86
0.86
The optimum concentration of hydrochloric acid was found to
( 123) . be 1-2 molar. Katz et al also reported that the solut~ons
more than 3M with respect to hydrochloric acid gave low results
i
due to the reduction of small amounts of chromium(VI) to chromium(III).
9. Effect of Cerium(IV)ions on the absorption of Chromium(VI)
( 121) In the method suggested by Blundy 1 the excess of Ce(IV)
ions were destroyed by dropwise addition of sodium azide. Due
to the toxicity of sodium azide, an attempt was· made to destroy
excess Ce(IV) using sodium nitrite solution. Although the results
obtained were in good agreement with standard solutions, it was
found that there was no need to destroy the excess of Ce(IV),
because no difference in absorbance of Cr(VI) due to excess of Ce(IV)
ions was obtained. The results are shown in Table XVIII.
TABLE XVIII
Effect of Cerium ion on Chromium(VI)
Solutions extracted into HIBK Absorbances
Chromium(VI) (20)ig per ml) 0.86
Chromium(VI) (20/1!; per ml) plus Cerium(IV) 0.84
Chromium(VI) (20;Ug per ml) plus 0.85 Cerium(IV)plus sodium nitrite
10. Calibration curve for chromium(VI) extracted into HIBK
Blundy's(121 )method was carried out using 2~g per ml of
chromium(VI) standard solutions. Different volumes of working
standard solution were extracted into HIBK, and from the results
obtained the absorption-concentration curve was plotted and it
was found that the curve passes through the origin and was linear
up to 1~g per ml of Cr(VI).
Table XIX and Fig. III.
Calibration results are shown in
TABLE XIX
Calibration curve for Chromium(VI) extracted into MIBK
Cr(V1)concentr~tion ;g per ml in MIBK 2 4 10 14 20
Absorbance 0.1 0.18 0.49 0.61 0.84
11. Methods of separating iron(III) from Chromium(VI)
Due to the fact that iron(III) depresses the absorbance of
chromium(VI) several methods were tried to eliminate this interference
effect. The extraction of iron(III) using isopropyl ether
(124) suggested by Kodama was applied as follows:
Aliquots(10ml) of Cr(VI) working standard solutions were transferred
into two separating funnels. To one of them was added 2ml of
iron(III) (20000(g per ml) solution. The concentration of
hydrochloric acid was brought to 8M as suggested, by adding
concentrated hydrochloric acid, followed by addition of 25ml of
isopropyl ether. The solutions were mixed thoroughly for 3 minutes,
and left for the two phases to separate •. The aqueous layer was
run off into a second separating,funnel, and the organic layer
washed again with a few drops of HCl 8M and transferred to the
same separating funnel. The same procedure \~as repeated twice
more and the aqueous solutions ~tere left to evaporate to avoid
spraying fairly concentrated hydrochloric acid solution into the
flame. The final volumes were 5-10ml, The solutions were
cooled, and transferred to 50ml volumetric flasks. The beakers
were washed thoroughly ~ti th water and the·, combined . washings
were placed in the same flask and made up to volume with \~ater.
To compare the solutions obtained from this extraction
method, to 10ml standard working solution, were added 5ml of
concentrated hydrochloric acid to obtain the same acid
concentration. The solutions were sprayed into the air-acetylene
flame using the optimum conditions recommended for the aqueous
solutions:
acetylene flow:
air flow
burner height
4e3 (;m *
3.6 c.m * 7.5 mm
and the results are given in Table XX
• conversion to actual f101; rate >~ere based mainly on maker's recommendations.
TABLE XX
(124) Separation of iron(III) from chromium(VI) by Kodama's method.
Solutions sprayed into the flame Absorbance . .
Chromium(VI) (4o~g per ml) directly sprayed 0.25 into the flame
.
Chromium(VI) (40/"g per ml) after extraction 0.25 into isopropyl ether, sprayed into the flame
Chromium(VI) (4o~g per ml) plus Fe(III) (8oofg 0.25 per ml) after extraction into isopropyl ether, sprayed into the flame.
As was shown in Table . XX very satisfactory separation
of iron(III) from chromium(VI) was achieved and so an attempt
was made to use this method of separation after oxidizing Cr(III)
to Cr(VI) by Blundy' s method, and extracting the Cr(VI) into HIBK
followed by spraying into the flame. From the results obtained,
no absorbance due to Cr(VI) was observed in the flame. This was
because of the reduction of Cerium(IV) to cerium(III) by hydrochloric
84
2Ce4+ + . ( 1)
acid(B3), and hence hydrochloric acid cannot be used in an oxidation
which necessitates boiling with excess of cerium(IV) in acid
solutions. This method seemed to be disappointing, from the
point of view of the present work.
There seemed to be two possible approaches in continuing the
present work:
i) the use of Kodama's separation method and use of
oxidants such as potassium permanganate recommended by
( 119) Feldman ; and
ii) search for another separation method.
The second choice was tried and satisfactory results were obtained
by masking iron with fluoride. Other separation methods that were
tried were not satisfactory.
The separation methods tried were:
1) Separation of iron(III) by precipitating as Fe(OH)3 <83~
Aliquots (10ml).of working standard Cr(VI) were transferred
into two beakers to one of which was also added 2ml of Fe(III)
(2000~g per ml). •
The solutions were made acid with hydrochloric
acid, and then heated to boiling, and 1:1 w/v of ammonium nitrate
solution was added dropwise until a slight excess was present.
The solutions were boiled carefully for 1 minute and the
precipitate formed was allowed to settle, and was filtered through
a Whatmann filter paper No. 541 into another beaker. The solutions
were made ~ 1M with_ respect to hydrochloric acid, and Blundy's( 121 )
method was carried out. This method was unsatisfactory.
2) An attempt was made to separate iron(III) from chromium by
the 0 (126) method recommended by Meun~er. .• In this method cup-
ferron extraction was used. Cupferron forms a chelate with
iron(III), which is soluble in organic solvent such as chloroform,
ether and ethyl acetate. Since this reagent is not very stable
to~1ards heat and undergoes degradation, all chelation reactions
using this reagent should be carried out in the cold. The
solution was acidified using 1 + 9 sulphuric acid. After
extraction of chromium(VI) plus iron(III) Blundy's(121 )method
was carried out, and the solutions were sprayed into the flame.
In this method very low results were obtained.
3) The final procedure adopted was to mask the iron(III) with
sodium fluoride, this proved to be highly effective up to at least
a thousand fold ratio of iron to chromium. Purushottam and ( 84)
eo-workers had shown that ammonium bifluoride suppresses
interference by iron when aqueous chromium samples are sprayed
into the flame.
The following procedure is recommended for the determination
of chromium in steel using sodium fluoride as masking agent. In
this method no difference in results was observed when ~he
fluoride was added before or after the oxidation step, but the
results given here were determined with fluoride added before the
oxidation step.
Analysis of steel samples
The procedure adopted was as follot~s:
A suitable amount (0.5g) of the steel sample was weighed out using
micro balance. into a 250ml conical flask covered with a watch glass.
An aliquot (50ml) of sulphuric acid solution was added to
• (127) d1ssolve the steel sample. • The flask was gently heated
until the evolution of hydrogen bubbles ceased. The flasks were
removed from the heat, allowed to cool slightly and the carbon
and carbide residues removed by dropwise addition of hydrogen
peroxide (100 volume). The solutions were boiled gently to
decompose excess hydrogen peroxide. When cool, the solutions
were diluted in 100ml graduated flasks. The size of aliquots
needed for.various steel samples were as follows:
a) for steels containing less than 0.1% of chromium,
pipette 20ml of the sample solution.
b) for steels containing 0.1-0.3 of chromium, use 10ml of
sample solution.
c) for steels _containing 0.3-0.5 of chromium, use 5ml of
sample solution.
A series of standard solutions covering the range 0-20~g per
ml of chromium(VI) were taken from standard working solutions,
extracted into MIBK, applying Blundy's method- (121 ). Then_the
solutions were sprayed in order pf increasing concentration,
distilled water and MIBK being sprayed bet11een each solution.
The zero was reset as necessary. The steel solutions were sprayed
one at a time. The standards were then sprayed again, followed
by the sample solutions. The mean of the two readings for each
standard chromium solution was used to prepare the calibration
graph. The mean absorbance of the two readings from each steel
solution used with the calibration graph, gave the·chromium
concentration for each analysed solution. The percentage chromium
in each steel sample was then calculated. The results were in
good agreement with the certificated values for the steels.
1) Determination of chromium in steels
Based on the observations above, the procedure described
in the experimental section was developed and applied to a wide
range of standard steels. The procedure used for the steel
analysis is fairly rapid and appears satisfactory as the results
show. The results are given in Table XXI, and show that the method
gives sufficiently accurate results for a range of steels.
The standard samples were stable for 3-4 hours, but the steel
samples should be sprayed into the flame within 5 minutes of
extraction. The absorbance of the steel samples began to decrease
after 5 minutes. .The reason for this decrease in absorbance is not
clear. The same standard solutions could be used for several
steel samples provided that a net~ calibration curve was plotted
in each version.
2) Final procedure
Aliquots of steel samples were dissolved in 50ml sulphuric
acid solution and carbide residues oxidized by addition of hydrogen
peroxide. Solutions were diluted to 100ml. The volumes recommended in
paragraph.3;2.5. were pipetted out, and diluted to 20ml with the
sulphuric acid solution. Sodium fluoride (4ml) was added. The
solutions were mixed and 25ml of ammoniumhexanitratocerate was
added. The flasks were immersed in boiling t~ater for 25 minutes.
They were then removed and cooled to~ 10°C. Then the solutions
were transferred to a 100ml calibrated separating funnel. The
TABLE XXI
Analysis of Steel Samples
Steel type BCS Chromium found Certificated value No, % %
Low alloy 251/1 0.51, 0.51, 0.51, 0.50, 0.53, 0.57, 0.51 0.51.
Low alloy 252/1 0.44, 0.41, 0.44, 0.42 0.43, 0.42, 0.41
Mild steel 273 0.070, 0.073, 0.07C 0.070
Mild steel 325 0.23, 0.22, 0,22 0.22
Mild steel 321 0,106, 0.11, 0.1' 0.106 0.09' 0.1.
Hild steel 322 o.o4, o.o4, 0.039, 0.039 0.03
Mild steel 324 0.07, 0.073, 0.07 0.07
flasks were washed with distilled water, then 10ml of 8M hydrochloric
acid were added.while the solutions were cooling. Solutions were
diluted up to 32ml with water, which made each solution molar with
respect to hydrochloric acid. The solutions were mixed and
exactly 20ml of MIBK (saturated with M· hydrochloric acid) were
added to each flask. After they had been shaken for 1 minute,
the layers were allowed to separate and the aqueous layers were
run off and discarded. The hexone layers were then sprayed into
the flame using optimum conditions. Distilled water.and MIBK
were sprayed between each solution.
3.2.6. Conclusion and discussion.
Chromium analysis are frequently performed by atomic
absorption spectrophotometry and a large number of elements are
known to interfere with these analysis. The principal reported
interferences are Fe, Ni, Ag, Co, Mn, Al, Nand Ti(43,45•46 •47).
The work carried out in this study has only considered the
interference of iron on chromium. This interference observed using
an air-acetylene flame as a reduction of the chromium absorption
signal, is completely eliminated with the present procedure.
As can be seen in Fig IV. addition of iron to chromium
solutions causes dep~ession of chromium absorption. The extent
of the depression depends on the flame conditions. The depressive
effect of iron is more significant in a fuel-rich flame and becomes
considerably less important in a lean air-acetylene flame. In
the. optimum instrumental conditions the degree of depression was
found to be 50'/o.
The effects of three acids were studied. In the case of nitric
and sulphuric acids no absorption due to the extraction of
chromium(VI) into MIBK was obtained, but by the addition of
hydrochloric acid into both solutions extractions were complete.
In fact this is because of the presence of chloride ion rather than
the effect of hydrochloric acid( 123)so the addition of chloride
ion was found to be essential for this extraction procedure.
The solvent extraction of chromium(VI) with MIBK is a convenient
90
and rapid method for isolating chromium from other elements ( 1 31 ~.
Introduction of this organic solvent in place of water increases
the absorbance of chromium(VI) by sixty fold as is shown in Fig. V.
Comparison of the efficiency of ammonium chloride, sodium
chloride, sodium sulphate as a salting out agent in the presence
of iron is given in Table(XV1 and Fig.(VI), As can be seen iron
interference is completely removed under the optimum, more fuel-
lean flame conditions using sodium sulphate but sodium chloride
and ammonium chloride were not very effective because the normal
chromium signal was not restored. The effect of 8-hydroxyquinoline
was also studied, and it was found that this reagent ~1as very
hydrogen ion dependent.
The effect of acidity on the extraction of chromium(VI) into
MIBK was studied and the optimum hydrochloric acid concentration
was found to be 1-2M. .· ( 123) In fact as reported by Katz and eo-workers
in solutions more than 3t-1 with respect to hydrochloric acid,
low results due to the reduction of small amounts of chromium(VI)
to chromium(III) were obtained.
Although an interference effect in the presence of 1000~g per •
ml of cerium upon chromium is reported. (91) ,in the present method
as shown in Table(XVIII)no interference was observed in the
presence of cerium concentrations of up to 3000~ per ml.
From the absorbance-concentration curve (Fig. 1), it can be
seen that the curve was linear up to 12}"g per ml of Cr(VI) and
passed through the origin.
Although the methods for separation of iron(III) from Cr(VI)
were satisfactory, they were not applicable in the present method
91
due to the oxidation step which followed.
Finally the method described gave reliable results for
the determination of chromium in a range of steels.
92
CHAPTER 4
DETERMINATION OF INORGANIC PHOSPHATE IN BIOLOGICAL SYSTEMS
~1. I~r~uclioo
A wide variety of procedures have been proposed for the
colorimetric determination of inorganic phosphate in biological
fluids. Most of the methods available are based on one of the
following:
1) Measurement of the intense and characteristic yellow colour
of molybdophosphoric acid,
2) Reduction of the molybdophosphate complex into molybdenum
blue,
3) Salt formation between molybdophosphate anions and basic
dye cations.
4.1.1.Direct measurement of inorganic phosphate
The measurement of inorganic phosphate via the yellow
molybdovanadophosphate was first mentioned by Kitson and Mellon(132)
for the determination of phosphorus in steel. Later, in 1946,
the same method was applied for the determination of phosphate in
serum samples by Simenson,Wertman, Westover and Meh1.< 133) In
this method ammonium vanadate was used with molybdate to determine
the phosphate by formation of molybdovanadophosphoric acid. In
1971 Robinson, Roughan and Wagstaff( 134)reexamined the method
after dilution and dialysis of the serum sample with 1% v/v
sulphuric acid containing 1 ml of octan-2-ol per 1. The
93
dialys~ate was added to ammonium metavanadate in nitric acid
which had been previously mixed with ammonium molybdate solution.
The absorbance of the yellow complex formed was measured at
403nm against a reagent blank. They reported that the results
obtained were in good agreement with the established molybdophosphate
methods.
In the case of measuring unreduced molybdophosphate
methods developed for determining inorganic phosphate, polyoxy
ethylene sorbitan monooleate has been used by Ferlan( 135)to
keep the serum protein. In this method the intensely yellow
colour due to molybdophosphoric acid and the monooleate was
measured and no preliminary reduction step was necessary. The
wavelength suggested was 405 or 370nm. Interferences observed
were due to arsenic and by large amounts of silicon.' and iron(III).
Daly and Ertingshausen( 136)applied a similar procedure for
determining inorganic phosphate in human blood serum. They found
that the unreduced molybdophosphate complex absorbs ultraviolet
light. By using an acidified ammonium molybdate "Tween 80 11 reagent
and a centrifugal analyser they developed a direct method. The·
wavelength suggested was 340nm. These authors also have checked
many organic polymers as to their suitability for keeping serum ·
protein in solution. "T~1een 80" polyoxyethylene sorbitan monooleate
was reported to be the most effective in preventing protein
precipitation and turbidity. A simple serum phosphorus analysis by
continuous flow·ultraviolet spectrophotometry without reduction of
the phosphomolybdate complex is described by Amador and Urban( 13?).
In this method the dilute sample is dialyzed into dilute (1jb v/v)
sulphuric acid, and is then mixed with an ammonium molybdate-sulphuric
acid - "Tween 8011 solution. The absorbance is measured at 340nm
94
with a miniaturized Auto-Analyzer manifold and a linear absorbance
spectrophotometer. Even this method outlined above quantitate
the unreduced phosphomolybdate heteropolyacid 1 but have not yet
been accepted as routine procedures.
4.1.2. Reduction of molybdophospnate complex.
Most of the commonly used methods for the determination of
inorganic phosphate are based on Fiske and Subbarbw (FOUS) 1s(138)
method published over 50 years ago. The principle and operation
o£ the method is simple. The filtrate obtained after precipitating
proteins by means of trichloroacetic acid, is treated with an
acid molybdate reagent which reacts with inorganic phosphate to
form phosphomolybdic acid. This is followed by reduction of
hexavalent molybdenum by means of 1 12 1 4 aminonaphtholsulphonic
acid to give a blue compound which was esti~ated colorimetrically.
The modifications to the method use either a different reducing agent
or perform the reduction under different conditions. Gras and
Kolck(139)have studied the factors affecting the molybdenum blue
calorimetric method for phosphate determination and a set of optimum
parameters suggested. The parameters studied were the concentration •
of the reagents (sulphuric acid and molybdate), the amount of the
reducing agent (tin(II) chloride) added, the order in which the
reagents were added, the temperature at which the solution was
kept, and the time interval elapsed between the time the last
reagent was added and the time the photometric measurement was
taken.
Inorganic phosphate in biological samples is conventionally
determined by use of these molybdenum blue reactions. These methods
could be classified as follows depending on the treatment of the
95
biochemical system:
a) Automated methods using an AutoAnalyz,er.
b) Deproteinization by using either trichloracetic acid
or perchloric acid.
c) Direct methods, (i.e. deproteinization was not carried
out).
Automated methods
A simple and sensitive automated method for the determination
of inorganic phosphate in serum, urine and other biological fluids
is given by Hoppe-Seyler, and Gundlach( 140>. In this method
the materials acidified with hydrochloric acid are dialyzed
against hydrazine sulpl'\ate, and ammonium molybdate was mixed
into the dialyzate; The reduction of t'he molybdic acid occurs
when the sample solution was heated at 95°C for 7 minutes. The
absorbance of t he coloured solution was read at 660nm. Amic,
Lairon and Hauton(141
)also dialyzed biological fluids into 0.5M
sulphuric acid, follo>ling the addition of molybdate reagent and
ascorbic acid solution. 0 The solutions were kept at 95 G for
1 minute, and the absorbance was measured at 660nm.
A simple; sensitive and reliable method is presented( 142)
for the direct photometric measurement of inorganic phosphate in
serum and urine. The reagents required are reported to be stable,
inexpensive and minimal in number. The molybdophosphate complex
formed in an acid medium is reduced by p-methylaminophenol sulphate,
subsequent alkalinization with ethanolamine produces a clear blue
colour, which is measured at 660nm, against a reagent blank. In
this method bilirubin interfered slightly. Other reducing agents
96
which have been examined for automated analysis include tin(II)
chloride(143)and iron(II) ammonium sulphate-t'hiourea, (144>.
Methods involving deproteinization
Reducing agents proposed by several authors for determination
of phosphate via molybdophosphate reduced to molybdenum blue after
deproteinization of blood, urine and serum samples using trichloro
acetic acid are as follows. Negrin( 145)reported that when
hydroiodic acid (HI) is used as a reducing agent in place of
4-amino-3 hydroxynaphthalene-1-sulphonic acid(138), a '2.5 fold
increase in sensitivity was observed and production of stable
colour was obtained. Guirgis and Habib( 146)used a new reducing
agent, metamizol (sodium 1-phenyl-2,3 dimethyl-5-pyroazolone-4-
methyl-amino methane sulphonate). This reagent was employed for
the assay in a protein free filtrate of serum or plasma or
(147) directly on a diluted urine specimen. Parekh and Jung ~
combined the molyb· ,'lie acid solution with the trichloroacetic acid
(TCAA) as one reagent in order to save reagent preparation time
and avoid one reagent addition step in the analysis. A new
reagent, P-phenylenediamine-hydrochloric acid was used for colour
development, which resulted in a ~ery stable molybdenum blue complex
obeying Beer's law. Ascorbic acid was used (148-149,152)for reducing
molybdophosphate followed by stabilization'of the coloured product
with citrate and arsenite. Another variant of Fiske and SubbaRow's
method was studied for estimation of inorganic phosphate in serum
by Nath and Debnath( 151). They used freshly prepared 4-amino-3-
hydroxynaphthalene-1-sulphonic acid in 5% w/v sodium sulphite as
reductant. They suggested that sodium sulphite was preferred to
the bisulphite solution used by Fiske and SubbaRow because of the
more stable colour that has been obtained in this method.
97
------------------------------------------------------------------------------- - ~
A rapid micro method for determination of phosphate in biological
t 0 1 ° 0 b J 0 k ( 150) I tho th d th 1 ma er~a s ~s g~ven y aen~c e. • n ~s me o e samp es
were digested with boiling perchloric acid, followed by addition
of ~/o w/v ammonium molybdate and 1% w/v of p-methylaminophenol
sulphate in 3% ~v sodium bisulphite solution. The absorbance
was measured at 578nm after 20 minutes delay at room temperature
for stable colour development to take place.
Direct methods
Determination of phosphate in biological fluids without
preliminary treatment for deproteinization has been the subject
of study by a few workers. A new calorimetric procedure is that
proposed by Morin and Prox(153)for the determination of serum
inorganic phosphate with a-phenylenediamine dihydrochloride
as a reducing agent and dimethylformamide and poly(vinylpyrrolidine)
as a catalyst with relatively low acidity. The procedure requires
neither elevated temperature nor deproteinization, and yields a
stable molybdenum blue in 5 minutes. The recovery was reported
to be 9o//o and the relative standard deviation, 0.6%. This
method is the most sensitive phosphate procedure reported to
date. Another direct method was'devised by Goodwin(154). In this
method inorganic phosphorus formed a molybdophosphate complex in
the presence of borate, and the complex was reduced with ascorbic
acid. The resulting suspension was dissolved in sodium carbonate
solution and the absorbance was measured at 720nm. Recently a
direct method was given for determination of inorganic phosphorus
in serum and urine( 155)based on the reaction of phosphorus with
acid molybdate at 37°C and reduction of the resulting molybdophosphate
with ascorbic acid. In this method addition of arsenite and citrate
containing a non-ionic surfactant (Polysorbate 20, 60 or 80)
prevented further reaction of molybdate and avoids the need for
deproteinization of the sample. The method was reported to be
simple and rapid.
1f.1.3. Ion-association complex formation between molybdophosphate and basic dyes
Several workers have studied the fact that molybdophosphate
complexes react with basic dyes, which are well known to have high
molar absorption. The most sensitive method thus far reported
for the determination of inorganic phosphate using the solvent
extraction method seems to be the procedure based on the formation
of an insoluble Methylene Violet molybdophosphate(156>. The
precipitated molybdophosphate complex, after being washed several
times, dissolved in acetone before being determined colorimetrically.
Though this method using Methylene Violet is highly sensitive, a
careful and troublesome manipulation is required in order to obtain
a reproducible amount of dye-molybdophosphate complex by the complete
washing away of the contaminating dye.
An extraction method has been described for the determination
of traces of phosphate in the form of molybdophosphate, in which
Safranine is used as the ion-association complexing agent, which
can be extracted by aromatic ketones, particularly acetephonone.
The phosphorus content of the extract is determined by a differential
(157) spectrophotometric method • In this method acetophenone-o-
dichlorobenzene 3:1 v/v was used. Babko, Shkaravskii and Kulik( 158)
studied the reaction between molybdophosphate complex ions and
basic dyes, and the possibility of using basic dyes for the
extraction-photometric determination of phosphorus. They have
99
examined crystal Violet, Methyl Violet, Malachite Green, Auramine,
Iodine Green, Rhodamine 6I, Neutral Red, Safranine, Toluidine Blue
and Basic Bright-Green for this purpose. In order to select
the best solvent for extraction of dye-molybdophosphate complex
and the most sensitive dye for determining phosphorus, they pave
plotted the absorption spectra of molybdophosphate complex-basic
dye compounds extracted into the various solvents and calculated
the apparent molar absorptivity. The compound which has the
highest molar absorptivity was the molybdophosphate complex with
Crystal Violet extracted into 1:1 v/v n-butanol and cyclohexanol.
In this extraction-photometric method, the free molybdate also
reacts with basic dyes to form a precipitate which is extracted
in the same way as the molybdophosphate-basic dye compound.
Consequently, excess molybdate ion must be removed before adding
the dye. The removal of this excess molybdate is possible by
shaking the organic phase with dilute nitric acid. On shaking
with a.hydrochloric acid solution of potassium permanganate, any
free dye is completely decolorized, while the phosphomolybdate-
basic dye compound is not affected. The maximum sensitivity
obtained by this method, using Crystal Violet, was 0.01;4g phosphorus
(159) . . per ml. Trautner determ1ne~phosphorus traces in copper
anodes as the molybdophosphate-Crystal Violet complex in butyl
acetate-acetone mixture. The sample ~1as dissolved in 50',6
v/v nitric acid, oxidized with hydrogen peroxide, and treated 1~ith
ammonium molybdate. Molybdophosphoric acid was extracted into
butyl acetate, treated with Crystal Violet and mixed with acetone.
The detection limit was found to be ~.4ppm phosphorus. The same
( 160) . author determ1ned phosphorus in tin-lead solder py extraction
from a 0.8-1.3 molar nitric acid solutions as molybdophosphoric
100
acid into butylacetate. The molybdophosphoric acid was precipitated
with acidic Crystal Violet, the precipitate was dissolved in
butylacetate-acetone and the absorbance was measured at 59Dnm.
The determination of inorganic phosphorus in biological systems as
a dye-phosphomolybdate complex was studied as far back as 1947.
Soyenkoff( 1G1)has been · d th "b"l"t of th 1" t" exam1ne e poss1 1 1 y e app 1ca 1on
of dye salt of phosphomolybdate with Quinaldine Red for the
quantitative determination of small amounts of phosphate in a serum
filtrate over the concentration range of 0.02 to 0.2mg of phosphorus
per 1. Itaya and Ui(162) examined a series of basic dyes as to
their analytical use. They suggested that Malachite Green was
most useful for determination of phosphate in blood serum. In
this method the deproteinization 1ms applied using perchloric acid.
They also have shown that the sensitivity of the method was far ·
greater than other procedures such as those based on the formation
of molybdenum blue, being 30 times more sensitive than the Fiske
SubbaRow ( 138) method and 12 times than that of the Nartin-Doty• s
method~ 163) The effect of detergents such as sodium lauryl
sulphate or polysorbitan to prevent the coloured complex from
sedimentation was studied also. Addition of Tween 20 after the
colour development kept the colour stable for 48 hours. The
addition of detergent before the addition of the colour developing
reagent, caused retardation in the complex formation. Van Belle(164)
made comparative studies of the method based on the formation of
the molybdophosphate-Methyl Green 00 complex with Itaya and Ui's(162)
Malachite Green method. As a result of this comparison he found
that Methyl Green 00 was to be preferred to Malachite Green for
several reasons, namely:
101
I
I
I
I
a) Methyl Green 00 was more soluble in water and it
did not precipitate on contact with the acid molybdate
reagent,
/
b) lower concentrations of Methyl Green 00 could be used>
c) Methyl Green 00 did not stain the plastic tubes of the
apparatus,
d) a comparatively mild acid concentration could be used
(1.25 molar hydrochloric acidh
e) a relatively short reaction time (25 minutes) was required.
This method was reported to be a sensitive reaction for automated
determination of inorganic phosphate in serum samples. The
coefficient of variation \<as found to be 0.91%. 0 ( 165) Bast~aanse
also described a modification of Itaya and Ui' s method using a 10\•er
concentration of Malachite Green (0.04~/o compared with 0.2%)
and a different stabilizing agent, Sterox resulting in an analytically
more suitable determination. The advantage of this_ stabilizing agent
compared to Tween 20, used by Itaya and Ui,<162) -is that Sterox
can be added to the reagent consists of ammonium molybdate and
Malachite Green while Tween 20 must be added after the colour has
developed. The author reported that this method could be applied
to serum and urine samples, and gave precise and highly repr{Jducible
results. Traces of inorganic phosphorus in biological material,
particularly serum and urine, ~<ere determined satisfactorily by
the application of the method of Itaya and Ui. In this method( 1GG)
dextran was used in place of Tween 20. This modification was
reported to be more sensitive and the reagents more stable than
methods involving the reduction of molybdophosphate into molybdenum
blue, Hohenwallner and Wimmer( 1G7)proposed a calorimetric
102
micromethod for the determination of inorganic phosphate avoiding
deporteinization. In this method they used Malachite Green and
Sterox SE as the stabilizing agent. This method gave absorbance
values lower than those obtained by methods involving depr,9teinization
and it was suggested that organic phosphates may be hydrolyzed
during deproteinization using strong acids such as trichloroacetic
acid or perchloric acid. In order to prove their ideas about this
hydrolysis of organic phosphate during dep~<oteinization, they
tried to obtain as mild a deproteinization as possible, by desalting
by addition of cold saturated ammonium sulphate solution. After
this desalting they obtained nearly the same values as with the
Malachite Green method without deproteinization, and so they believe
that phosphate groups were hydrolyzed from organic compounds during
deproteinization with strong acid, but not during desalting with
cold saturated ammonium sulphate solution.
Malachite Green has also been used by Altmann, FUrstenau,
Gielewski and Scholz( 1GS)for the determination of phosphate in
natural water. They stabilized the suspension of the !1alachite
Green-12-molybdophosphate ion-association complex by the addition
of polyvinylalcohol. \1hen the excess of the Malachite Green has
• reacted in acid solution to form colourless products, the remaining
colour is a measure of the phosphate present. Stevens and Yeomans(1G9)
tried to adapt the method supplied by
phosphate analysis kit which is based
Roche Products Ltd in their
. (162) on Itaya and U1's method,
directly for use with the Vickers H300 Analyzer using 11alachite
Green and measuring the absorbance of molybdophosphate~alachite
Green ion-association complex at 610-660nm. They found that
the method described in the Roche kit hand book was unsatisfactory
for their purpose. The method that they have suggested involves
•
103
the initial dilution of plasma with deionized water and the
addition of reagents containing Tween 20 without protein precipitation.
Because phosphorus is present in the blood as both organic and
.inorganic phosphorus, and the phosphorus present in the
trichloroacetic acid filtrate of blood consists of the inorganic
phosphate as well as many of the organic phosphates, higher results
could be obtained by methods involving trichloroacetic acid for
deproteinization. In this method using Vickers Analyzer, lower
results compared to the results obtained by a Technicon Auto
Analyzer (using the same standards for both systems) explained by
the high specificity of the method for inorganic phosphate. A
highly sensitive method using the same basic dye for the determination
(170) . . of phosphate in serum and urine is given by Kallner , us~ng
polyvinylalcohol as a protective colloid in order to prevent
precipitation of the dye salt formed. In this method no
deproteinization was applied and therefore the method was suitable
for a one step procedure that allowed the analysis of. several
hundred samples per hour. Malachite Green has also been used
. (1?1) . by Anner and Moosmayer who descr~bed the application of
Altmann et al 1s(168
)method to biological systems, reducing the
reaction mixture from 38ml intb 2ml. They determined the inorganic
phosphorus content of desheathed rabbit vagus nerve applying
trichloroacetic acid and triethanolamine for extraction of phosphate
from the tissue. They have reported that phosphate could be
measured in the presence of phJphorylated compounds. The coloured
complex was stable for several days.
The review of the literature showed that Crystal Violet has
not been recommended by many workers as a reagent for determination
of phosphorus based on the direct measurement of molybdophosphate
104
complex with Crystal Violet. Franzke et al( 173)proposed a method
for determination of small amounts of phosphate in biological
material. In this method phosphatide containing mixture was
separated by TLC on Kieselgel G, with chloroform-methanol-H2o
(65:25:4) v:v:v as solvent. The isolated compounds were mineralized
by treatment with perchloric acid followed by spectrophotometric
determination of phosphorus as the molybdophosphate-Crystal Violet
complex.
4.2. Experimental
Although the basic dyes generalJ.y give very sensitive analytical
methods, their selectivity is not always satisfactory. Some degree
of selectivity can be achi"eved by the choice of reaction medium
in addition to the choice of dye. The present work has been concerned
with the suitability of various dyes for the determination of
. . ( 162) phosphate based on the Itaya and u~•s method.· They have shown
that it was possible to measure the inorganic fraction of plasma
phosphate by makinguse of the change in absorption spectra obtained
when it is allowed to react with acidified solutions of basic dyes
attached to molybdate. The dye was freed from the complex as the
phosphate preferentially combines with molybdate.
4.2.1. Reagents
1) Potassiumdihydrogen phosphate:
a) Stock solution: dissolve 1.4341 g of dried potassium
dihydrogen phosphate in 1 1 distilled water made up to
1 1 in a volumetric flask. This solution contains
b) Working solution: dilute 1 ml of stock solution into
105
1 1 with distilled water. This solution contains
,r 3-vg per ml. Po4 ,
2) Ammonium molybdate: 4.2% w/v (NH4)6Moo24 .4H2o was made
by dissolving 4.2g of ammonium molybdate in 100ml of
4.5 - 5.0N hydrochloric acid.
3) Tween 20 1.~/o w/v aqueous solution.
4) Malachite Green 0.2% w/v aqueous solution.
5) The colour-developing reagent: One volume of 4.2%
ammonium molybdate in 4.5-5.0 N hydrochloric acid is
mixed with three volumes of o.~/o Malachite Green. After
20-30 minutes, the mixture is filtered and stored at
room temperature. This reagent solution is stable for
at least 3 weeks.
4.2.2. Procedure
Into 1 ml of standard phosphate solution was added 5ml of the
colour-developing reagent, 0.2ml of 1.~/o Tween 20 and the colour
developed was measured at 660nm against reagent blank.
4.2.3. An experimental assessment of the determination of phosphat?( 162) with Malachite Green according to the method of Itaya and u~.
This method was published in 1966 and it was claimed that
the method was a very sensitive technique for determination of
phosphate on the basis of the principle that Malachite Green at
lower pH forms a complex with phosphomolybdate with a marked shift
of the absorption maximum from that of the original dye. Although
the same method using Malachite Green has been tried as recommended,
no significant shift due to the formation of the phospho~olybdate-
106
Malachite Green was observed. The results are given in Table
XXII, and shown graphically in Fig. VII.
TABLE XXII
Assessment of Itaya and Ui's method . .
vlavelenr,th 610 620 630 640 650 660 670 680 690 700
Absorbance .
Sample • 630 .682 .660 -572 .4?io .360 .21!7 .220 .1?0 .1!12 against H2o
Blank .420 .465 .430 -330 .215 .122 .075 .040 .025 .015 against H2o
Sample .224 .232 .242 .255 .255 .245 .225 .195 .145 .100 against blank
.
4.2.4. Application of Itaya and Ui's method to different dyes.
A similar method was applied using the same reagents
except that various dyes were used and were added individually. The
dye solutions were all 0.~/o w/v. For selection of a suitable dye,
the colour before and after the addition of 1fg per ml of inorganic
phosphate into the acidified solution of basic dyes containing
ammonium molybdate was considered. These are listed in Table XXIII.
Among the Sevron basic dyes studied by Burgess(58), the
most promising ones were found to be Sevron Red L and Sevron Red
... GL. As the structure of Sevron Red GL was given by the manufacturers
(E.I. Du Pent de Nenours & Co) the latter was purified by him and
studied as a prospective reagent for anions. The anions studied
were SbCl6-, T1Cl
4-, GaC1
4-, InBr
4-, Reo
4-, Auc1
4-. Of all those
107
0.7
Gl u0.6 c Cl
..0 .... 0
]0.5 <(
0.4
0.3
0.2
0.1
Fig. 7
AssQsmQnt of Itaya and Ui's mQthod
0 Sample? against H 0 2
s etank against Hp x Sample? against blank
__ .,.-_ _,.,__...;~:--- )(
X--)(-
10 20 30 40 so 60 70 80 90 100 Wav~-!C?ngth
TABLE XXIII
Colour of the basic dyes in an acid solution with and without addition of molybdophosphate.
Dye Colour of the
without phosphate
Safranine 0 purple
Brilliant Green yello~rish brown
Acid Fuchsine red
Malachite Green brown-green
Victoria Blue brmm-yellovr
Crystal Violet orange-yellov1
Sevron Red L red
Sevron Red GL red
solution
with phosphate
reddish purple
green
red
brown-green
blue
green
red
red precipitate
anions tested tetrachloroaurate(III) was found to be the most
suitable. In his method for defining gold, tetrachloroaurate-
Sevron Red GL ~ras extracted into a-dichlorobenzene.
The absorption spect~a of the system indicated in
Table XXIII were recorded using a Unicam Sp 8000 spectrophotometer
against their own blanks using 1cm silica cells. From the
results obtained in the present study, the method did not seem
to be satisfactory. For this reason attempts \<ere made to adapt
a solvent extraction finish to this procedure.
108
.
l
4.2.4. AppUcation of solvent extraction to Itaya and Ui' s method using different dyes
Itaya and Ui's method was carried out using the same reagents
as before and the dye-molybdophosphate complexes formed were
extracted by 5ml aliquots of organic solvents using separating
funnels. The solutions were shaken for 1 minute and after the
layers had separated completely the organic layers were run off
and the absorbance characteristics were investigated and subsequent
quantitative measurements were made at the absorbance maximum for
each dye. Reagents blanks were run in parallel; the absorbances
obtained by measuring the extracted phosphomolybdate complex
being measured against appropriate blanks. If the absorbances were
too high, the organic phases were diluted with the same solvent
so that the final absorbances were always less than 1.2. Reagent
blanks were treated the same way and their values were subtracted
from the total absorbance to give the net absorbance. The dye-
molybdophosphate complexes behaved quite differently with the
various solvents, some did not extract the complex at all, and
in case of chlorinated solvents, the blank absorbances were very
high and even in some cases higher than the absorbances due to
the dye-molybdophosphate complex. This is because certain of
the dyes themselves are relatively soluble in organic solvents.
Solvent mixtures were also examined because mixing a solvent
into which the coloured dye-phosphomolybdate complex reacts readily
with one which is a poor extractant may have made adjustment of
the sensitivity and range possible. Initially the observations
were of a qualitative nature, as this was sufficient to determine
whether any significant amount of dye was extracting into the
109
organic phase. When the qualitative observations seemed to
be satisfactory, then they 11ere carried out quantitatively as
indicated below. The results obtained are shown in Table ~XIV).
As a result, an attempt to produce an extractive photometric
method for the determination of phosphate based on extracting the
dye-molybdophosphate complex Has unsuccessful. The reason for
this was either because of the very high blahk absorbance due to
chlorinated solvents, or to the non extractability of the dye-
mo1ybdophosphate complex.
4.2.5.' Investigation of the determination of phosphate ;,ith Crystal Violet
Altmann et al( 168)developed a method for the determination
of inorganic phosphate in water based on the measurement of
complex formation between Malachite Green and molybdophosphate.
In the present study the same method was applied using Crystal
Violet, Sevron Red GL, Sevron Red 1 and Brilliant Green but from
the results obtained, Table (XXV) no improvement was achieved.
But in the case of using Crysta~ Violet and using nitric acid in
place of sulphuric acid higher molar absorptivity was observed, by
lowering the concentration of sulphuric acid (from 3ml to 2ml) higher
absorbance compared to Altmann's method was achieved, but when
hydrochloric acid was used in both cases precipitates were obtained.
By using Crystal Violet it was found that above a 20 per cent
increase in, sensitivity was obtained, and by heating the mixed
solutions the development time was reduced to twenty minutes.
110
TABLE XXIV
Extraction tests of various dye-molybdophosphate complex into various solvents.
The dyes used are Sevron Red GL(SRGL), Sevron Red L(SRL), Crystal Violet(CV), Victoria Blue(VB),
Brilliant Green(BB) and Safranine O(SO).
SRGL Organic solvent used qual.-quant.
Methylisobutyl ketone GE-NA
a-dichlorobenzene GE-NA
Benzene NE . Chloroform NE
a-dichlorobenzene NE· nitrobenzene
Ghlorobenzene-nitro- GE-NA benzene 1/1 v/v
Toluene NE
Ether NE
1,2-Dichloroethane GE-NA
Key:- CE NE
complete extraction - no extraction
' phm - phase miscible - no absorbance
SRL qual,-quant.
GE-NA
GE-NA
GE-NA
CE-NJ,
GE-NA .
GE-NA
GE-NA
GE-NA· _c
GE-NA
NA IC = intense colour due to chlorinated
solvent
Dye CV VB BG
qua.L,-quant. qual.-quant. qual.-quant.
GE-NA phm. GE-NA
CE-OC GE-NA GE-NA
GE-NA NE NE
CE-IC CE-IC GE-NA
CE-IC CE-IC GE-NA
CE-IC CE-IC phm.
NE GE-NA NE
NE GE-NA NE
GE-IC GE-IC GE-NA
qual. qualitative observations.
quant. quantitative observations.
~u qua.L. -quant.
phm.
GE-NA
NE
GE-NA
phm.
phm.
GE-NA
NE
phm.
4.2.5.1. Reagents.
1) Standard phosphate solution
a) Stock solution: Dissolve 0.7529g of Analar disodium
hydrogen phosphate.12H2o in distilled water, and make
the solution up to 500ml in a graduated flask. This
solution contains o.4mg phosphate per ml.
b) Working solution: Dilu~e 5ml of stock solution to 1 1
. with water. This solution contains ~g Po43- per ml,
and should be freshly prepared daily.
2) Nitric acid solution 24% v/v
3) Polyvinyl alcohol solution 1% w/v
Dilute 10g of polyvinyl alcohol (Mol.Wt. 14000, BDH)
in water by gentle heating. Dilute the resulting solution
to 1 1 with distilled water. Mix and filter it through
a Whatman No. 54 filter paper.
4) Crystal Violet solution:
Dissolve 0.051 g of Crystal Violet (B.P. grade) in water,
add 125ml of 10 g per 1 polyvinylalcohol solution, and
dilute the resulting solution to 250 ml with water.
The concentration of the dye in this solution is 5 x 10-411.
5) Sodium molybdate solution, 0.1M
Dissolve 6.049 g of Na2Hoo4 .2H2o in 250ml of distilled
water and mix the solution thoroughly.
112
4.2.5.2.Basic Procedure
Aliquots of the working standard phosphate solution were
pipetted into 100ml conical flasks. The volumes were made up
to 10ml with distilled water. Polyvinyl alcohol solution (15ml)
nitric acid solution (3ml), Crystal Violet solution (5ml) and
sodium molybdate solution (5ml) were added in turn with mixing to
each flask. The solutions 1<ere read after 60 minutes
spectrophotometrically against the blank at 560nm in a 1cm silica
cells.
TABLE XXV
Application of Altmann et al's method using different dyes
Altmann et al's method was carried out as recommended and
the spectrum were recorded using a Unicam SP 8000 spectrophotometer
against appropriate blanks. The results are given below.
4.2.5-3-
Dye Absorbance
Malachite Green 0.576
Crystal Violet 0.628
Brilliant Green 0.49
Sevron Red GL 0.032
Sevron Red L 0.038
Study:th~ effect of temperature on the formation of molybdophosphate-Crystal Violet complex.
In Altmann's method the effect of temperature is reported to
be negligible between 20°-40°C, but in the present method when
0 solutions were kept in a water bath for 20 minutes at 35 c, the
113
development time was reduced to twenty minutes. The results
are shown in Table (XXVI).
TABLE XXVI
Effect of heating time on the absorption spectrum of molybdophosphate - Crystal.
Violet
Time that solutions left Absorbance in water bath (minutes)
25°C 35°C 45°C
5 0.785 0.888 0.790
10 0-779 0.817 0.782
20 0-771 0.744 0.778
30 0.722 0.743 0.766
4o 0-712 0.743 0.758
4.2.5.4. Standard Curve and absorption spectrum
55°C
The dye salt
precipitated
The method carried out as g~ven in 4.2.5.2. The solutions
after being mixed, were left for 20 minutes at 35°C, and the
solutions were read against·a blank. The absorption maximum was
found to be at 555-565nm and 560nm was selected for quantitative
measurements. The standard curve is shown in Fig.VIII ; a
straight line was obtained for absorbance against the concentration
of inorganic phosphate over the range from 0.05 to 0.5)'g per ml.
Stability of colour of the molybdophosphate-Crystal Violet
at 35°C in different time intervals were determined and the
114
(\! 0.4 u a
. .Ll L.
~ ~ 0.3
0.2
0.1
Fig.S
Calibration curve for determination of inorganic
phosphate in biological systems using Crystal
Violet
1 2 3 4 Vol of P~=l2,...g/mllml
results obtained are given in Table(XXVIIl.
TABLE XXVII
Colour stability of molybdophosphate-Crystal Violet complex with time
Time(min) 5 10 15 20 25 30 absorbances of:
blank against 0.202 0.107 0.087 0.086 0.084 0.083 water
Sample against 1.09 0.924 0.860 0.830 0.824 0.826 water
Sample against 0.88 0.817 0.773 0.744 0.740 0.743 blank -
The colour ~1as found to be stable after 20 minutes being left at
40
0.083
0.824
0.742
4.2.5.5. Comparison of the present method with Altmann et al's method
Altmann et al's method was carried out as recommended as well
as the ,Present method, and fr9m the results obtained, above 20 per
cent increase in sensitivity was" achieved by using Crystal Violet.
Altmann's method Present method ..
Number of readings 10 15 Average absorbance value 0.576 0.739
IR.2 /.Standard deviation 3.49 5.2 .
99't6 confidence limit on average 3.6 3-99 value .
Apparent_~ole1 absorptivity (1 mol cm ) 1.04 X 105 1.34 X 105
115
4.2.6. Possible application of the present method on determination of phosphate in biological systems.
In a study of the possible applications of this procedure
to serum samples, possible interferences were investigated. The
2+( ) 2+( effect of Ca 10mg per 100ml , Mg 2mg per 100ml) and bovine
albumine (7g per 100m1\172\as investigated. The phosphate
concentration is reported to be 3.5mg per 100ml in serum. The
results are given in Table XXVll/ •
t . . t• . 1 d (138) • The depro e~n~za ~on ~n serum samp es was ma e us~ng
trichloroacetic acid. To 1ml of the serum sample was added 9 ml
of trichloroacetic acid (TCAA), the solution was mixed well, left
for a few minutes, centrifuged for 5 minutes, and filtered if
necessary. In fact the same results were obtained without
centrifugation, and filtration was found to be sufficient. In
the present method the filtration method was applied. The sample
used was "Seronorm" analytical values Batch No. 113(BDH). Results
are given below:
Number of readings 10
Average value 0.353
Standard deviation 2.63
9~~ confidence limit on average value 2.70
116
r------------------------------------------------------------------------------
TABLE XXVIII
Study of the effect of Interfering ions
The basic procedure given in 4.2.5.2. was carried out
by addition of ions that might interfere in this method. The
results obtained are:
Interfering ions ~dded 20f1g per 38ml Po4 -
Mg++(2mg per 100ml)
Ca++(10mg per 100ml)
into
Bovine albumine (7g per 100ml).
Absorbances
0.735, 0.740, 0.735, 0.732
0.738, 0.734, 0.731; 0.737
0.733, 0.733, 0.741, 0.730
0.731, 0.735, 0.739, 0.74o
lf.2. 7. Checking the method using an unknown sample
Wellcomtrol Quality Control Sera (Assayed Code numbers
BC01-02-03) samples were used as the unknown samples. The ,.....
cert,.ificated ranges were given in the table attached. The
Wellcomtrol sera samples were diluted with exactly 10ml of distilled •
water by means of a bulb pipette, the bottle was swirled to aid
· dissolution of the freeze-dried plug and the solution was left
for 20 minutes with intermittent swirling to ensure that no serum
adhered to the side of the bottle. After that, the bottle was
inverted and the solution was mixed thoroughly to make sure that
all traces of dry material had dissolved before any of the serum
was withdrawn.
Aliquots (0.5ml in case of sample No: 1 and 3, 1ml in case of
sample No: 2, using 0.5ml and 1ml bulb pipettes) were pi petted into a c ·'
117
I I
I
10ml volumetric flask containing 7-8ml of 10% v/v trichloracetic
acid. The flasks were mixed after each addition, and were brought
up to volume with 10% trichloracetic acid, mixed well, left for
a few minutes, then filtered through a filter paper into a flask.
An aliquot (5ml) of filtrate was pipetted into a 25ml volumetric
flask and made up to volume with distilled water, 5ml aliquots
were tried for each experiment as described in 4 • .2. 5. :2 • and
the results are given as follows:
a) The determinations made immediately after dissolution of the Wellcomtrol sample:
Batch No. of Average value Standard deviation 99";6confidence No readings mg per 100ml limit
1 11 4.88 0.17 0.16
2 8 2.75 0.045 0.056
3 11 5.37 0.12 0.12
b) The determinations have also made 24hours or more after dissolution of the \ifellcomtrol sample:
Batch No. of Av·erage value Standard deviation 9CJJ!, confidence No. readings mg per 100ml limit
1 29 4.81 • 0.15 0.079
2 21 1.65 0.11 0.666
3 27 . 5.39 0.15 0.080
~
The cert~1ificated values given for these batches are: 5.0, 2.8, 5.5mg
per 100ml respectively.
113
4.3. Conclusion and discussion.
Numerous procedures have been proposed for the calorimetric
determination of inorganic phosphate in biological fluids. Most
of these procedures are based on the measurement of the intense
and ch~racteristic yellow colour of 12-molybdophosphoric acid,
or on reduction of the 12-molybdophosphate complex to molybdenum
blue, which is measured colorimetrically. A few workers have proposed
the determination of phosphorus on the basis of salt formation of
12-molybdophosphate with Crystal Violet( 159,160-173l. Crystal
Violet, which has a violet colour in neutral aqueous solution,
forms an insoluble blue-violet dye salt with 12-molybdophosphate,
and this may be kept from precipitating by addition of a stabilizing
surfactant such as Tween 20( 162). B th t· f ·a th y e ac ~on o ac~ , e
Crystal Violet in true solution is further protanated to a green
divalent form, whereas the Crystal Violet in colloidal solution
remains unreacted and can be measured colorimetrically. Traces
of inorganic phosphate in biological material, particularly sera,
have been estimated, apparently satisfactorily, by the application
of these procedures.
. (168) In the present study a method by Altmann et al , using
Crystal Violet in place of Malachite Green, and stabilising the
colloid with polyvinyl alcohol, was tried and was found to
give good results. Altmann's method has the disadvantage of a
1-hour colour development period which could detract from its use
in manual as well as in automated analysis. In the present study
other dyes and solution conditions have been investigated in an
attempt to reduce this development time. By using Crystal Violet
it was found that above a 20 per cent increase in sensitivity was
119
obtained, and by heating the mixed solutions the development
time was reduced to twenty minutes.
A study of the possible applications of this procedure to
serum samples, possible interferences were investigated. The
effect of Ca2+, Mg2'~'' and bovine albumine was examined and no
interference effect was observed in the determination of
phosphate using Crystal Violet.
This method has given good results and may be considered
to be an alternative method to that in which molybdenum blue is
measured.
120
CHAPrER 5
STUDY OF THE POSSIBILITY OF IMPROVING THE PHENYLFLUORONE METHOD FOR THE DETERMINATION OF TIN
5.1. Introduction
Since the development of the first spectrophotometric method
for tin, in 1952(1?4), about fifty different calorimetric reagents
have been used to varying extents for this determination. The
most significant of these reagents are dithiol, gallein, Catechol
Violet and phenylfluorone.
5.1.1. Phenylfluorone.
Various 2,3,7-trihydroxyfluorone dyes substituted at the C9
carbon atom have been reported to be useful as chromogenic reagents
for the photometric determination of various metal ions.
The absorption spectrum of phenylfluorone as well as the
absorption spectra of seven organic reagents and their complexes
with tin(IV) have been compared at different pH values by Babko
et a1( 175l. The optimum pH for tin(IV)-phenylfluorone complex
formation was found to be 1.0. 'At pH)2 anionic form of
phenylfluorone appeared. Picasso and Pizzimenti( 1?G)have used
this reagent for the determination of tin in steels in which the
concentration of alloying elements does not exceed 376. The control
of pH was found to be particularly critical (1-1.2) and the
possibility of elimination of interferences was also studied.
It was found that only molybdenum caused interference. Luke( 177)
isolated the tin by an acid sulphide separation and a double
carbamate chloroform extraction. An improvement in the specificity
121
of the phenylfluorone method can be achieved by developing the colour
in acid solution in the presence of oxalate ion and hydrogen
peroxide as masking agents. Quantitative tests showed that oxalate
reduces the intensity of the tin-phenylfluorone colour and
peroxide increases the intensity of the colour, hence the two
effects cancel each other and the colour produced in the presence
of the two masking agents is the same as that produced in their
absence. Increase in sensitivity is also obtained by extracting
the tin~Phenylfluorone complex into a small volume of methyliso-
butylketone. In this method germanium(IV) and antimony(Ill) give
positive interference and zirconium(IV) and hafnium(IV) cause
1 . htl 1 lt t b bt · d Luke{ 1?8)has determ<ned s ~g y ow resu s o e o a~ne • • tin
in organic and inorganic samples by applying the double carbamate
method. In this method, extractable metals such as copper, bismuth,
and mercury are separated from arsenic(V) and antimony(V) or tin(IV)
by solvent extraction with a chloroform solution of diethylammonium
diethyldithiocarbamate from sulphuric acid (1 + 9).· Following this,
the arsenic, antimony and tin are reduced to their lower valance
states with thioglycolic acid and then separated from nonextractable
metals by repeating the carbamate extraction. To minimize
interference from other metals, it was found desirable to develop
the colour at pH 1.8 instead of 3.1 as previously suggested. At
p!l 1.8 very satisfactory results have been obtained. However when
Bennett and Smith(179)re-examined the procedure they found that
the optimum wavelength was 530nm at pH 3.8. Luke(1SO)isolated
the tin by solvent extraction as the iodide and consequently
simplified the spectrophotometric methods for determination of tin
in steel i.e. it was possible to dispense with the sulphide
separation, the double carbamate extraction and the use of oxalate
122
and peroxide as
phenylfluorone.
complexing agents in colour development with
( 181). Cluley 1ntroduced pqenylfluorone as a
calorimetric reagent for germanium in 1951 and used gum arabic
as dispersant, and this has since been used almost universally
( 182) 0
in the tin procedure as well. Luke and Campbell stud1ed
the.isolation of germanium from other metals by carbon tetrachloride
extraction. In this method very rapid colour development is
achieved after adjusting the pH to 3.1. An extractive
separation and spectrophotometric determination of tin in
biological materials using phenylfluorone is given( 183)and the
conditions for increasing the stability and ensuring the best
reproducibility in the formation of tin-phenylfluorone were also
investigated and improvements achieved with respect to the
stability of the complex and accuracy of the results.
5.1.2. Effect of surfactants on reagent-metal complex system
A consideration of the effect of surface active agents on the
absorption spectrum of reagents of this type and on those of their
reagent-metal complex is very important.
' ( 184) • . 0
Dagnan, West and Young have exam1ned the sensitivity
effect of cetyltrimethylammonium bromide (CTAB) on the colour
reaction between tin(IV) and Catechol Violet. The optimum pH
for colour development was found to be 2.2. Ashton et al(1B5)
also studied the effect of CTAB on the tin-Catechol Violet complex
and developed a procedure which gave good results for steel
analysis and postulated a mechanism for the reaction. The molar
absorptivity for the sensitized reaction was found to be 92,000
l mol-\m-1 •
123
Gelatin has been used by Plotnikova( 1BG)in determination of
tin in· niobium-tin alloys using phenylfluorone. The alloy
samples were dissolved in sulphuric acid followed by hydrogen
bromide. Tin(IV) bromide was then distilled at ~ 200°C, and
ammonium hydroxide was added to an aliquot of the distillate
until alkaline (to Congo Red). To this solution, gelatin and
alcoholic phenylfluorone was added and the absorbance was
measured after 30 minutes. Phenylfluorone in methanol has also
been used for determination of tin(IV)( 187l. In this method
the addition of excess cetylpyridinium chloride and oxalic acid
were found to enhance the colour development of the tin(IV)-
phenylfluorone complex. The pH was adjusted to 1.0 using 3%
sulphuric acid, the solution allowed to stand for 20 minutes at
0 45 C and the absorbance was measured at 530nm.
5.1.3. Some studies in dimethylformamide(DMF)-water system
Dimethylformamide is an easily purified solvent and does
disso·lve a 'iarge number of inorganic compounds. The dielectric
constant of DMF (36.7 at 25°C) is such that ion-association does
not predominate, yet it is low enough that it is a good
differentiating solvent in term& of the types of conductance
equations which are most valid for the system( 188). Buncel et
(189-190) al have reported that aqueous DMF systems containing
hydroxide ions are unstable, the hydroxide ion being consumed
fairly rapidly under mild conditions with the liberation of
dimethylamine and formate ion. They have reported that DMF has
the property of greatly enhancing the rate of many reactions and
the use of aqueous-DMF media was suggested by the practical
consideration of increased solubility of reagents and products.
124
Due to the effect of sulphuric acid on DMF( 1SS) 1 hydrochloric acid
has been used to prepare the tin solution in the present work.
In the case of sulphuric acid, appreciable hydrolysis to formic
acid causes large errors. Wild( 191 )using a partially non-aqueous
system, namely 6~~ DMF reported some improvement in determination
of tin using a 1.5/1 v/v DMF-water solution of phenylfluorone.
In the present work by using 1~~ D!1F much better results have been
5.2. Experimental
5.2.1. Ethanol-water system:
Phenylfluorone reagent solutions were prepared by
dissolution of the sample in ethanol (2~~ and 40%) with the aid of
a few drops of concentrated hydrochloric acid. The absorbance
spectrum of phenylfluorone was measured at different pH values by
adding small volumes of hydrochloric acid (M/10) and sodium hydroxide
(M/10) as appropriate. The molarity of the reagent was always
6 -6 .24 X 10 M. The results obtained are shown in Table XXIX •
• An increase in pH leads to a decrease in absorbance. The blank
absorbance was quite high and the absorption spectrum at pH values
greater than 3.4 had two peaks.
125
TABLE XXIX
Variation of absorbance of ethanolic solutions of p·henylfluorone
with pH.
2~~ ethanolic ( I = 460nm) 1\ max
pH 1.47 1.63 2.13 2.54 3.42 6.42 11.31
absorbance 0.458 0.446 0.421 0.396 0.362 colour changes ·Two into pink peaks
40% ethanolic ( \ = 465nm) 1\ max
pH 1.22 1.31 2.26 2.7 3.5 4.42
absorbance 0.477 0.469 0.422 0.382 0.352 0.328 Two Two
peaks peaks
These ethanolic phenylfuorone solutions have been used for
determination of tin(IV) using phenylfluorone, v1ith and without
·addition of surface active agent: In the case of using 2~~
ethanolic solution as well as· 4~~. in the absence of surfactantf
the dye and complex bands overlapped and even after addition of
cetyltrimethylammonium bromide as surfactant, the complex bands
did not move to longer wavelength and no appreciably higher
absorbance was obtained. The results indicated that 2lf~ and 40%
ethanolic phenylfluorone were not suitable results for the req.uired
purpose.
126
10.48
red colour
5.2.2. Dimethylformamide systems
In carrying out the investigation, special regard was given
to reproducibility of results and molar absorptivity and the
stability of tin(IV)-phenJ'lfluorone system with time. All
measurements were carried out on Unicam SP8000 and SP600
spectrophotometer. The pH meter used was a Pye 290 with a
combined glass electrode.
Reagents
1) Standard tin solutions(~92 ):
a) Stock solution: Dissolve Q.500g analytical-reagent
grade granulated tin in 150ml hydrochloric acid (1/1 v/v) contained
in a 400ml beaker covered with a watch glass, by using gentle
heating. Wash the watch glass >ri th water and cool the solution.
Transfer the solution to 500m2 volumetric flask, dilute it to the
volume with distilled water. This solution contains 0.001g per
ml. of tin(IV).
b) Working solution: Transfer 10ml of stock solution
into a 1000ml. volumetric flask, add 100ml. concentrated hydrochloric
acid carefully, and dilute to volume with water. This solution
contains 10j1g per ml. of tin(IV).
2) Phenylfluorone (organic reagent, Hopkin and Williams Ltd).
Dissolve 0.05g phenylfluorone in dimethylformamide BDH
laboratory grade, add 1ml concentrated hydrochloric acid, dilute
to volume in 500ml graduated flask with DMF.
127
3) Cetyltrimethylammonium bromide (BDH laboratory reagent),
0.2'~ w/v in distilled water.
4) Acidified water, pH 1.1: Adjust the pH of water to 1.1
with dilute hydrochloric acid. If this solution has been stored,
it is advisable to check its pH before use.
5.2.4.Procedure:
Add in sequence 2ml aliquots of diluted tin(IV) solution,
5ml. of phenylfluorone in DMF, and 20ml. of cetyltrimethylammonium
bromide solution to a 50ml beaker. Mix the solution using a
magnetic stirrer, and adjust the pH value (using a glass electrode),
to 1.1 by adding molar hydrochloric acid by means of dropping
pipette. Transfer this solution to a 50ml. volumetric flask,
carefully wash the electrode and the beaker with small amounts
of dilute hydrochloric acid prepared to have a pH of 1.1. Dilute
to volume with the same acid solution. Read the absorption
spectrum on a SP600, after 30 minutes, using 1cm silica cells at
525nm. against water as reference. A blank solution is prepared
using the .same procedure but without adding tin(IV). The absorbance
spectrum of the complex is found'by difference.
5.2.5.stability of the tin(IV)-phenylfluorone complex with time
The stability of the tin(IV) phenylfluorone system in DMF~
water media and in the presence of CTAB as dispersing agent was
checked by measuring the absorbance at the various time intervals.
It was found that the absorbance was constant after nearly 30
minutes, for the full development of the colour for two hours. ·The
results are shown in Table XXX.
123
07
c.> u c
05
Eo5 '-0 Vl .c <(
04
03
02
01
Fig.9
Effect of C TAB on the colour r ec:Jction between tin!IV) and phenylfluorone
m Tin- phenylflo crone
0 Tin-phPnylfluorone sensitiz<?d with C TAB
450 500 550 600 "nm
01 u c
0-4
03S
Q.3
c 0.2S .0 ,_ 0 cJ)
.0 <( a2
QjS
o.1
nos
Tin(IV)- phenylfluorone system: The visible absorption spectra at various C TAB concentrations .,
li
' Cl
\
o) 2.2x 10- 3 M
11) 2.1 X 10-3 M
o) 1.6S x10-3 M
x) 1.1 x 10-3 M
Cl) o. ss X 1 0- 3 M
Fig.1 0
TABLE XXX
The time interval of the stability of tin(IV)-phenylfluorone system in DMF-water media in presence of CTAB
Time 0 5 10 15 20 25 (minutes)
Absorbance 0.316 0.318 0.320 0.324 0.325 0.326
Time 30 40 6o 120 (minutes)
Absorbance 0.327 0.327 0.327 0.327
5.2.6. The precision of the determination.
A series of determinations were performed using the method
described in 5.2.4. The molar absorptivity calculated from
10 determinations was found to be 95,000 1 mol-1cm-1 • The
standard deviation and confidence limits were also calculated and
the results obtained are:
Number of readings • 10
Average absorbance value 0.321
Standard deviation 5.1
9'Ji~ confidence limit on 5.22 average value
Conclusion and discussion.
Phenylfluorone appears to be the most popular reagent
for tin used in the British Steel Industry at the present time.
In view of this, an investieation of the tin-phenylfluorone 1•as
129
0.8
0.7 w u c 0.6 0 .c 0 0.5 Cl)
.c <( 0.4
Q3
0.2
0.1
F"ig,11
Calibration curve fortiniiVl.phenylfluoron/DMF
Different volume of tin+ 5 rrl of I 0.01% in D M F + 0.2 ml
cone. HCI + 20 ml C TAB I 0.2% in H20) I 50 ml vol flask
readings were mode at 525 nm ogoin~t water.
1 2 3 4 5
Volume of10/'g mt-1 tiniiV)
carried out, and a spectrophotometric method for a rapid
determination of tin was described. \-/hen phenylfluorone in
dimethylformamide was used as an analytical reagent, the
reproducibility of results was better than when the reagent is
used in water. The sensitivity of the colour reaction between
tin(IV) and phenylfluorone has been increased by the addition of
CTAB. (Fi~ IX). The spectrum of the tin-phenylfluorone complex
varies markedly with increasing concentration of quaternary
salt at a fixed pH. Fig(X),presents some typical data to illustrate
the effects observed when the quaternary concentration is
progressively increased while the acidity of the solution is
maintained at pH 1.1. The various spectra are for different
concentrations of the quaternary salt in the solution. The
recommended concentration is 2.1 to 2.2 x 10-3H. Higher
concentrations lead to an increasing turbidity due to the
insolubility of the CTAB at those concentrations. Beer's Law
is obeyed from 0-S~g of tin(IV) per ml. Fi$(XI). The final
method is simple and is suitable for lo~1 concentrations.
Although a higher apparent molar absorptivity is obtained in
the present procedure compared l<i th the method using Catechol
Violet( 1&35l, but is not consid~red to be an improvement since
its success is dependent on the critical control of pll and CTAB
concentration.
The method which has been developed for the determination
of tin could be regarded as a method which may be useful under
certain circumstances for steel analysis. The study was made
using standard tin solutions and interfering metals have not been
0 ( 1 &30) cons1dered, but Luke has studied interfering metals and
their removal.
130
CHAPTER 6.
THE DEVELOPI1ENT OF AN ION-SELECTIVE ELECTRODE RESPONSIVE TO PERIODATE.
6.1. Introduction.
The response of a glass membrane electrode to change
in the activity of hydrogen ions in solutions was mentioned as
long ago as 1909(212 ).
Hith the development of electrode technoloey over the
past 15 years, efforts have been made by different workers to
introduce new types of electrodes which were easy to prepare
and use and in addition 11ere more selective and durable from
the chemical and mechanical points of vie11 respectively than
glass membrane electrodes. This work led to the invention of
new types of electrodes based on different membranes. These
membranes take the form of either ion-conductive crystals or
inert hydrophobic membranes, the latter being either impregnated
with a solid ion-sensor or saturated with a solution of a
particular active material dissolved in a non-aqueous phase.
Such membranes by virtue of the ion-conductivity of the sensor
present in them distinguish a particular ion in solution and
respond to its activity. Electrodes with such properties are
known as Ion-selective Electrodes. In the last few years
a considerable number of such electrodes responsive to a
wide range of different cations and anions have been developed.
The construction of these electrodes is n?t restricted to a
particular material, shape, size or design.
131
The development of this new analytical sensor has given an
increasing importance to analytical potentiometry as the range
of the technique has been extended greatly and has created a
great deal of interest both theoretical and practical aspects
of the subject. The ion-selective electrode forms an
electrochemical half cell responding to a particular ion in
the ·external solution under test.
The classification of membrane electrodes has been
. · th (fi:S2) db Al S"b .(194) d b of d1scussed by Pa an an y - 1 aa1 . an a num er
electrodes responsive to different ions have been developed
· th" 1 b t b F t 1 (202,203,204,205,206,62,63,64) 1n 1s a ora ory y ogg e a •
Due to the rapid development of the field of ion-selective
electrodes, a vast number .of papers and reviews have appeared·
in the literature in a relatively short time. The most
h · · ·a a t b th f B k( 195- 196) compre ens1ve rev1ews are cons1 ere o e ose o uc ,
(197) (198) . (199) Moody and Thomas , Koryta and Cov1ngton • Also,
·two text books, "Ion-Selective Electrode", and "Selective Ion-
Sensitive Electrodes" are the main texts on the subject.
In this short review only electrodes developed in this
laboratory will be discussed.
A silicone-rubber electrode responsive to cationic
detergents, based on hexadecyltrimethylammonium dodecylsulphate,
. (202) was descr1bed • Nernstian response v1as obtained over the
range 10-3 to 10-5M of hexadecylpyridinium bromide and
hexadecyltrimethylammonium bromide. The electrode was used
in the potentiometric titration of sodium dodecylsulphate,
sodium tetraphenylborate, ammonium reineckate, potassium
132
hexacyanoferrate(III) and potassium dichromate using
hexadecyltrimethylammonium bromide as titrant. A liquid state
(203) electrode selective to molybdenum 11as developed , based
on bis-tetraethylammoniumpentathiocyanatooxomolybdate(V)
dissolved in nitrobenzene: a-dichlorobenzene (2:3). This
electrode had Nernstian response over the range 10-2 to 5 x 10-SM
molybdenum. Iron, vanadium, tungsten, niobium and rhenium,
11hich form thiocyanate complexes, interfere.
Brilliant Green 11as used successfully to.prepare electrodes
selective to tetrathiocyanatozincate(II) (61 ), perchlorate(62),
. (6 ) and tetrafluoroborate 3 ions. The tetrathiocyanatozincatc
electrode 11as used to determine zinc(II) in the presence of
-1 -4 thiocyanate ion over the range 10 to 10 H of zinc(II). The
electrode did not respond to zinc or thiocyanate ion 11hen either
of them 11as present in the solution alone. No interference was
observed froin copper(II), lead, nickel and copper(I) but iron(III)
and cobalt(II) interfered. From the active material, 11hich ~Jas
the ion-association complex formed bet11een the tetrathiocyanato-
zincate ion and Brilliant Green, liquid state, heterogeneous
silicone rubber and carbon pas~e electrodes 11ere also prepared
and examined. The best performance 11as observed using the liquid
state electrode. The perchlorate electrode, which was based on
Brilliant Green perchlorate, responded to perchlorate over the
range 10-1 to 10-4H of perchlorate with no significant interference
from bromide, acetate, chloride or fluoride but 11ith a little
interference from iodide, hydrogen carbonate and nitrate ions.
The electrode 11as used in potentiometric ti trations of perchlorate
with tetraphenylarsonium chloride. The tetrafluoroborate electrode
133
-1 -4 -responded in the range of 10 to 10 M of BF4 • The only
serious interference being the perchlorate ion. Fogg et al(Z04)
have also assessed a silicone rubber membrane containing
potassium zinc ferrocyanide as ion-selective electrode sensor
for the determination of alkali metal ions. The slope of the
calibration graph for potassium ion t<as found to be 59mV per
decade change in concentration t<ithin the range 5 x 10-4 to 10-1M
0 at 25 C. Similar membranes prepared by the same authors from
PVC responded similarly ;Ji th a slight improvement in selectivity.
The basic dyes, Sevron Red L, Sevron Red GL, Flavinduline 0
and Phenazinduline 0, have been applied in liquid-state ion
selective electrodes for the determination of antimony and
thallium(64l. This electrode is response to antimony(III) as
well as to antimony(V), and behaves in a Nernstian manner.
Ion selective electrodes for the determination ·of perrhenate(Z05)
and tetrachloroaurate(ZOG)using Brilliant Green and Safranine 0
respectively have been developed by Fogg et al.
Current interest in research in the development of ne1-1
ion-selective electrodes for a variety of ions lies mainly in
their useful application in potentiometric analysis which is
• comparatively simple, inexpensive and readily automated. The
present 11ork on ion-selective electrode was undertaken in order
to study some onium compounds as active materials for ion-selective
electrodes. To our knowledge no previous attempt has been made
to prepare an ion-selective electrode responsive to periodate.
Therefore the possible preparation of a liquid state ion-selective
electrode based on the onium-periodate ion-association complex
was attempted.
134
6.2. Experimental
6.2.1. Possible precipitation and extraction of salts of different onium ions with periodate into a-dichloro
benzene.
A series of 10-2M onium compounds (Haybridge research
chemical) were used as precipi tants for 10-211 sodium periodate.
For those onium compounds which showed any sign of precipitation,
further study was performed to see >lhether the periodate was
extractable into a-dichlorobenzene. The results are summarised
in Table (XXXI).
6.2.2. The electrode assembly.
The liquid state electrode assembly used in this
study has been described earlier by Fogg et al(11 ). The
electrode body was fabricated from P. T .F .E. tube (-~11 internal
and 1.;[-11 external diameter). A et" section was cut from the end
of the tube and threads were cut into two parts so that they
can be screwed together and hold a rubber membrane of et" diameter.
Electrical contact with the back of the membrane was made with
an .;-n diameter carbon rod >lhich just fitted the tube and which
was held firmly in place by a narrow nylon screw passing through
the main body of the electrode. Connection with the pH-meter
was made through a coaxial cable connected to the carbon rod
by means of a conducting thermosetting silver resin preparation
(Johnson Matthey Metal Limited). A commercial natural rubber
(Harboroue;h Rubber Company) of 2.70mm in thickness as well as
a natural rubber, especially prepared for this purpose in the
Department of Polymer Technaloe;y, were used to prepare the
membranes for the electrode.
135
-----Coaxial wire
'1----Teflon
' • I
--Carbon rod
--Cap
Natural-rubber membrane liquid-state- electrode .•
TABLE(XXXI)
Possible precipitation and extraction of salts of different onium ions with periodate into a-dichlorobenzene •
.
Onium Compound . Precipitation observation
Extraction Observation
Triphenylmethallylphosphonium chloride p phase miscib
Tris(trichlorophenyl)ethylphosphonium p CE . iodide
Tetraphenylarsonium chloride p . NE. .
3,5 di-t-butyl-4-hydroxybenzyltriphenyl- p NE phosphonium bromide
Tetraphenylphosphonium chloride p NE
Sodium tetraphenyl boron NP NE
Tetrabutylphosphonium chloride NB NE
Tri-n-butylmethylphosphonium iodide p CE
(Methyl)-triphenylphosphonium bromide NP NE .
Triphenyl-n-butylphosphonium bromide p NE
Tri(3 chlorophenyl)methylphosphonium iodide p NE
Tri-n-butylbenzylphosphonium chloride NP NE
Diphenyleneiodonium bisulphate NP NE •
Key:
P precipitate
NP No precipitation
CE complete extraction (by fading the yellow colour in
NE no extraction
136
aqueous phase and appearing in organic phase).
I
le!
I
1
I
It is apparent from the table that only Tris(trichlorophenyl)
ethylphosphonium iodide and Tri-n-butylmethylphosphonium iodide
gave satisfactory results.
6.2.3. Preparation of the membrane and the use of the electrode.
A solution of the onium periodates in a-dichlorobenzene
was prepared as follows:
A ~-11 disc rubber membranes used in the present
study were soaked in a saturated solution of the particular
onium-periodate in a-dichlorobenzene overnight or longer
depending on the onium compound used. Before use the soaked
rubber was dried on a tissue and inserted directly into the
electrode body. The membrane was used in· the eiectrode assembly
described above. The resulting ion-selective electrode was
used in an electrochemical cell which may be shown diagramatically
as:
Ion-Selective electrode Sample solution
Reference electrode
The temperature of the cell was maintained at desired temperature
using a flow-through jacket beaker, a small >later pump and
a thermostatically controlled water bath. This system controlled
the temperature of the solution in the beaker within ~ 0.2°C.
An electromagnetic stirrer was used for stirring purposes. The
potential measurements were made with n Radiometer PHM64 Research
pH-meter versus a saturated calomel electrode.
137
6.2.4. Evaluation of the electrode.
Analytical reagent grade sodium periodate was
-2 dissolved in tridistilled 11ater to give a 10 M standard
stock solution. Subsequent solutions were prepared by serial
dilution. It l<as not possible to prepare solutions of concentration
of 10-2M of tris(3-chlorophenyl)ethylphosphonium iodide and
tri-n-butylmethyl-phosphonium iodide due to the problem produced
from the low solubility of these compounds in water so saturated·
solutions were prepared and 2:1 v/v ratios of onium-periodate
are extracted into a-dichlorobenzene. The response of these
electrodes are shol<n in Table ')(X)( 1/ a_ •
TABLE XXXI!a.
Response of the electrodes to1~ards periodate solutions.
These potential values were obtained immediately and only
gave indication of the possible usefulness of the electrode.
The potential dr.ifted over a period of minutes and this is discussed
later.
• Potential(mV) Concentration(M) TCEPI TBMPI
10-2 35.2 -62.4
10-3 -74.1 -70.1
10-4 -109 -74.2
10-5 -130 -78.0
10-6 -138.4 -80.5
It is apparent that a very 10~1 potential changes per decade
were obtained when TBMPI 11as used. Therefore further studies
"ere carried out using TCEPI.
138.
6.2.5. The effect of length of time of soaking in organic solvent.
Table(XXXII) shows the potential concentration slope fo""
(()3:...10-2M sodium periodate solution at different time intervals
of electrode being soaked in a-dichlorobenzene.
TABLE( XXXII)
Effect of potential with time.
Soaking time Potential concentration slope (days) mV per decade after:
0 3(min) 10(min)
1 49.1 38.3 44.5
2 48.0 42.1 33.7 '
5 44.3 3!3.7 36.3
15 40.2 3:;\.1 32.9
30 39.5 39.1 37.2
6.2.6. Stability of the electrode response.
In order to see the effect of age of conditioning
• on electrode response, a check was made at different time
intervals. The results are recorded in Table(XXXIII).
6.2.7. Electrode response.
The results obtained in 6.2.4. were made with no
conditioning. The electrode 11as now studied with an electrode
-2 which had been conditioned in 10 M periodate solution for seven
days. The results obtained are shown in Table(XXXIV).
139
TABLE(XXXIII)
Stability of electrode response.
Days. 0 1 2 4 5 6 7 8 10 13 15 19
Slope(mV per decade) 39.5 43.2 45.5 48.6 46.3 49.1 53.2 55.4 53.6 54.7 53·3 52.3
Days 29 33 42 48 54
Slope(mV per decade) 55.4 55.1 53·6 54.8 55.6
From the above it is clear that maximum response (above 54mV per decade) is only obtained after
7 days of conditioning in 10-~ periodate solution.
25
54.5
TABLE(XXXIV).
Response of the periodate-TCEPI electrode to periodate,
- \
!04 concentration (H) ·' Potential(mV)
just after the after 7 days be11;1g electrode prep- left in 10-2]1 ro
4-
aration
10-2 -104.1 -142.9
10-3 - [};.) .:q -Tqr.:g
10-:4 - j g!. 1 -245. s
10-5 - ::coq-; o - 271.8
10-6 0 ~
-21 if, g -233·0
From the above it is clear that the electrode gave nearly 38mV
-2 -4 per decade change in concentration in the range 10 to 10 H of
periodate solution without conditioning and nearly 54mV per
decade change in concentration in the range 10-2to 10-311 periodate
solution after seven days conditioning.
6.2.8. Study of the interfering ions.
Possible interference by certain ions was studied
by observing the effect of concentrations in the range 10-6 - 10-2J.I
of sodium sulphate, nitrate, chloride and iodate as well as
potassium chloride on the value of the potential obtained with
periodate electrode in contact with an 10-2J.I periodate solution.
The change in potential >las very small (0-3mV) indicating
negligible interference by these.ions.
141
6.2.9. Applications of TCEPI-periodate electrode in potentiometric titrations.
Solutions of 10-2M and 10-3M of periodate were titrated
with 10-2M solutions of tetraphenylarsonium chloride, glycerol(20?)
and o-phenylenediamine( 20S) and 10-311 solution of CTAB using the
periodate selective electrode to detect the end point. The
end points in each case were determined by means of first
derivative curves.
All the solutions ~<ere prepared in tri-distilled 11ater
and used for the study of the electrode performance. It was not
feasible to prepare solutions of concentrations of 10-~l CTAB
due to problems of precipitations, and therefore in this procedure,
50ml aliquots of 10-3H periodate was used against 10-3H CTAB as
titrant. In the case of the potentiometric titrations using
tetraphenylarsonium chloride, a-phenylenediamine and glycerol,
50ml aliquots of 10-2H periodate were titrated. Typical
potential-volume data for these titrations are given in Table
XXXV, XXXVI, XXXVII and Fig. XII, XIII, XIV and XV.
6.3. Conclusions and discussion.
The liquid state electrode described earlier is based on
a water insoluble periodate-tris(3 chlorophenyl)ethylphosphonium
ion-association complex. 1'he electrode gave nearly 54mV per
decade change in concentration in the range 10-2 to 10-311 of
periodate ion after being soaked in the organic solvent for
-2 30 days and being conditioned for at least 7 days in 10 H
periodate solution. The electrode gave a stead.y response
142
TABLE( XXXV)
Potentiom.etric titration of sodium periodate with glycerol.
I II
Amount of glycerol Potential added (ml). (m V)
Amount of glycerol Potential added (ml). (m V)
- -110.0 - -117.5
3 -115.1 3 . .-122.0
6 -119.2 6 -126.6
12 -130.4 12 -137.6
18 -145.2 18 -153-2
21 -156.8 19 -156.4
24 -176.4 20 -160.5
25 -190.1 21 -165.0
27 -226.9 22 -169.8
28 -230.2 23 -175.6
29 -233.0 24 -185.4
30 -235.0 25 -203.0.
31 -235.8 26 -212.8
32 -237.0 27 -230.0
33 -237 .o 28 -240.9
34 -236.6 30 -256
35 -.:.235. 9 35 -262
38 -236.1 4o -263
4o -235.0 45 -258.3
143
TABLE( XXXVI)
Potentiometric titration of sodium eriodate with tetraphen larsonium .chloride I) and o-phenylenediamine(II).
(I) (II)
Amount of tetraphenyl Potential Amount of o-phenyle- Potential arsonium chloride (mV) nediamine added (m V)
added(ml). (ml)
- -144.3 - -192.0
5 -149.1 1 -193.2
10 -153.0 3 -195.7
12 -162.7 5 -198.8
15 -168.4 8 -203.9
18 -176.3 10 -208.0 -
20 -182.5 12 -212.1
21 -214.1 15 -218.3 -
22 -268.0 18 -226.8
25 -298.6 20 -233.4
26 -304.3 21 -237-9 .
-- ---27 -308.6 22 -242.8
28 -312.3 23 " -241>.0
·-30 -323.5 24 -255.3
4o -334.7 25 -262.9
45 -336.0 26 -269.0 '
27 -275.6 .
30 -274.8
35 -277.0
144
TABLE(XXXVII)
Potentiometric titration of 10-3M sodium periodate with 10-311 CTAB.
Amount of CTAB added Potential (ml). (m V)
- -185.4
2 -~90.4
5 -200.5
6 -205.0
7 -209.5
s -213.&
10 -224.8
12 -240.7
14 -255.5
16 -263.8
18 -267.9
20 -269.4
25 • -270.4 -
30 -270.8
40 -271.2
12 0
140
:> 1.60 -E --0
~1.80 (!) -0 a.
2.00
2.20
240
Fig. 12
Potl:!ntioml:!tric titration of 50ml of 10-2 M sodium pc:!riodatl:! with 10-2 M glycl:!ro\
10 20
m at 35°C (!I)
0 at 25° C I I l
··~ . . ......
30 40 50 Volume:! of glycl:!rol addc:!d(m\1
• 1. '
1 40
-; 160 E = 180 Cl
c 200 01 .... 0 2 20 n.
240
260
280
300
320
Fig. 13
PotC?ntioml2tric titration of 50ml of10-2M sodium pC?riodatC? with10-2M.TPAC .
10 20 30 40 50 Volum12 of TPAC added(mll
0
""' c (\) 2 00 ..... 0
0..
220
240
2 60
280
Fig.14
Pote?ntiometri c titration of 50 ml of 10-3 M sodium pe?riodcrte? with 10-3M c TAB
10 20 30 Volume of C TAB added (m\)
40
180
-> 200 E -~
~220 ... D L
240
260
Fig.15
Potentiomcatric titration of 50 ml of 10- 2 M
sodium pcariodatewith10-2M o-phenylcanediamine
• 10 20 30 40
Volume of a-phenylenediamine added(mll
immediately on contact with the periodate solution. The slope
of the periodate response fell to 26mV at 10-4 to 10-5M.
No interference effect was observed in the presence of
sulphate, nitrate, chloride and iodate anions as well as
potassium cation. The potentials were almost reproducible
on the same day of working, but for a precise measurement
it is adviseable to calibrate the electrode every day because
electrode has shown drifts in potential at the rate of approximately
3mV per day.
The electrode has been used for potentiometric titrations
(209) . (207) of periodate with tetraphenylarsonium , CTAB, glycerol
and o-phenylenediamine( 20S). From the values of the end points
obtained, the stoichiometry of the reaction between periodate
with CTAB, a-phenylenediamine, tetraphenylarsonium and glycerol
was found to be 2:1 ratio of periodate/titrant. In the case of
using glycerol, the results obtained were in good agreement with
Malaprade( 20?)reaction and very satisfactory results were obtained.
No information was found to explain the reason for reaction taking
place between a-phenylenediamine and periodate • •
From the literature survey, it was found that although
Laurie et al(209)studied the solubility of periodate-tetraphenyl-
arsonium in aqueou~ solutions, they gave no information about the
stoichiometry of the reaction. Buist et al(210 )observed the
dimerization of periodate in aqueous solution. Further confirmation
of dimer formation came from potentiometric titrations of periodic
acid at various concentrations and temperatures.
dimer was suggested to be the dimesoperiodate ion
146
The probable
lf(041-0-104) •
Owing to lack of time in the present studies, it was not
possible to study this mechanism further. 11ore lvork will have
to be done to explain these results properly. It is possible
that the end point of the periodate-tetraphenylarsonium titration
could be due to the formation of a salt RH3r2o
9, but no salts
of this form have been reported previously.
This electrode could be used for the reactions involving
the selective oxidation of organic
groups attached to adjacent carbon
compounds having hydroxyl
(207) atom , for example the
estimation of glycerol in fermentation solutions(211 ).
147
CHAPTER VII
FINAL CONCLUSION AND DISCUSSION.
The ion-association-extraction technique is widely used
in analytical procedures. A large cation is used to extract the
anion to be determined from water into an organic phase as an ion
pair. Two types of reagents are in common use, the onium
compound and the basic dyes.
The original aim of this study was to extend further the
applications of these ion-assocation methods and to improve
existing methods, possibly by the introduction of new onium
compounds. The onium compound that had not previously been used
for analytical purposes was prepared and tested for this purpose.
The use of onium compounds in solvent extraction procedures
for the determination of dichromate \~ere investigated involving
an investigation of the possibility of applying the solvent
extraction technique to atomic absorption.
The use of CTAB with phenylfluorone in the determination
of tin(IV) and basic dyes to t~e determination of phosphate
was investigated.
Finally the possibility of using onium salts as an ion
selective electrode responsive to periodate was investigated.
A general survey of the extraction of dichromate ion ~1i th
different onium salts revealed a possible method for the
determination of dichromate. This was investigated further using
various onium compounds, and different organic solvents. A
148
I
I
I
procedure has been described for the determination of dichromate
as the TBBP-dichromate ion-association complex extracted into
chloroform. In the present study, in spite of the low standard
deviation as well as the reproducible results obtained, the apparent
molar absorptivity was similar to that for other methods for the
spectrophotometric determination of dichromate and ~Jas not an
improvement on these methods.
The extraction method has also been used in the development
of a new atomic absorption method of determining chromium.
Amongst the reported interferences, perhaps the most inconvenient
and difficult interference to overcome is that of iron on chromium.
This interference is completely eliminated with the present
procedure by addition of sodium fluoride to complex the iron(III).
Solvent extraction of chromium(VI) from aqueous solution into
11IBK increased the absorbance of chromium(VI) sixtyfold. The
effect of nitric and sulphuric acids was studied and no absorption
due to the extraction of chromium(VI) into 11IBK was obtained in
the presence of these acids, but by addition of hydrochloric acid
into both solutions, complete extractions were obtained. This
effect is clearly because of tne presence of chloride ion being
essential for the extraction. The optimum hydrochloric acid
concentration was found to be 1 - 2!1. The low results obtained
in higher concentrations of hydrochloric acid ,;as due to the
reduction of small amounts of chromium(VI) to chromium(III).
The absorbance-concentration curve was linear up to 12}1 per ml
of chromium and passed through the origin. This method has
been applied satisfactorily to several British Chemical Standard
Steels. The procedure has the advantages of using an air-acetylene
1.49.
flame and being free from interference from iron.
During this work it was found that, although standard
solutions of Chromium(VI) were stable for 3-4 hours, the solutions
prepared from steel samples had to be sprayed into the flame
within 5 minutes of extraction, because the absorbance of these
solutions decreased after this time. The reason for this is
not kno1m but could be due to the presence of other metals
present in the steel samples. Further 110rk would need to be
carried out to explain the reason for this decrease.
A .. sensitive method for rapid determination of inorganic
phosphate in biological systems is described 11hich is based on
a reaction between molybdophosphate and Crystal Violet.
Tetrachloroacetic acid is used for the extraction of phosphate
from biological materials particularly sera without interference
in the photometric determination. The present method requires
a protective colloid .in order to avoid precipitation of the
dye salt formed. Polyvinyl alcohol was found to give excellent
results. Other materials commonly found in plasmas do not
interfere. In this study the method of Altmann et al was
investigated and the optimum experimental conditions were •
obtained. The method of Altmann et al has the disadvantage of
a one hour colour development period. In the. present study an
attempt was made to reduce this development time. By using
Crystal Violet in place of Halachite Green a 20 per cent increase
in sensitivity was obtained, and by heating the mixed solutions,
the development time VIas reduced to 20 minutes.
Previous workers have studied phenylfluorone as an analytical
reagent. In the present v1ork, it VIas found that the use of
150
phenylfluorone in D~F instead of water, improved the
reproducibility of the method. The sensitivity of the colour
reaction between tin(IV) and phenylfluorone has been increased
by addition of CTAB. The visible adsorption spectra of the
tin(IV)-phenylfluorone-CTAB system under conditions of various
CTAB concentration was obtained. A CTAB concentration of
2.1 - 2.2 x 10-3M is that used in the solutions for the
spectrophotometric determination of tin, and found that this
concentration was the optimum for maximum colour development.
The role of CTAB in this reaction is explained by dispersion of
the insoluble ion-association complex. Beer's law is obeyed
from 0.2 - o.8~g per ml of tin. The final method is simple
and is suitable for the determination of low concentrations of
tin(IV). Its main disadvantage is that the concentration of
CTAB and control of pH are particularly critical.
Finally a liquid state electrode based on the water
insoluble periodate-tris(3 chlorophenyl)ethylphosphonium
ion-association complex was prepared. The electrode gave nearly
54mV per decade change in concentration in the range 10-2 to
.10-3M of periodate ion. No interference effects were observed
in the presence of sulphate, nitrate, chloride and iodate anions
as well as potassium cation. The electrode has been used for
potentiometric titrations of periodate with tetraphenylarsonium,
glycerol, CTAB and a-phenylenediamine. From the values of the
end points obtained, the stoichiometry of the reaction bet1~een
periodate with these titrants was found to be 2:1 ratio of
periodate/titrant. Satisfactory results ~1ere obtained in the
potentiometric titration of glycerol with periodate. Owing to
lack of time in the present study it was not possible to study
151
the mechanism of these reactions, although dimer formation
of periodate in aqueous solutions may provide the solution.
Further study of this system tvould be required to confirm
these findings.
It seems·that a great deal of work has been done on the
determination of anions using onium compounds and basic dyes.
The use of readily available reagents of these types has been
considered by many workers and any advances in onium salt
reagents or basic dyes ;rill probably utilise compounds that
have to be synthesised. Diphenyleneiodonium bisulphate was
considered to be a possible useful reagent in the present study
but unfortunately this reagent proved to be disappointing due
to its low solubility in water._ It is possible that other
ne\" onium compounds may have slight advantages over present
reagents.
Amongst traditional basic dyes, the potential for research
in the solvent extraction spectrophotometric field is almost
exhausted, but many of the methods already published, particularly
those involving bulky anions and ternary complexes, may ;rell
prove suitable for adaption to ion-selective electrodes. Some
work has already been undertaken in this field and in this
laboratory by Fogg et al but many systems remain to be
investigated.
152.
-----Coaxial wire
:t---Carbon rod
--Cap
~--Membrane
Natural-rubber membrane liquid-state electrode .•
REFERENCES
1. NERNST, W., z. Phys, 1891, 8, 110 •
2. HILDEBRANb,J .H. and ROTARIU, G.H., Analyt.Chem., 1952, 24, 770 •
3. BJERRUM, N., K. DANSKE VIDENSK. SELSK., 1926, 7, No. 9 "'Selected papers", p. 108 Einar !1unksgaard, Copenhagen, 1949.
4. MURATA 1 K., YOKOYAI1A, Y. and IKEDA, S., Analyt.Chem., 1972, 44, 4, 805.
5. HURATA, K. and IKEDA, S., Bunseki Kagaku, 1969, .:@_, 1137 •
6. STARY, J., The Solvent Extraction of 11etal Chelates, 1964 Pergamon, London.
7. FREISER, H., Analyt.Chem., 1962, 34, 64R; 1964, 36, 93R; 1966, 38, 131R.
8. Idem., ibid., 1968, 40, 522R.
9. BROOKS, R.R. and SMYTHE, L.E., Talanta, 1975, 22, 495.
10. KATZ, S., Am.Lab., 1972, 19 •'
11. BAILES, P.J. and WINWARD 1 A., Trans. Inst.Chem.Eng., 1972, 50(4) 240·
' 12. STARY, J., MTP Int.Rev.Sci: Phys.Chem., Ser.one, 1973, 12, 279 •
13. IRVING, H., Chem.Ind., 1974, (16), 639.
14. 11ERKIN, E.N., IVANOVSKII, !1.D. and BORBAT, V.F;, Radi9khimya, 1966, §., 705.
15. GINDIN, L.l1., BOBIKOV, P.I., PATYUKOV, G.l1., ROZEN, A.M., KOUBA, E.F •. and BUGAEVA, A. V., Ekstraktsiya, Teoriya, Premenenie, Apparatura, Sb.Statei 1962, 2, 87.,
16. KERTES, A.S., Recent Advances in Liquid-Liquid Extraction, Edited by Hanson, c., First edition, 1971, Pergamon Press.
17. FREISER, H., An Introduction to Separation Science, edited by Karger, B.L., Snyder, L.R., and Horwath, C., 1973, l~iley, J. Publication.
18. BERTHELOT, 11. and JUNGFLEISCH, J., Ann.Chim.et.Phys., 1872, 26, 396.
153
19. BOWD, A.J., THORBURN BURNS, D, and FOGG, A.G., Talanta, 1969, .!.§., 719.
20. HEAL, H.G., J.Chem.Edu,, 1958, 35, 192.
21. WILLARD, H.H. and SMITH, G.M., Ind.Eng.Chem.Anal,Ed., 1939, 11, 186.
22. TRIBALAT, S. and DUVAL, C., Anal.Chim.Acta, 1952, 6, 138.
23. TRIBALAT, S., ibid.' 1949, 3, 113.
· 24. RICHARDSON, M.L., Analyst, 1962, 87, 435.
25. MURPHY, J.W. and AFFSPRUNG, H.E. Anal.Chem., 1961, 33, 1658.
26. NEEB, R., Z.Anal.Chem., 1957, 157, 24 •
27. Idem., ibid., 1960, 177' 20 •
28. AFFSPRUNG, H.E., BARNES, N.A., and POTRATZ, H.A., Anal.Chem., 1951' 23, 1680.
29. · AFFSPRUNG 1 H.E. and MURPHY, J.W., Anal.Chim.Acta, 1964, 30, 501 •
30. FOGG, A.G., l1ARRIOTT, D.R. and THORBURN BURNS, D., Analyst, 1970, 95, 854.
31. FOGG, A.G., KUMAR, J.L. and THORBURN BURNS, D., Analyst, 1975, :!..9£' 311 •
32. TRIBALAT,.S., Anal.Chim.Acta, 1950, 4, 228.
33. NEEB, R., Z.Anal.Chem,, 1957, 154, 17 •
. 34. DWER, F.P. and GIBSON, N.A., Analyst, 1951, 76, 548.
35. RICHARDSON, M.L., Analyst, 1962, 87, 435·
36. WILLARD, H.H. and PERKINS, L.R., Anal,Chem., 1953, 25, 1634.
37. GIBSON, N.A. and WEATHERBUilll; D.C., Anal.Chim.Acta, 1972, 58(1), 149 •
38. BOWD, A.J. and THORBURN BURNS, D., Micro.Chim.Acta, 1965, 1151
39. Idem., ibid., 1966, 539.
4o. Idem., ibid., 1966, 1051 •
154
41. Idem., ibid.' 1967, 539.
42. Idem., ibid.' 1967, 560.
43. Idem., ibid. 1967, 564.
44. BOWD, A.J., Ph,D. Thesis University of London, 1967.
45. SCHWAB, G.M. and PECHLIVANIDIS, J., Svensk.Kem.Tidskr., 1947,59,130 •
46. BERINGER, F.M., DREXLER, M., GINDLER, E.N. and LlJ}lPKIN, C.C., J.Amer.Chem.Soc., 1953, 75, 2705.
47. HIGGENS, C.T., M.Sc. Thesis, LUT, 1969.
48. GIBSON, N.A. and WHITE, R.A., Anal.Chim.Acta, 1955, 12, 115
49. Idem., ibid.,
50. ENCYCLOPAEDIA BRITANNICA, 1970, 70, 814 •
51. VENKATARAMAN, K., The Chemistry of Synthetic Dyes, 1952, Academic Press (New York).
52. STOCK, J.T. and PURDY, vi.C., Chemist Analyst, 1959, 48{1), 22 •
53. WILLCOX, A., Ph.D.Thesis, LUT, 1974 '
54. BLYUM, L.A. and PAVLOVA, N.N., Zav.Lab., 1963, 29(12), 1407.
55. BLYUM, I.A. and OPARINA, L.I., ibid., 1970, 36(8), 897 •
56. FOGG, A.G., BURGESS, C. and THORBURN BURNS, D., Talanta, 1971' 18, 1175.
57. EEGRIIVE, E., Z.Anal.Chem., 1927, 70, 400
58. BURGESS, C~, Ph.D.Thesis, LUT, 1971.
59. FOGG, A.G., BURGESS, C. and THORBURN BURNS, D., Analyst, 1970, 95, 1012.
60. ENTWISTLE, J.R. And HAYES, T.J., Abstracts of papers, IUPAC, "International Symposium on Selective Ion-Sensitive Electrodes" Cardiff, U.K. April (1973), paper 17.
61. FOGG, A.G ... DUZINKE\ITCZ, M· and PATHAN, A.S., Anal.Lett., 1973, 6(12), 1101.
155
62. FOGG, A.G., PATHAN, A.S. and THORBURN BURNS, D., Anal.Chim.Acta, 1974, 73, 220.
63. Idem., Analyt,Letters, 1974, z, 545.
64. FOGG, A.G., AL-SIBAAI, A.A. and BURGESS, C., Analyt.Letters, 1975, _Q(2), 129.
65. SANDELL, E.B., Calorimetric Determination of Traces of Metals, 1949, Interscience, New York.
66. BERNHARDT, H.A., U.S. Atomic Energy Comm. Rept.MDDC, 1947, 1541.
67. DEAN, J.A. and BEVERLY, M.L., Analyt.Chem., 1958, 30, 5, 977•
68. BOOK, R. and HUMMEL, C.H., Z.Analyt.Chem. 1 1963, 198, 176 ·-
69. BOCK, R. and JAINZ, J., Z.Analyt,Chem., 1963, 198, 315.
70. ZEIGLER, M. and POHL, K.D., Z.Analyt 1Chem., 1964, 204, 413.
71. SHEVCHUK, I.A. and SIMONOVA, T.N., Ukrairt.Khim.Zh., 1964, 2£, 983,
72. CHALMERS, R.A. and DICK, D.M., Anal.Chim.Acta., 1965, 32, 117•
73. MAECK, I~.J., KUSSY, M.E. and REIN, J.E., Analyt.Chem., 1962, 34, 1602.
74. BARRESWIL, L.C.A., Ann.Chim.Phys., 1947 1 20(2) 264 •
75. SASTRI, M.N. and SUNDAR, D.S., Chemist Analyst, 1961, 50, 101
76. SASTRI, H.N. and SUNDAR, D.S., Z.Anal.Chem., 1963, 95, 343 •
v
77. ADAM, J. and PRIBIL, R., Talanta, 1971, j.Q, 91.
78. RICHARDSON, H.L., Analyst, 1962, 87, 453.
79. BACZUK, R.J. and DUBOIS, R.J., Anal.Chem., 1968, 40, 685.
80. COLLETT, J. and McGREEN. D.J., J.Am.Chem.Soc., 1956, 78, 3819.
81. LEE, P. Ph.D. Thesis, 1967.
156
82. HALSH, A., Spectrochim Acta, 1955, 1• 108.
83. GIAMMARISE, A., Atomic Absorption Newsletter, 1966, z, 113.
84. PURUSHOTTAH, A., NAIDU, P.P. and LAL, S.S., Talanta, 1973, 20, 631.
85. BARNES, L., Analyt.Chem. 1966, 38, 1083.
86. GOHEZ COEOO, A. and JIHENEZ SECO, J.L., Rev.Het.(Hadrid), 1968, 4(3), 316(Spanish),
87. THOHERSON, D.R., and PRICE, \v.J., Analyst, 1971, 96, 321.
88. GUNKAGA, J., LEVITNER, C.H. and HAAS, H.G., Clin.Chim.Acta, 1974, 57(1), 77.
89. NALL, H.R., BRUHHEAD, D. and WHITHAH, R., Analyst, 1975, 100(1193), 556.
90. KINSON, K., HODGES, R.J. and BELCHER, C.B., Anal.Chim.Acta, 1963, 29, 134.
91. HURLBUT, J.A. and CHRIS\'IELL, C.D., Anal.Chem., 1971, 43,(3), 465.
92.
93.
HISATOHI, K., t1ATS!Jl'10TO, H. and KUGE, Y., Hizu Shori Cijustsu, 1974, 15(9), 857(Jap).
ELROD, B.B. and EZELL, J .B., At.Abs.Newslett. 1969, 8(2), 4o. '
94. VOGLIOTTI, F.L., At.Abs.Newslett,, 1970, 9(6), 123.
95. SHIHADA, T., TANAKA, K., FUKUI, S., and KANNO, S., Eisei Kagaku, 1973, 19(1), 135(Jap).
96. ROOS, J.T.H., Spectrochim.Acta, 1972, 27B, 473.
97. ROOS, J.T.H. and PRICE, 'II.J., Spectrochim.Acta, 1971, 26B, 441.
98. SCRIBNER, B.F. and HULLIN, H.R., J.Res.natn.Bur.Stand., 1946, 37, 379.
99. STRZYZEHSKA, B., Spectrochim.Acta 1 1971, 26B, 39, 51.
100. OTTAWAY 1 J.M. and PRADHAN, N.K., Talanta, 1973, 20, 927.
157
101. THOMERSON, D.R. and PRICE, IV.J., Analyst, 1971, 96, 321.
102. WALLIS,J.B., Nature, 1965, 207, (4998), 715.
103. MARKS, J.Y. and WELCHER, G.C. Analyt.Chem., 1970, 42, 1033.
104. IIUSLER, J., Atom.Abs.Newsletters, 1971, 10(2), 60•
105. YANAGISA\vA, M., SUZUKI, M. and TAKEUCHI, T., Anal.Chim.Acta, 1970, 52, 386.
106. SASTRI, V.S., CHAKAROBARTI, C.L. and VIILLIS, D.E., Talanta, 1969, 16, 1093.
107. DEAN, J..A. and LADY, .:t.H., Anal.Chem., 1955, 27, 1533.
108. BRYAN, H.A. and DEAN, J.A., Anal.Chem., 1957, 29, 1289.
109. ALLAN, J .E., Spectro.Chim.Acta., 1961, .lZ• 467.
110.PRICE, W.J., Analytical Atomic Absorption Spectrometry, 1972, Hyden & Son Ltd, London, New York, Rheine.
111. COKER, D.T., OTTA\VAY, J.M. and PRADHAN, N.K., Nature Physical Science, 1971, 233 0 69 •
112. KIRKBRIGHT, G.F., PETERS, H.K. and \VEST, T.S., Talanta, 1967, 14, 789.
113. CLOWLEY, T.G., FASSEL, V.A. and KNISELEY, R.N., Spectro.Chim. Acta, 1968, 23B, 771.
114. COKER, D.T. and O'rTAWAY, J.M., Nature 'Phys.Sci., 1971,~,156.
115. MORRIS, L.D., M.Sc. Thesis, L.U.T. 1971.
116. FUKAMACHI, K., MORHIOTO, M. and YANAGA~IA, M., Japan.· Analyst 1972, ~{1), 26 (Japanese)•
117. YANAGISAWA, M., SUZUKI, M. and TAKEUCHI, T., Japan I1ikroChim.Act<! 1973, (3), 478 (Japanese) •
118. KONO, T., Ind.Res.Inst.of Iwate, Tonan-Mura, Shiwa-Gun, Japan Analyst, 1973, 22(1), 55 (Japanese) •
119. FELDMAN, F.J. and PURDY, W.C., Anal.Chim.Acta, 1965,33, 273.
120. FELDMAN, F.J., KNOBLOCK, E.C. and PURDY, W.C., Anal.Chim.Acta, 1967,38,(4), 489.
158
121.BLUNDY, P.D., Analyst, 1958, 83, 555.
122. KUZNETSOV, V.I., Uspekhi Khimii, 1954, 23, 6, 654.
123. KATZ, S.A., McNABB, W.A. and HAZEL, J.F., Anal.Chim.Acta, 1962, 27, 405.
124. KODAMA, K., Hethods for Quantitative Inorganic Analysis, 1963, John Wiley and Son Ltd.
125, VOGEL, A.I. A Text Book of Quantitative Inorganic Analysis, 1961, Third edition, Longmans
126. HEUNIER, P., Compt.Rend., 1934, 199, 1250.
127. ASHTON, A., Ph.D. Thesis, 1973, L.U.T.
128. BIANCIFIORI, C., BORDONALI, C. and BESAZZA, G., Chim.Ind. (Milan), 1968, 50, 423.
129. BELLING, P.J., Effluent Hater Treat, J., 1969, _2, 314.
130. Analytical Hethods for Atomic Absorption Spectrophotometry Perkin-Elmer Corp. Norwalk,Conn., 1968, pp. 1-24, Cr. 1-Cr 6 •
131. WEINHARDT, A.E. and HIXON, A.N., Ind.Eng.,Chem., 1951, 43, 1676 •
132. KITSON, R.E. and MELLON, H.G., Ind.Eng.Chem.Anal, 1944, 16, 379. (C.A. 38 No. 3926). --
133. SIHENSON, D.G. WERTHAN, M., lo/ESTOVER, L.H. and HEHL, J.W.1
J.Biol.Chem., 1946, 166, 747.
134. ROBINSON, R., ROUG!!AN, !1. and WAGSTAF'F, D.F., Ann.Clin.Biochem., 1971' §_(5)' 168. .
135. FERLAN, Z., Z.Anal.Chem., 1972, 260(4), 291•
136. DALY, J.A. and ERTINGSHAUSEN, G., Clin.Chem., 1972, 18(3), 263.
137, A!1ADOR, E. and URBAN, J., Clin.Chem., 1972, 18(7), 601.
138. FISKE, C.1l., and SABBAROW, Y., J.Biol.Chem., 1925, 66, 375.
139. GROS, I.AND KOLCK, H., Chim.Anal., 1972, £(2) 1 98.
140. HOPPE-SEYLER, G.F. and GUNDLACH, G., Z.Klin.Chem.Klin.Biochem., 1968, §_(4), 256.
159
141. AMIC, J., LAIRON, D. and HAUTON, J., Clin.Chim.Acta, 1972, 40(1), 107.
142. DRE\ffiS, P.A., Clin.Chim.Acta, 1972, 39(1), 81•
143. HIRSCH, C. , KUPFER, S. and ORESKES, I., Clin.Chim.Acta, 1967, 18(2), 322.
144. YEE, H.Y. and BLACK\vELL, L., Clin.Chem., 1968, .J.!!:(9), 898.
145. NEGRIN, A., Clin.Chim.Acta, 1964, 10(3), 262.
146. GUIRGIS, F.K. and HABIB, Y.A., Clin.Chem., 1971, .:!2(2), 78.
147. PAREKH, A.C. and JUNG, D.H., Clin.Chim.Acta, 1970, 27(3), 373.
148. BAGINSKI, E.S. ar.d ZAK, B., ibid, 1960, _2(6), 831f.
149. BAGINSKI, E.S., HARIE, S.S. and ZAK, B., Hicrochem.J., 1974, 12.(3), 285.
150. JAENICKE, L., Analyt.Biochem., 1974, £1(2), 623.
151. NATII, R.L. and DEBNATH, B., J.Inst.Chem(India), 1968, 40(2), 73.
152. HUXTABLE, R. and BRESSLER, R., Analyt.Biochem., 1973, 54(2), 604 •
153. HORIN, L.G. and PROX, J., Clin.Chim.Acta., 1973, 46(2), 113.
154. GOODWIN, J.F., Clin.Chem., 1970, 2£(9), 776.
155. ROJKIN, H.L. and ZACCARA, F.A., Revta Asoc.Bioquim., 1974, 39, 215.
156. AKAHATSU, S. and IHAI, K., J.Biochem. 1952, 39, 203.
157. DUCRET, L. and DROUILLAS, H., Anal.Chim.Acta, 1959, ~. 86.
158. BABKO, A.K., SHKARAVSKII, YU.F~, and KULIK, V.I., Z.Anal.Khim., 1966, ~(2)' 196.
159. TRAUTNER, A., Hetalloberflache, 1971, 25(7), 234.
160. idem., Hetallwissenschaft und Technik, 1972, 26(5), 467.
161. SOYEt~OFF, B., J.Biol.Chem., 1947, 168, 447.
160
162. ITAYA, K. and UI, M., Clin.Ghim.Acta, 1966, 21(3), 361.
163. MARTIN, J.B. and DOTY, D.H., Anal.Chem., 1949, ?.1.• 965.
164. VAN BELLE., H. Analyt.Biochem., 1970, 33(1), 132.
165. BASTIAANSE, A.J., and HEIJERS, G.A.M., Z.Klin.Chem.Klin.Biochem. 1968, _£(1)' 48 •.
V V .I ' 166. STEPANOVA, I., Clin.Chim.Acta., 1967, ~(2), 330 (ger) •
167. HOHENWALLNER, W. and WIHMER, E., Glin.Chim.Acta, 1973, 45(2), 169.
168. ALTMANN, H.J., FURSTENAU, E., GIELE\vSKI, A. and SCHOLZ,L., Z.Anal.Chem., 1971, 256, 274 (ger)·
169. STEVENS, J .F. and YEONANS, F .R.i\., Medical Laboratory Technology, 1975, 32, 19.
170. KALLNER, A., Glin.Ghim.Acta, 1975, 59(1), 35.
171. ANNER, B. and MOOSMAYER, M., Anal.Biochem., 1975, 65(1), 305.
172. CATLOVE, E., HARRIS. E.K., and WILLIAM. G.Z., Clin.Ghem., 1970, 16{12)' 1028.
173. FRANZKE, C., HOLLSTEIN, E., BRANDT1 P., and ZIETZE, H., Nahrung, 1973, !L(6), 679 (ger)•
174. W!LLIAMS, F.R. and WfiTTEHEAD. J., J.Appl.Ghem,, 1952,~, 213.
175. BABKO, A.K. and KARNAUKHOVA, N.N., Zhur.Anal.Khim., 1967, ~(6) ,868.
176. PICASSO, G. and PIZZIMENTI, ·A., Met.Ital., 1963, 55, 361.
177. LUKE, C.L., Analyt.Chim.Acta, 1967, 37, 97.
178. LUKE, G.L., Analyt.Chem., 1956, 28, 1276.
179. BENNETT, R.L., and SMITH. H.A., Anal.Chem., 1959, 31(11), 1441
180. LUKE, C.L., Anal.Chim.Acta, 1967, 39(3), 4o4.
181. GLULEY, H.J., Analyst, 1951, 76, 523.
182. PATHAN, A.S., Ph.D. Thesis, L.U.T., 1974.
183. GORENC, D., KOSTA, L. and BYRNE, A.R., Vestn.Slov.Kem.Drus., 1974, 21(1-2), 9.
184. DAGNAL, R.M., \•/EST, T.S., and YOUNG, P., Analyst, 1967, 92, 27.
185. ASHTON, A., FOGG, A.G., and THORBURN BURNS, D., Analyst, 1973, 98, 202.
186. PLOTNIKOVA, O.M., SELEZllEVA, N.A., and REDKINA, A.I., IzV. Akad.Nauk.Kaz.SSSR. Ser.Khim., 1975, 25(1), 74(Rus).
187. YAMAZAKI, M., MORI, T., and ENOKI, T., Bunseki Kagaku, 1973, 22(1), 112, (Jap.).
188. LAGOWSKI, J.J., Chemistry of Non Aqueous Solvents, 1967, _?;, Academic Press, New York, London.
189.· BUNCEL, E., and SYMONS, E.A., Chemical Communication, 1970, 164.
190. BUNCEL, E., KESHARKY, S. and SYJ10NS, E.A., ibid., 1971, 120.
191. WILD, M.J., B.Sc. Project report, L.U.T., 1971.
192. CROAL, G., G.K.N. Method, Private communication.
193. KATZ, S.A., !1cNABB, VI.M., and HAZEL, J .F., Anal.Chim.Acta, 1961' 25, 193.
194. Al-SIBAAI, A.A., Ph.D. Thesis, L.U.T., 1975.
195. BUCK, R.P., Anal.Chem., 1972 1 44, 270R.
196. BUCK, R.P., Anal.Chem., 1974, 46, 28R.
197. MOODY, G.J., and THOMAS, J.D.R., Talanta, 1972, 12.1 623~
198. KORYTA, J., Anal.Chim.Acta, 1972, 2!, 329.
199. COVINGTON, A.K., Critical Reviews in Analytical Chemistry, 1974, 3, 355.
200. DRUST, R.A. 1 "Ion-Selective Electrodes", N.B,S. Publication, 1969, 314.
201, MOODY, G.J., and THOMAS, J.D.R., "Selective Ion-Sensitive Electrodes", 1971, Merrow Public Co, England.
202. FOGG, A.G. 1 PATHAN, A.S, and THORBURN BURNS, D. 1 Anal.Chim.Acta, 1974,69, 238.
162
203. FOGG, A.G., KUMAR, J.L. and TI!ORBURN BURNS, D., Anal.Lett., 1974, z, 629 •
. 204. FOGG, A.G., PATHAN, A.S. i and THORBURN BURNS, D., Anal.l,ett., 1974, z, 539·
205. FOGG, A.G., and AL-SIBAAI, A.A., in press.
206. FOGG, A.G., and AL-SIBAAI, A.A., in press.
207. HALAPRADE, L., Compt.rend., 1928, 186, 382; Bull.Soc.Chim., France, 1928, 43(4), 683.
208. JOHAR, G.S., Z.Anal.Chem., 1974, 269(2), 123.
209. LAURIE, S.H., IVILLIAHS, J.H. and NYMAN, C.J., J.Phys.Chem., 1964, 68(6), 1311.
210. BUIST, G.J. and LE\VIS, J.D., Chemical Communications, 1965, 4, 66.
211. LA11BERT, 11., and NEISH, A.C., Can.J.Res., 1950, 28, 83.
212. HABER, F. and KLEHENSIE','/ICZ, Z •. , Z.Phys.Chem., 1909, 67, 385.
