BCHET-141

ANALYTICAL METHODS IN
Indira Gandhi National
Open University CHEMISTRY
School of Sciences

Block

2
CHROMATOGRAPHY AND ION EXCHANGE

UNIT 5
General Aspects of Chromatography 79

UNIT 6
Adsorption Chromatography: 93

UNIT 7
Ion Exchange Chromatography 103
Course Design Committee

Prof. N.K. Kaushik, (Retd.) School of Sciences,

Department of Chemistry, IGNOU, New Delhi
University of Delhi, New Delhi
Prof. M.S. Nathawat
Prof. B. S. Saraswat (Retd.)
School of Sciences, IGNOU, Prof. Sunita Malhotra
New Delhi Prof. B.I. Fozdar
Prof. Javed A. Farooqi
Prof. Nafisur Rehman, Prof. Sanjiv Kumar
Department of Chemistry, Prof. Lalita S. Kumar
A.M.U. Aligarh Prof. Kamalika Banerjee

Block Preparation Team

Prof. Sunita Malhotra Prof. N.K. Kaushik, (Retd.), Editor
School of Sciences, IGNOU Department of Chemistry,
New Delhi University of Delhi, New Delhi

Course Coordinator: Prof. J. A. Farooqi

Print Production
Mr. Rajiv Girdhar Mr. Hemant Kumar Parida
Assistant Registrar (Pub), Section Officer (Pub),
MPDD, IGNOU, New Delhi MPDD, IGNOU, New Delhi

Acknowledgements: Sh. Sarabjeet Singh for CRC preparation

Material partially adapted from the MCH-001 and MCH-002 Course of PGDAC
Programme.

March, 2022
@ Indira Gandhi National Open University, 2022

ISBN:

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BLOCK 2: CHROMATOGRAPHY AND ION EXCHANGE
In this Block, we will discuss the general aspects of chromatography and ion exchange
methods. Unit 5 will begin with a discussion on classification of chromatographic methods. In
this unit, we will also explain basic principle of paper chromatography and its applications.

In Unit 6, the principle and efficiency of the column chromatography technique will be
described. This will be followed by a brief account of the mechanism of separation operating
in this technique. The important features of adsorption and partition chromatographic
techniques will then be compared.

Unit 7 on ‘Ion Exchange Chromatography’ covers the classification of ion exchangers. The
details of mechanism of exchangers ion exchangers and applications of ion exchange
chromatography have also been explained.

Expected Learning Outcomes

After studying this block, you should be able to:

 discuss the classification of various chromatographic techniques according to the

different criteria;

 state the importance of paper chromatography, column chromatography and ion

exchange chromatography;

 explain the principle of paper chromatography, adsorption chromatography and ion

exchange chromatography, and

 discuss the applications of paper chromatography, adsorption chromatography and ion

exchange chromatography.
Unit 5 General Aspects of Chromatography

UNIT 5

GENERAL ASPECTS OF
CHROMATOGRAPHY

Structure
5.1 Introduction Efficiency of the Technique
Expected Learning Outcomes Mechanism of Separation
5.2 Classification of Development of Chromatograms
Chromatographic Methods
5.4 Summary
5.3 Partition Chromatography:
5.5 Terminal Questions
Paper Chromatography
5.6 Answers
Principle

5.1 INTRODUCTION
This is the first unit of this Block on ‘Chromatography and Ion exchange’. Here,
we will cover the general aspects of chromatography. The unit will begin with a
discussion on classification of chromatographic methods. This will be really an
eye opener for you to realise that what a vast variation of this technique is
possible .A wide variety of options has resulted in this technique with time as
newer developments kept on taking place and increasing need of separation
of various types of substances was felt.

We will also explain the technique of paper chromatography which is classified

as partition chromatography because the mechanism of the separation
involved in this technique is partition. Under this technique, we will focus on
the principle of separation as well as the efficiency of this technique. The
mechanism involved will also be discussed and the development of the
chromatograms will be explained. We will also discuss the characteristics of
the mobile phase and the concept of Rf value.

Expected Learning Outcomes

After studying this unit you should be able to:

 discuss the classification of various chromatographic techniques

according to the different criteria; 79
Block 2 Chromatography and Ion Exchange

 state the importance of paper chromatography and explain the various

terms involved in it;

 explain the principle of paper chromatography;

 describe the mechanism of separation in partition chromatography;

 explain how the chromatogram is developed in paper chromatography;

and

 discuss the applications of paper chromatography.

5.2 CLASSIFICATION OF
CHROMATOGRAPHIC METHODS
‘Chromatography’ was introduced by a Russian Botanist, Mikhail Tswett in
1906. It is now a journey of more than a century and a lot of interesting
developments have been introduced in using such techniques .A lot of
variations with regard to the use of equipment and materials used have taken
place from time to time.

Chromatography was used by Tswett to separate the plant pigments using a

simple glass column packed with finely divided calcium carbonate. He used
petroleum ether to separate various pigments which appeared as bands at
different heights in the column after separation. You will also study about
Mikhail Tswett column chromatography in Unit 6.
(1872-1919)
Thus, the technique of chromatography involves the separation of the
components of a mixture by their distribution between two phases- one being
the stationary phase of large surface area and the other being the mobile
The technique of phase. Here, the mobile phase moves on the stationary phase in a definite
chromatography is a
direction. Depending upon the type of stationary support, nature of mobile
physical method of
separation. It is based
phase and the mechanism involved in the separation; several types of
on molecular weight of chromatographic methods are possible, see Fig.5.1.
a substance and its
Let us see the following possible variations when each of the above mentioned
adsorption or partition
coefficient. Thus, aspects are taken into account:
substances with
(i) Depending upon the type of stationary support or more accurately the
lesser molecular
masses diffuse more
shape of the stationary support, chromatographic technique can be two-
quickly than those with dimensional or three-dimensional.
higher molecular
The techniques of paper chromatography and thin layer chromatography
masses.
are two further subdivisions of two-dimensional chromatography because
the stationary phases used in these two types of chromatography are,
respectively paper and a solid support coated on a plate of glass, plastic or
metal. The shapes of all these supports are two-dimensional; hence, these
two techniques are classified under two-dimensional chromatography.

The technique of column chromatography is considered under the three-

dimensional chromatography. Column chromatography involves the
stationary support present in a column which is three-dimensional in
80 shape.
Unit 5 General Aspects of Chromatography

(ii) According to the nature of mobile phase-whether the mobile phase is

gas, liquid or supercritical fluid, the respective chromatographic
techniques are called gas chromatography, liquid chromatography or
supercritical fluid chromatography.

(iii) We can also classify chromatographic techniques into different classes

according to the mechanism involved in separation. Accordingly, we
can name these techniques as partition, adsorption, ion exchange or
size exclusion chromatography.

Classification of Chromatography

On the basis of physical

On the basis of shape of state of mobile phase On the basis of interaction of
stationary support solute to the stationary phase

Liquid Super Critical Fluid

Chromatography Chromatography
Two-dimensional Three-dimensional Adsorption
Ion Exchange
Chromatography
Chromatography
Gas
Chromatography
Partition Size Exclusion
Column Chromatography Chromatography
Chromatography

Paper Thin Layer

Chromatography Chromatography

Fig. 5.1: Classification of Chromatography

We will be dealing with partition chromatography i.e. paper chromatography in

this unit itself. The adsorption chromatography using column chromatography
will be dealt in Unit 6 while ion exchange chromatography will be described in
Unit 7. The size exclusion chromatography, however, is not a part of this
course and will not be dealt here.

Thus, all the available chromatographic techniques cannot be classified using

a single criterion, as the different combinations of stationary and mobile
phases can be used and the mechanism operating in separations may also
differ. Hence, it is the nature of mobile phase which is taken as main criterion
to classify the technique. The other factors mentioned above are discussed as
details or sub-criteria under this main class.

Thus, when the nature of mobile phase is taken as the main criterion, then the
gas, liquid and supercritical fluid chromatographies are considered. The
gas and supercritical fluid chromatography are beyond the scope of this
course. Therefore, we will keep our discussion focused to the liquid
chromatography only.

When the mobile phase is liquid, the stationary phase can be either a liquid
supported on a solid or a solid and this gives rise to the liquid-liquid
chromatography or liquid-solid chromatography. In liquid-liquid
chromatography, the distribution of components of mixture takes place by their 81
Block 2 Chromatography and Ion Exchange

partitioning of the components of the mixture in the two liquid phases, as was
the case in the solvent extraction.

Further, two-dimensional and three-dimensional chromatographies are

possible as shown in Fig. 5.2, according to the shape of the stationary phase
taken. Each of these variations may further involve upward or downward
direction of movement of the mobile phase, thereby widening the options to
ascending and descending modes of operation. This is illustrated below in
Fig. 5.2.

Liquid Chromatography
Mobile phase: Liquid

Liquid-liquid Liquid-solid
chromatography chromatography
(stationary phase: (stationary phase:
liquid) solid)

Two-dimensional
chromatography Three-dimensional chromatography
e.g. column chromatography

Paper chromatography Thin layer chromatography

The other variations available are

. Ascending ,descending and horizontal modes
. Normal phase and reverse phase chromatographies
. High Performance Liquid Chromatography (HPLC)

Fig. 5.2: Types of liquid chromatography

Normally, polar support is taken as the stationary phase and non-polar mobile
phase is chosen. Such a combination is referred to as normal-phase
chromatography. But, if the polarities of the two phases are reversed, then it is
called reverse-phase chromatography.

Various forms of chromatographies explained above can also be performed

under high pressure leading to high performance or high pressure liquid
chromatography (HPLC).

Let us now focus our attention on paper chromatography which involves

partition mechanism.

SAQ 1
Name two types of two-dimensional chromatography.
82
Unit 5 General Aspects of Chromatography

5.3 PARTITION CHROMATOGRAPHY: PAPER

CHROMATOGRAPHY
The foundation of the paper chromatography dates back to 1944 when
R. Consden, A.H. Gordon and A.P.J Martin reported the separation of
amino acids using paper. The amino acids present in only 200 g of wool
could have been separated using paper chromatography. The importance of
chromatography was recognised when Nobel prize was awarded in 1952.

The technique of the paper chromatography is very simple and useful. Let us
now understand its principle.

5.3.1 Principle Archer John Porter

Martin
Paper chromatography is a kind of liquid-liquid partition chromatography in
which the substances are distributed between two liquids-the stationary (1st March 1910-28th
phase(usually water that is held in the fibres of the paper, also called July 2002)
water –cellulose complex containing 20% water) and the moving liquid or the
He shared Nobel
mobile phase ,also called the developing solvent. Prize in Chemistry for
year 1952 with the
The paper chromatography involves the use of paper which is made up of
Richard Synge for the
highly pure cellulose .The examples of such a paper are - Whatman papers invention of partition
with no. 1,2,3,3 MM, 31 ET etc. These papers are having uniform physical chromatography.
characteristics and are very low in organic and inorganic impurities. Some
more types of modified papers are also used about which you will study later.

Here, it would be interesting to know that paper chromatography can be done

in ascending mode, descending mode or in circular fashion. The simplest one
is ascending mode paper chromatography for which the set up is shown below
in Fig. 5.3.
Lid

Richard Laurence
Millington Synge
Paper
th th
(28 Oct 1914-18
August 1994)
Solvent
He shared Nobel
Mixture or Prize in chemistry for
impure compound (A)
the year 1952 with
A.J.P. Martin for the
Fig. 5.3: Set up for the ascending mode paper chromatography invention of partition
chromatography.
The sample containing the mixture of compounds to be separated is taken in a
suitable solvent and using a capillary, a fine small spot is put on the line drawn
on the paper. Let this spot be marked as A. The sample could also be an
impure compound which could be tested by paper chromatography for how
many impurities are present. 83
Block 2 Chromatography and Ion Exchange

The paper is then kept in the development chamber as shown in Fig. [Link]
solvent which is the mobile phase is then allowed to rise by the capillary action
over the spot of mixture of compounds . After some time, the solvent takes
along with it, the different components of the mixture (of spot A) to different
heights on the paper leading to their separation.

Similarly, we can have descending paper chromatography and circular paper

chromatography .The experimental set ups for these two methods are shown
in Fig. 5.4. a) and b) below.

Glass tank
Trough for solvent

Spot of sample
Start line

Paper suspended in trough

a)

Inverted petridish
Sample spot
Circular Whatman paper
Cotton wick
Petridish Solvent

b)

c)
Fig. 5.4: a) Descending paper chromatography

b) Circular paper chromatography

c) Separated compounds by circular chromatography

In descending paper chromatography, the solvent tank contains the solvent

which moves from upside to downward direction on the paper. The spots of
the sample are put on the paper and the same is placed in the development
chamber as shown in Fig. 5.4 a).
84
Unit 5 General Aspects of Chromatography

In circular paper chromatography, as shown in Fig. 5.4 b), the spots of the
sample are placed on the paper in circular fashion. Then, the moving phase or
the mobile phase which is a liquid (i.e. a solvent or a mixture of solvent) is
taken in a Petri dish. At the centre of the paper, a very small hole is made
using a pin. A wick is then placed in this hole whose other end dips in the
mobile phase.

The solvent rises to the centre of the paper through the cotton wick .The
mobile phase then carries along it, different components of the sample in a
circular fashion. Thus the different separated components are obtained as
rings on the Whatman paper and not as the spots.

Having understood the basic principle of paper chromatography, let us now

study about its efficiency.

5.3.2 Efficiency of the Technique

As discussed above, the techniue of paper chromatography is not only simple
and easy to perform but it is also efficient in separating the components of a
mixure or in obtaining the pure compound from an impure sample.

For obtaining the pure components of a mixture or for getting a pure

compound from its impure sample, quantitative paper chromatography has to
be performed. A 30 cm x 30 cm sheet of paper is taken and several spots of
the sample are put in a line near the base of the paper. The paper is then kept
in the development chamber as shown in Fig. 5.5. The mobile phase is then
allowed to rise by the capillary action over the spots of sample.

Fig.5.5: Quantitative paper chromatography

The separated components appear as separate bands at different heights on

the paper. The relevant portions of the paper are cut for obtaining different
components in pure form. Each of the component is then separataly extracted
from the paper by dissolving it in a suitable solvent. The removal of the mobile
phase or the solvent yields the pure components.

The technique of paper chromatography has been efficiently used for the
qualtative analysis of several types of naturally occurring and synthetically
obtained reaction mixtues. The separation can be effectively carried out and 85
Block 2 Chromatography and Ion Exchange

the identification of different components can be done by putting the spot(s) of

the known standard sample(s) alongwith the sample to be tested. The spots of
the same compound (whether present in a mixture and as a pure sample) will
appear at the same level on the paper after the separation has occurred.
Hence,the componenets of the sample can be correlated with the standard
compounds.

Paper chromatography is a very efficient technique and it can be used to

separate very closely related compounds such as isomers, homologues and
species with different valency or oxidation states etc.

Some examples of separations using paper chromatography are as follows:

(i) Identification of metal ions such as Ni2+, Co3+, Zn2+ and Mn2+

(ii) Separation of mixture of pesticides containing heptachlor, BHC, aldrin

etc. into its pure components

(iii) Checking of foods and drinks for adulterants using the standard
samples of pure components and adulterents.

After understanding the efficiency of paper chromatography ,let us now study

the mechanism by which such separations take place.

5.3.3 Mechanism of Separation

In the beginign of this section, we have mentioned that cellulose paper is used
as solid support in paper chromatography and the mobile phase is a suitable
solvent which passes over the sample carrying along with it, the different
components of the sample to different extents. But how does this happen?

Which mechanism is operating in such a separation?Let us find out.

So far we have not deliberated upon the water present in the cellulose
[Link], this has the actual role in deciding the mechanism of separation. In
fact, it is this water which is acting as the stationary phase. Here, the mobile
phase is also a liquid. Thus, the components of the mixture will be separated
between two liquid phaes and hence,paper chromatography is a liquid-liquid
chromatography .

Therefore, the mechanism involved here is the partition of the componens in

two different liquid phases. Every time the mobile phase moves, there is an
equilibrium between the two liquid phases and the components partition
themselves in the two phases.

The components which are held strongly by the stationary phase will be slow
in moving alongwith the mobile phase while those which are weakly held by
the stationary phase will be moving faster along with the mobile phase.

Thus,different components of the mixture will move with different rates along
with the mobile phase. This eventually leads to their separation in due course
of time.

However, this simple mechanism of partitioning is not taken as the only mode
of separation in paper chromatography. It is now believed that hydrogen
86
Unit 5 General Aspects of Chromatography

bonding, interactions between solutes and cellulose support also play an

important role in separation.

It is also possible to use modified cellulose papers in paper

chromatography. The chemically modified papers such as carboxylated and
acetylated papers are available. The carboxylated papers have increased
carboxyl content and are suitable for the separation of amines and amino
acids. The aceylated paper is more hdyrophilic and can be used for reverse
phase chromatography.

The paper to be used in paper chromatography can also be loaded with cation
or anion exchange resins. In such a case, the mechanism of separation would
be via excange of ions.

It is also possible to use papers impregnated with silica or alumina, in which

case, the mechanism of separation would involve adsorption of the
components of the mixture on paper to different extents leading to their
separation.

Hydrophilic papers can be obtained by treatment with methanol, glycerol,

glycol etc.

Having understood the mechanism of separation,try to answer the following

SAQs.

SAQ 2
What are modified cellulose papers?Give examples.

SAQ 3
What is the general mechanism of separation in paper chromatography? How
does it get modified in case of papers impregnated with silica gel?

5.3.4 Development of Chromatograms

We have discussed above about the ascending, descending and circular
forms of paper chromatography. Whatever form we choose from these, before
starting the separation using paper chromatography ,the general procedure
involves the saturation of the development tank with the vapours of the solvent
used. The development tank containing the mobile phase is covered with the
lid/cover and kept undisturbed for sometime to allow the vapours of the mobile
phase to saturate the tank .

The substance to be analysed is dissolved in a suitable solvent and its spot of

is put on the paper. Let the position of this spot be represented by the line at
point A. The paper is then cap in the development chamber and it is then
allowed to come in contact with the mobile phase. The system is again left
undisturbed for sometime.

The components present in the substance are then carried along by the
mobile phase. Their rates of movement will be different as their inteactions
with the stationary and mobile phases will be different. 87
Block 2 Chromatography and Ion Exchange

Here, it is also important to note that the nature of solvent used as the mobile
phase has an important role in the development of paper chromatogram.

The solvent or the mobile phase should be pure, free from impurities and dry.
The following other criteria should also be followed while chosing the mobile
phase.

 The solvent system should not react chemically with the components of
the substance to be analysed.

 The composition of the solvent system should not change during the
course of separation. So, as far as voltility is concerned, we should prefer
non-volatile solvents.

 The solvent system or the mobile phase should be so selected that it is

able to separate different components present in the sample. The polarity
of the solvent system can be changed for getting the good separation.

As in the case of paper chromatography, here also the water present in the
cellulose acts as the stationary [Link], here in paper chromatography ,
the starionary phase is polar in [Link] can use a relatively less
polar solvent such as ethanol, acetone, formamide, amines or a suitable
mixture of such solvents as the mobile phase.

Some typical examples of such mobile phases are as follows:

 Isopropanol-Ammonia-Water (9:1:2 v/v)

 n-Butanol-Acetic acid-Water (4:1:5 v/v)

 Water-Phenol

 Formamide-Benzene

For the separation of cations, some of the commonly used mobile phases are
as follows:

 Methanol

 Methyl ethyl ketone containing 30% (v/v) water and 1% (w/v) potassium
thiocyanate

 Pyridine containing 10% (v/v) water

 Acetone containing 5% (v/v) water and 8% (v/v) hydro

 chloric acid.

The components of the substance get separated after leaving the paper in the
development chamber for some time. The paper is then taken out from the
development chamber and a line is marked on it with a pencil at the level to
which the mobile phase has travelled. Let this line be represented by B. The
solvent is then allowed to dry. Remember that the position of the original spot
was represented as point A and a line can be drawn at this level, see Fig. 5.7.

If the separated components of the mixture are coloured, their spots will be
clearly visible on the paper, see Fig. 5.6. Otherwise, a suitable locating agent
is sprayed on the paper to detect the components.,
88
Unit 5 General Aspects of Chromatography

Fig. 5.6: Chromatogram

The examples of some locating agents are as follows:

 Dimethyl glyoxime (for Ni)

 Dithizone

 Potassium chromate

 Ammonium sulphide

 Iodine vapours (for organic compounds)

 Starch solution (for carbohydrates)

 Ninhydrin solution (for amino acids)

Sometimes, the spots of the components are visible under UV light.

Once the spots of the separated components are located, these then need to
be characterised. One such way is to calculate their Rf values and match
these values them with the Rf values of their standard or pure samples which
are usually run simultaneously on the same paper under identical conditions.

For this purpose, let us first understand what is an Rf value.

The Rf value

It is the retardation factor and is defined as the following ratio:

Distance travelled by a solute

Rf  … (5.1)
Distance travelled by the solvent

If we now look at the chromatogram shown in Fig. 5.6, then, we can calculate
the Rf values for the separated components.

Let us understand this the calculation of the Rf values with the help of a
hypothetical chromatogram shown in Fig. 5.7. Let there be two components 1
and 2 which have been separated by using the technique of paper
chromatography (or TLC). Let the distance travelled by component 1 is AY
and that travelled by component 2 is AX.
89
Block 2 Chromatography and Ion Exchange
55

4
4
B
33

22 X 2

11 Y 1

00 A
Sample spot of
a mixture of
components 1 and 2

Fig. 5.7: Calculation of Rf value from chromatogram

Therefore, their Rf values can be calculated as given in the following

equations:

Distance travelled by component1 AY

For component 1, Rf   … (5.2 a)
Distance travelled by mobile phase AB

Distance travelled by component 2 AX

For component 2, Rf   … (5.2 b)
Distance travelled by mobile phase AB

It must be taken care that the distances AY and AX are measured between the
line A and the centre points of spots Y and X, respectively.

Many times, the sample of the available standard substances are placed along
with the mixture (or unknown substance). Then the chromatogram is obtained.
The separated components of the mixture are then matched with the spots of
the standard samples and the components are characterised, see Fig. 5.8.

Solvent front

Pure A
Pure B

Mixture of A and B

Spot of standard A Spot of standard B

Fig. 5.8: Development of Chromatogram along with the use of standard samples

The Rf values are characteristic of the solute under a given set of conditions.
They may change with the change in the solvent system, temperature,
presence of impurities and sometimes with the development time. Hence,
these need to be carefully compared.

Hence, we can say that paper chromatography can be used for the qualitative
analysis of different samples and identification of components present in a
mixture.

Now you can answer the following questions.

90
Unit 5 General Aspects of Chromatography

SAQ 4
Which component shown in Fig. 5.7 has smaller Rf value?

SAQ 5
Give any two characteristics of a mobile phase.

Let us now sum up, what we have learnt in this Unit.

5.4 SUMMARY
In this unit, we have learnt the following main aspects of chromatography:

 The chromatographic techniques can be classified in various ways

according to the shape of the stationary support, nature of the mobile phase
and the mechanism of separation.

 A variety of chromatographic techniques have evolved with variations in

their operational aspects over the time.

 Paper chromatography is a very simple and useful chromatographic

technique.

 Various modes of paper chromatographic techniques are available-

ascending, descending and circular.

 The mechanism of separation using paper chromatography is mainly

partition. However, other variations, i.e. adsorption and ion exchange using
the modified papers, are also possible.

 The development of chromatograms is an important skill and several

aspects related to placement of spots of samples, use of suitable mobile
phases and proper handling of chromatogram are to be taken care of to
achieve the good separation of the components. The spots of the sample
and the reference substances should not dip in the solvent.

 The Rf values are characteristic of the substances but several factors may
affect these values.

5.5 TERMINAL QUESTIONS

1 Name different types of mechanisms operating in the chromatographic
techniques.

2. What are the polarities are two phases in reverse-phase chromatography?

3. Differentiate between ascending and descending modes of paper

chromatography.
91
Unit 6 Adsorption Chromatography

UNIT 6
ADSORPTION CHROMATOGRAPHY

Structure
6.1 Introduction 6.7 Development of
Chromatograms: Frontal
Expected Learning Outcomes
Analyses, Elution and
6.2 Classification Displacement Methods
6.3 Principle 6.8 Summary
6.4 Efficiency of the Technique 6.9 Terminal Questions
6.5 Mechanism of Separation 6.10 Answers
6.6 Adsorption and Partition
Chromatographies

6.1 INTRODUCTION
In Unit 5, you have studied about the classification of chromatographic
methods and about partition chromatography, i.e. paper chromatography in
detail.

In continuation to that, in this unit, you will study about one more type of
chromatography, i.e. column chromatography in which the separation of the
components of a mixture takes place by the adsorption mechanism.

We will begin the discussion of this unit by recapitulation of the classification of

chromatographic methods dealt in Unit 5. Here , we will describe the principle
and efficiency of the column chromatography technique. This will be followed
by a brief account of the mechanism of separation operating in this technique.
The important features of adsorption and partition chromatographic techniques
will then be compared.

Finally, we will explain the development of chromatograms by frontal analyses,

elution and displacement methods.

Expected Learning Outcomes

After studying this unit you should be able to:

 explain the classification of chromatographic techniques;

93
Block 2 Chromatography and Ion Exchange

4. Give some examples of areas of application of paper chromatography.

5. List the factors which can affect the Rf value.

5.6 ANSWERS
Self Assessment Questions

1. Paper chromatography and thin layer chromatography.

2. Papers can be modified chemically to yield carboxylated and acetylated

papers.

3. It is partition mechanism in paper chromatography when different

components partition themselves between the two phases. In case of
papers impregnated with silica gel, the mechanism involves adsorption of
the components on the stationary phase to different extents.

4. Component 1.

5. It should be pure and should not react with the substance to be

separated.

Terminal Questions
1. Partition, adsorption, ion exchange and size exclusion.

2. In reverse-phase chromatography, non-polar support is taken as the

stationary phase and a polar mobile phase is taken.

3. Refer to Sec. 5.3.1.

4. Paper chromatography has applications in the following areas:

 In the identifiction of metal ions

 In the analysis of mixtures of organic compounds

 To assess the purity of a given sample-organic and inorganic –both

types

 In the analysis of foods, drinks, pharmaceuticals, pollutants,

fertilizers, drugs etc.

5. (i) Temperature change, (ii) change of solvent and (iii) the presence of
impurities can change the Rf value.

92
 Block 2 Chromatography and Ion Exchange

 discuss the principle of column chromatography which is a form of liquid-

solid chromatography;

 describe the efficiency of the column chromatography technique;

 explain the mechanism of adsorption involved in the separation of

components of a mixture using column chromatography;

 compare the important aspect of adsorption and partition

chromatographic techniques; and

 describe different methods of development of chromatograms.

6.2 CLASSIFICATION
You may remember from Unit 5, Sec. 5.2 that according to the shape of the
stationary support, chromatography can be classified as two-dimensional or
three-dimensional chromatography. A portion of Fig 5.1 is shown below in
Fig.6.1.

Both thin layer chromatography and column chromatography are the liquid-
solid chromatography techniques as these involve the liquid mobile phase
and a solid stationary phase.

Classification of Chromatography

On the basis of physical

On the basis of shape of state of mobile phase On the basis of interaction of
stationary support solute to the stationary phase

Two-dimensional Three-dimensional

Column
Chromatography

Paper Thin Layer

Chromatography Chromatography

Liquid-solid chromatography

Two-dimensional chromatography Three-dimensional chromatography

Thin layer chromatography Column chromatography
(using solid suport on a plate
made of plastic/glass/metal)

Fig. 6.1: Types of Liquid-solid Chromatography

The technique of thin layer chromatography is classified as the two-

dimensional chromatographic technique but its discussion is beyond the scope
of this course. Hence, in this unit, we will be dealing mainly with the column
94 chromatography which is a three-dimensional chromatographic technique.
Unit 6 Adsorption Chromatography

6.3 PRINCIPLE
The column chromatography involves the use of liquid mobile phase and a
solid stationary phase; hence, it is a type of liquid-solid chromatography as
mentioned above.

A solid stationary phase is used in liquid-solid chromatography. Several

stationary supports such as silica gel, activated alumina, magnesium
carbonate, hydrated calcium silicate, talc, fuller’s earth, sucrose and powdered
cellulose can be used in liquid-solid chromatography. Thus, the availability of a
large number of materials offer a wide range of flexibility in choosing the
desired stationary phase in terms of the surface area, particle size and type of
adsorbent.

As shown in Fig. 6.2 a column which is a long tube which is filled with the solid
support. The mixture to be separated is first dissolved in a suitable solvent and
its slurry is prepared which is then placed at the top of column. Different
components of the mixture get adsorbed on the stationary phase to different
extents.

When the mobile phase is allowed to pass over stationary phase, the
component of the mixture which is least adsorbed on the stationary phase gets
carried along the mobile phase at a faster rate than those components which
are strongly adsorbed.

Fig. 6.2: Set up for the Column Chromatography and column showing different
separated components.

The mobile phase is continuously added from the top of the column and
allowed to move out of the bottom coloumn by opening the stop cock. The
different components of the mixture keep moving downwards as bands in the
column. These are then collected as different fractions in the flask placed at
the bottom of the column. The different components are then obtained from
the eluent by removal of the solvent.

SAQ 1
Name any two stationary phases which can be used in column
chromatography. 95
 Block 2 Chromatography and Ion Exchange

6.4 EFFICIENCY OF THE TECHNIQUE

Column chromatography is a very useful and convenient technique, both for
the effective separation of components of a mixture and for the purification of
an impure compound. Virtually, any type of mixture – occurring naturally (in
plants, animals or in any other source) or obtained synthetically from chemical
reactions, can be separated into its components using column
chromatography. The sample size needed for such a separation could vary
from a few milligrams to grams.

Since a lot of stationary phase materials and different mobile phases-whether

single solvents or suitable mixtures, can be used in column chromatography, it
has the applications in many areas and many different types of compounds
can be separated in pure form from their mixtures or impure samples.

The following points are to be kept in mind for this technique.

1. Type of Adsorbent or the Stationary Phase

Different types of compounds are adsorbed to different extents on

different stationary phases.

Unsaturated, aromatic and polar compounds such as alcohols, acids and

amines are adsorbed on polar adsorbents such as metal oxides and
magnesium silicate.

Silica gel and magnesium silicate are acidic adsorbents and they adsorb
bases. Alumina adsorbs both acidic and basic substances.

Kieselguhr is a weak adsorbent and shows no selectivity for adsorption

of polar compounds. Similarly, sucrose is the weakest adsorbent while
charcoal is the strongest adsorbent.

The adsorbents can be activated by heating when these lose water and
other adsorbed molecules. Silica gel can be activated by heating from
100oC to 120oC for one hour while alumina is to be heated at 400oC.

The following other characteristics of the adsorbent are also important:

 It should be colourless, easily available and inexpensive.

 It should not react with the sample.

2. Particle size of the Stationary Phase

Its particles should be of uniform size. The finely divided nature of

stationary phase leads to better separations. Particle size from74 to149
is suggested with the mesh size of 100 to [Link] still smaller size, the
column packing may not be efficient and it may cause irregular zones.
Also, it will be difficult for the solvent to pass through such a packed
column.

3. Surface Area of the Stationary Phase

The larger the surface area available, the larger will the possibility of
interaction of molecules adsorbed on the stationary phase with the
96 mobile phase; hence, better will be the separation.
Unit 6 Adsorption Chromatography

Also, the smaller size will provide more surface area. But, the size has to
be optimally chosen as too small particle size also slows down the
mobile phase and can have cracks in the column, as mentioned above.

4. Length of the Column

The longer the column, the better will be the separation. The ideal length
to width ratio for the column is 20:1 or 30:1.

5. Sample to Adsorbent Ratio

Ideally, the sample to the adsorbent ratio is chosen as 1:20 to 1:50.

Otherwise, more concentrated samples may not lead to clear
separations.

6. Choice of Mobile Phase

The mobile phase should be appropriately selected. It should not be

viscous and it should not react with the components. The flow rate of the
solvent or the mobile phase should be uniform and not very fast. This will
give better separation of bands of different components of the mixture
without much tailing.

SAQ 2
How does particle size of stationary phase affect the separation in column
chromatography?

6.5 MECHANISM OF SEPARATION

The separation of components of mixture using adsorption chromatography
(such as by use of column chromatography) involves their adsorption on the
stationary support. The mobile phase then displaces the different components
selectively one by one.

The following aspects, thus, need to be considered for the separation process:

 How strongly the component is adsorbed to the stationary phase?

 How much is the surface area of the stationary phase?

 What is the binding strength of the mobile phase is there to get adsorbed
on the stationary phase and how much it can displace the adsorbed
components(s)?

So, as the process of adsorption is involved in such separations, the different

forces involved are hydrogen bonding, van der Waals forces and dipole-dipole
interactions.

Hence, the polarity of the mobile phase will also affect how effectively the
separations can be carried out. The separations will also be affected by the
nature of the stationary phase used. For example silica gel which is polar in
nature; polar solvents may have better interactions with it as compared to the
non-polar ones. 97
 Block 2 Chromatography and Ion Exchange

Having understood the details of the adsorption chromatography, let us now

compare the two chromatography methods which have studied in this unit and
the previous unit.

6.6 ADSORPTION AND PARTITION

CHROMATOGRAPHY
So far we have studied that both the adsorption and partition chromatographic
techniques have the following commons points:

 These chromatographic techniques are useful in separating the

components of a mixture or in the purification of compounds.

 Both these techniques involve stationary and mobile phases and the
mobile phases in both these techniques are liquids.

However, the main differences between these techniques are as follows:

 In the adsorption chromatography, the separation of components occurs

on the basis of their adsorption while in partition chromatography, the
separation of components takes place by their partitioning between the
stationary and the mobile phases.

 Adsorption chromatography is a liquid-solid chromatography while partition

chromatography is a liquid-liquid chromatography.

 Adsorption chromatography requires more amount of solvent and more

time than partition chromatography.

Needless to mention here that suitable mobile phase should be used and
other factors need to be taken care of while separations using these
chromatographic techniques are meaningfully achieved.

So far, we have learnt about many important aspects of adsorption

chromatography. But, we are yet to know more about the development of the
chromatogram, i.e. how to obtain the separated components of the mixture in
the column.

Let us learn about it in detail. But before that answer the following SAQ.

SAQ 3
List the similarities between adsorption and partition chromatographic
techniques.

6.7 DEVELOPMENT OF CHROMATOGRAMS:

FRONTAL ANALYSES, DISPLACEMNET AND
ELUTION METHODS
The development of chromatograms is a very important aspect of
separations using column chromatography. The development of
98 chromatograms can be done in different ways. It would be very interesting to
Unit 6 Adsorption Chromatography

know about the difference between them and understand which method will be
more useful in which situation. Let us study each one of these ways is more
detail.

(i) Frontal Analysis

Let a mixture containing three compounds X, Y and Z, is to be separated by

using frontal analysis. See Fig. 6.3 a) in which such a mixture is shown in the
column as the top layer. The three compounds X, Y and Z are shown by red,
green and blue colours.

a) b) c)

Fig. 6.3: a) Frontal analyses, b) Displacement and c) elution methods with their
graphical representations.

 Note that before placing the mixture (X+Y+Z) on the column, the
column has been saturated with the mobile phase. A large amount of
the sample of X+Y+Z is introduced from the top. Then the mobile
phase is continuously added from the top and the components start
passing down the column along with the mobile phase. The least
.
adsorbed component X (represented as ) will reach the bottom and will
be eluted first.

 Next to it, in the second fraction, a mixture of X+Y will be eluted.

 Finally, the third fraction will contain X+Y+Z, i.e. all the three
components. Such a separation is shown in graphical mode also in
Fig.6.3 below a).

Thus, in such a mode of separation, only one component has been obtained in
the pure form. 99
 Block 2 Chromatography and Ion Exchange

Such separations were used long back when chromatography methods were
not that much advanced.

Also, this mode of development of chromatogram was useful when only one
component which was the desired component , was to be obtained in pure
form and it could not be separated from the mixture by other methods.

But, otherwise such a method is not very useful.

(ii) Displacement Method

Here, again we start with the same mixture but this time we add a displacing
agent with the mobile phase. Let all the components X, Y and Z are strongly
adsorbed to the stationary phase. Here, the displacing agent is so taken that it
is more strongly adsorbed than any of the components X, Y and Z. Thus, the
displacing agent will compete with X, Y and Z for the adsorption sites and
displace X, Y and Z into the mobile phase.

The order in which X, Y and Z are displaced will be according to the fact that
the least out of the three will be desorbed first and hence, eluted first. Hence,
will be desorbed first and eluted first. Let us say that it is the X which is eluted
first.

This will be followed by another fraction in which both X and Y will be eluted
together, i.e. Y will also start getting displaced along with X. After that, pure Y
will get eluted.

Next, a mixture of Y and Z will start coming out in the eluent. Lastly, pure Z will
get eluted. Then, at the end, the displacing agent will be passing out of the
column.

Here also, this method is not perfect for separation as the quantitative
amounts of pure X, Y and Z are not obtained at the end of the procedure. The
graphical representation of such a separation is shown in the lower part in b)
in Fig. 6.3.

Let us now study what happens in elution method.

(iii) Elution Method

Let us start with the same mixture of X+Y+Z taken at the top of the column.
Here, there are two options of elution-isocratic elution and gradient elution.

In isocratic elution, the solvent or the mobile phase having the same
composition (if a mixture of two or more solvents is taken) is used throughout
the separation process. Different fractions of components start appearing as
different bands in the column depending upon which components get eluted
first. Again, the one which is least adsorbed will come out first from the
column. Let this be X, here also.

But, if the other components e.g. Y and Z are adsorbed to quite different
extents; then, the use of a single solvent, i.e. mobile phase of the single
composition many not be able to move Y and Z down the column along with
the mobile phase. Here, the gradient elution is used .The solvents of different
100 polarities, i.e. with increasing order polarities are used to elute the different
Unit 6 Adsorption Chromatography

components Y and Z in the pure form with a reasonable gap or difference

between their bands in the column.

Thus, all the three components X, Y and Z could be separated in the pure form
quantitatively using elution method.

The graphical representation in c) in Fig. 6.3 clearly shows this.

Here, we can conclude that elution method, especially the gradient elution is
the most useful method for separating the components of a mixture using
column chromatography.

SAQ 4
Why is gradient elution more useful than the other methods of development of
chromatograms?

6.8 SUMMARY
In this unit, we have learnt that

 Adsorption chromatography is classified as the three-dimensional

chromatographic technique.

 Adsorption chromatography is based on the phenomenon of adsorption

of various components of a mixture on the stationary phase.

 The mobile phase on passing on to the stationary phase, carries along

with it, the least adsorbed component at a faster rate than those which
are strongly adsorbed on the stationary phase.

 The technique of column chromatography is very efficient method for the

separation of a wide range of mixtures.

 A variety of adsorbents and mobile phases are used in column

chromatography for separating different mixtures.

 Adsorption and partition chromatographic techniques have similarities as

well as differences and these are useful as per the requirement of the
separation.

 The gradient elution is more useful than the frontal analysis and the
displacement methods.

6.9 TERMINAL QUESTIONS

1. What factors need to be taken care of with respect to the mobile phase
to achieve better separations in column chromatography?

2. Discuss the activation of adsorbents.

3. Why is frontal analysis not a good method of the development of

chromatogram? 101
 Block 2 Chromatography and Ion Exchange

6.10 ANSWERS
Self Assessment Questions

1. Silica gel, alumina or any other.

2. Particle size of 74-149  is recommended.

The smaller the particle size, the better is the separation. But too small
particles may lead to irregularities in the packing of the column. Hence,
the particle size needs be carefully chosen.

3. These chromatographic techniques are useful in separating the

component of a mixture or in the purification of compounds.

Both these techniques involve stationary and mobile phases and the
mobile phase in both these techniques are liquids.

4. The gradient elution is more useful than the other methods of

development of chromatograms because by using elution method, all the
components of a mixture could be separated in the pure form
quantitatively.

Terminal Questions
1. The mobile phase should not be viscous and should not react with the
components of the mixture. Further, the flow rate should not be fast and
should be uniform.

2. The adsorbents can be activated by heating when these lose water and
other adsorbed molecules. Silica gel can be activated by heating from
100oC to 120oC for one hour while alumina is to be heated at 400oC.

3. This is so because in such a mode of separation, only one component

has been obtained in the pure form.

102
Unit 7 Ion Exchange Chromatography

UNIT 7
ION EXCHANGE
CHROMATOGRAPHY

Structure
7.1 Introduction 7.4 Mechanism of Ion Exchange
Expected Learning Outcomes 7.5 Summary
7.2 Ion Exchange Materials 7.6 Terminal Questions
7.3 Principle of Ion Exchange 7.7 Answers
Ion Exchange Capacity

7.1 INTRODUCTION
You have studied about the charomatographic techniques based on partition
and adsoprtion respectively, in Units 5 and 6 of this [Link] this unit ,you will
study about one more chromatographic technique called ion exchange
chromatography based on ion exchange .

Ion exchange has the popular application in water softening. The analytical
applications of ion exchange chromatography include the analysis of sugars,
amino acids, proteins, nucleotides, pharamaceuticals,clinical samples etc.

Ion exchange is also important in the recovery of metals from industrial

wastes, separation of rare earths, decontamination of cooling water from
nuclear reactors. Glen T. Seaborg also utilised this technique to identify
elements of the 5f series which were obtained in a sequence similar to that for
4f elements.

We will begin the disussion of this unit by describing the types of materials
which are used for ion exchange. Then, the principle of ion exchange
chromatography will be explained alongwith the ion exchange capacity.

Finally, we will focus our attention on the mechanism of ion exchange.

Expected Learning Outcomes

After studying this unit, you should be able to:
 explain the importance of ion exchange chromatography;
103
Block 2 Chromatography and Ion Exchange

 give examples of natural and synthetic ion exchangers;

 classify the exchangers as weak and strong ion exchanges;

 discuss the principle of ion exchange chromatography;

 define ion exchange capacity of a resin;

 explain the mechanism of ion exchange; and

 describe different factors affecting the retention time of ions on the resin
and hence, affecting the efficiency of separation.

7.2 ION EXCHANGE MATERIALS

The phenomenon of ion exchange was observed by two agricultural chemists
Thomson and Way in 1850 who observed the exchange of ammonium ions
with calcium ions in the soil. The use of clay minerals for ion exchange was
thus realised and their use for water softening was attempted.

Synthetic ion Ion exchange is a process in which there is an exchange of ions of like sign
exchange resins have between a solution and an insoluble solid in contact with the solution. Clearly,
found widespread the solid must contain ions of its own to exchange, in addition, it must have a
laboratory and permeable structure of large in addition, it must have a permeable structure of
industrial applications large specific surface area so that the solvent and solute ions can readily
for water softening, come in contact with the solid surface.
water deionisation,
During the course of time, more such materials were synthesised. The first
purification of
synthetic ion exchanger was prepared by two German chemists – Harm and
solutions and
Rumpler in 1903. Meanwhile, permutits were also tried. However, in 1935,
separation of ions.
Adams and Holmes – two English chemists observed that crushed
phonograph records exhibited ion-exchange properties.

Before taking up the detailed description of synthetic ion exchange materials,

let us first know about some natural ion exchangers.

Natural ion exchangers: These include zeolites having cation exchange

properties. Their examples being the following minerals:
Zeolites are
aluminosilicate (i) Analcite – Na [SiAlO6]2. H2O
minerals that contain
(ii) Chabazite – (CaNa) [SiAlO6]2. 6 H2O
alkali and alkaline-
earth metals. They (iii) Naturalite – Na2 [Si2Al2O10]. 2 H2O
are three-
dimensional, These have three dimensional networks with negatively charged lattice. The
microporous,
crystalline solids with alkali and alkaline earth cations which move freely, balance the negative
cavities and channels charge. These behave as counter ions and can be exchanged with other
in which cations, counter ions.
water or small
molecules can reside. –  3
They are often called The exchange of OH ions for Cl , SO4 and PO4 ions has been reported in
molecular sieves. montmorillonite, kaolinite and feldspar of sodalite and camerinite groups.

Zeolites have poor mechanical strength, less abrasive resistance and are
104
decomposed partially by acids and alkalis.
Unit 7 Ion Exchange Chromatography

Some types of coals are also used as cation exchangers after stabilising with
2+ 2+
metal ion solutions. These contain weak acidic and carboxylic groups and are Ca or Mg forms a
thus, weak cation exchangers. But, these can be converted to strong cation coating on the inner
exchangers by sulphonation by treatment with strong sulphuric acid. walls of a boiler and
cannot be easily
Let us now explore more about synthetic ion exchangers.
removed. This is
Synthetic Ion Exchangers known as scale
formation. Then,
High molecular weight copolymer beads made by copolymerisation of styrene zeolites are used for
with divinylbenzene are used. The use of ~80% divinylbenzene leads to cross 2+
removing Ca and
linking of the polymer to give it mechanical stability. Acidic and basic functional 2+
Mg ions from hard
groups are then bonded to this polymeric structure to convert it into weak or water before water is
strong cation as well as anion exchangers as shown below in Fig. 7.1. put in the boiler.
CH=CH2 CH=CH2 These zeolites are
known as molecular
sieves.

CH=CH2

Exchange Reaction
styrene divinylbenzene
2+
CH2 CH CH2 CH CH2 CH CH2 CH Na2Z (solid) + Ca
(solution) ⇄
+
CaZ (solid) + 2 Na

cross link
R (solution)
CH2 CH CH2 CH CH2 CH CH2 CH
Regeneration

CaZ + 2NaCl (10%

R R
R-functional group solution) 
CH2 CH CH2 CH CH2 CH Na2Z (solution) + CaCl2
(solution)

Fig. 7.1: Copolymer of styrene-divinylbenzene with R functionalised chain to

convert it into cation or anion exchanger.

Table 7.1 shows different R groups corresponding to strong and weak cation
and anion exchangers.

Table 7.1: Strong and Weak Cation and Anion Exchangers

Type of Ion Exchanger Functional Group, R Example

 
Weak cation exchanger Carboxylic group  COO ,  CH2COO
 
Strong cation exchanger Sulphonic group SO3 ,  CH2CH2SO3

 
Weak anion exchanger Amine  NH3 , CH 2CH 2 NH(CH2CH 3 )

 
Strong anion exchanger Quaternary amine  CH 2 N(CH 3 )3 ,  CH 2CH 2  N(CH 2CH 3 ) 3

105
Block 2 Chromatography and Ion Exchange

SAQ 1
Give examples of two natural ion exchangers.

SAQ 2
What is the role of divinylbenzene in the copolymer with styrene?

7.3 PRINCIPLE OF ION EXCHANGE

Ion exchange resins are porous and insoluble in aqueous medium. These
contain many ionic functional groups per molecule as shown above in Fig. 7.1.

When such an ion exchanger is immersed in an aqueous solution, containing

the ions to be exchanged, an equilibrium gets established.

Let us first understand this for cation exchangers. Let the resin, i.e., cation
exchanger is immersed in a solution of metal ions, Mn+ which are to be
exchanged. We can write the equilibrium for both the weak and strong cation
exchangers as follows:
n
n RzCOOH  M(aq) 
⇌ (RzCOO)n M  n H(aq) … (7.1)
Weak cation
exchanger

n
n RzSO3H  M(aq) ⇌ (RzSO-3 ) n Mn   n H(aq)

… (7.2)
(solid) (solid)
Strong cation
exchanger

Here, Rz denotes the resin.

The equilibrium constant, K or the selectivity coefficient, can be written for the
reaction shown in Eq. 7.2 as follows:

n
[RzSO3 ) n M] [H ] (aq)
K … (7.3)
[RzSO3H ] n [Mn  ] (aq)

Hence, terms in the square brackets indicate the surface concentrations (or
more strictly the activities).

Eq. 7.3 can be rearranged, in terms of the distribution ratio as follows:

Amount of M in stationary phase

R
Amount of M n  in aqueous phase

[RzSO3 ) n M] [RzSO3H ] n
 … (7.4)
n
[Mn  ] (aq) [H ] (aq)
106
Unit 7 Ion Exchange Chromatography

Thus, the equilibrium in Eqs. 7.1 and 7.2 can be shifted to left or right by
increasing [H+] or [Mn+].

Weak cation exchangers are used in the pH range 5-14 while strong cation
exchangers can be used in the pH range 1-14. Weak cation exchangers hold
the protons tightly in the low pH range and hence, exchange will not occur.

Weak acid cation exchangers will not completely exchange cations of very
weak bases. These are exchanged by strong cation exchangers only.

Weak acid cation exchangers, however, separate strongly basic substances

e.g., peptides or proteins and multifunctional ionic compounds.

The strong cation exchangers are mostly used for separating the complex
mixtures.

Let us now see what happens with the anion exchangers. The anion exchange
equilibrium for the strong and weak anion exchangers can be represented as
given below:

n RzNR3 OH  A n  ⇌ (RzNR3 ) n A  n OH

Strong anion exchange
and
n RzNH3 OH  A n  ⇌ (RzNH3 )n A  n OH
Weak anion exchanger

Here, the –OH ions are exchanged for the anion An. The strong anion
exchangers work in the range of pH from 1-12 while weak anion exchangers
work in the pH range 0-9.

The weak anion exchangers are used for strong acids.

Having understood the basic concepts of anion and cation exchangers, it is

important to understand about their capacity. Let us understand it in more
detail.

7.3.1 Ion Exchange Capacity

We have studied above that the counter ions of a resin are exchanged in an
exchange process. The counter ion content in the given amount of an
exchanger, is the fixed charges which are balanced by counter ion charges.

The capacity of an ion exchanger is the number of ion equivalents in the

specific amount of the material. But, the context of its usage needs to be
specified.

We define the total capacity of the resin as the number of ionic (or potentially
ionic) sites per unit volume or weight of the resin.

The dry weight total capacity is usually expressed in milliequivalents per gram
of anhydrous resin. It is expressed as meq/g dry H+ or Cl form.

The wet volume capacity is number of sites per unit volume of the water
swollen resin. It is the maximum theoretical capacity; however, the
performance of the resin is generally based on its volume. It may be
expressed as milliequivalents per millilitre. 107
Block 2 Chromatography and Ion Exchange

The operating capacity of a resin in a particular cycle is the net number of sites
utilised in a given volume in that cycle. It is expressed in the same terms as
total capacity or as a percent of total capacity.
One more capacity is dynamic or breakthrough capacity which is used in
column operation and depends upon the operating conditions.
The exchange capacity affects the retention of the solute. The higher capacity,
the higher retention and improved will be the resolution.

Besides the capacity of the resin, the particle size of resin particles is also
important. The preferred effective size is of diameter 0.4-0.6 mm. The
corresponding particle size distribution is between 20-50 mesh screens. Note
that hydrated particles (i.e., swollen resin) are present in the aqueous medium.
The decrease in the particle size, decreases the time required to attain the
equilibrium; hence, the efficiency for a given volume of resin increases. In
other words, lesser volume will be required with smaller particle size.

SAQ 3
Write the expression for the equilibrium constant for a weak anion exchange
resin.

7.4 MECHANISM OF ION EXHCNAGE

The stationary phase in the ion exchange chromatography is the matrix
containing the ionisable functional groups with fixed ions and carrying
oppositely charged displaceable ions.
The mobile phase consists of an aqueous buffer system with small amount of
organic solvent. Note that exchange of cations or anions occurs with the
counter ions of the resin, as the case may be, if we are separating a mixture of
cations or anions, respectively.
The selectivity coefficient determines the affinity of the resin for the particular
ion. The large value of selectivity coefficient indicates strong tendency for the
resin to retain the ion and vice-versa.

For a typical strong cation exchange resin, the decreasing order of selectivity
coefficients is
3+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ 2+ +
Al > Ba > Pb > Ca > Ni > Cd > Cu > Co > Zn > Mg > Ag >
+  + + +
K > NH4 > Na > H > Li

This sequence indicates that the highly charged ions bind more strongly as
compared to ions with lower charge. For the ions having similar charge, the
ions with smaller hydrated radii or those which are more polarisable bind more
strongly to the resin.

Similarly, the order for strong anion exchangers is as follows:

SO42  I  HSO4  NO3  Br  NO2  Cl  HCO3  CH3COO  OH  F

Here again, the highly charged ions and those with smaller hydrated radii bind
108 more strongly to the resin.
Unit 7 Ion Exchange Chromatography

With regard to the aqueous buffer mobile phase, the pH and the ionic
composition are related to the retention time. Gradient elution involving the
change in pH or ionic strength, is used for eluting different cations. Here, if dil.
HCl is used as the mobile phase, then the increase in the concentration of HCl
+
increases the elution rate as more H ions are now available to compete
successfully for the ion-exchange sites.

SAQ 4
Why is the selectivity coefficient important?

7.5 SUMMARY
In this Unit, you learnt that

 Ion exchange materials are useful materials for separating a variety of

substances containing complex mixtures.

 Ion exchange materials could be natural or synthetic ones. These are

categorised as cation exchange materials or anion exchange materials.
Further, these could be weak or strong ion exchangers.

 For cation exchange, there is a choice between strong acid groups

(RSO3H) or weak acid resins containing carboxylic acid (RCOOH) groups.
The former has wider applications. Anion exchange resins contain basic
functional groups attached o the polymer molecule. These are generally
amines; strong base exchanges are obtained with tertiary amines and
(quaternary ammonium salts) [RN(CH 3 ) 3 OH  ] and weak types with
primary and secondary amines.

 Synthetic ion exchange resins are high molecular weight polymeric

materials containing large number of ionic functional groups per molecule.

 Ion exchange capacity of an exchanger can be defined in different ways.

 The particle size of the resin plays an important role in the retention and
hence, separation of materials. The other features being the temperature
and the pH of the mobile phase.

 The mechanism of ion exchange involves exchange of counter ions of the

resin. The selectivity coefficient determines the affinity of the ion for the
resin.

 Aqueous buffers along with some organic solvent is used in elution.

 Gradient elution using change of pH and ionic strength can separate a

mixture into its components.

7.6 TERMINAL QUESTIONS

1. What are the limitations of zeolites as ion exchangers?

2. What functional groups are present in a strong cation exchanger and a

weak anion exchanger? 109
Block 2 Chromatography and Ion Exchange

3. Differentiate between dry weight total capacity and wet volume capacity of
a resin.

7.7 ANSWERS
Self-Assessment Questions
1. Analcite, chabazite or any other suitable example.

2. It acts as a cross linking agent.

[(Rz NH 3 )n A] [  OH] n
3. K
[Rz NH 3  OH] n [ A n 1 ]

4. The selectivity coefficient determines the affinity of the resin for the
particular ion. The large value of selectivity coefficient indicates strong
tendency for the resin to retain the ion and vice-versa.

Terminal Questions
1. Zeolites have poor mechanical strength, less abrasive resistance and
are partially decomposed by acids and alkalis.

2. Strong cation exchangers have  SO3 H group and weak anion


exchangers have  N H3 group.

3. The dry weight total capacity is total capacity of anhydrous resin

whereas wet volume capacity is the number of sites per unit volume of
water- swollen resin.

110