NAME: Kwame Sam-Asmah

INDEX: 9644606

DEMONSTRATOR: Darkwa Maxwell

EXPERIMENT: O.3.1.1

DATE: 13th October 2008.

TITLE: SEPARATION METHODS II

AIMS\OBJECTIVES:

 To separate the various components of a given sample.

 To calculate the R f values for a given sample.

INTRODUCTION:

Separation is a technique used in isolating and or purifying a chemical constituent or a group of

chemically related substances. During separation, either the interfering substances are removed or the
substance to be separated is isolated. For a successful separation, the substances to be separated must
differ in a chemical or physical property due to which the separation can occur. Some form of separation
includes ion exchange, electrophoresis absorption, adsorption and precipitation. In this experiment, the
separation method employed is chromatography.

Chromatography is an analytical technique, which involves the separation of components in a

sample by distribution of the components between two immiscible phases namely, a stationary phase and
a mobile phase. This technique is based on differences in solubility. The stationary phase consists of a fixed
bed of small particles with a large surface area while the mobile phase is a fluid that moves constantly
through or over the face of the stationary phase. An evaluation of the separation can be achieved by
determining the retardation of each separated compound. A factor namely the Retardation factor Rf for
each eluted substance is introduced to enable the qualitative evaluation of the chromatogram. The
retardation factor is defined as a measure of that fraction of the total elution time that it spends in the
mobile phase. It is also defined as the ratio of the distance travelled by the mobile phase in a particular
time.
R distance travelled bythe compound (¿cm)
f=
distance travelled by the solvent front (¿cm)

Chromatographic processes are classified according to the type of mechanism involved in the
separation, which is based on the type of stationary phase. The different types are sorption, ion exchange,
partition and exclusion. More than one of these four mechanisms may be occurring simultaneously in any
chromatographic separation. Chromatographic techniques can also be characterised according to the type
of instrumentation or apparatus used. These include Column Chromatography, Paper Chromatography,
Thin-layer Chromatography, Gas Chromatography and High-pressure liquid Chromatography. Paper and
Thin-layer chromatography are the chromatographic techniques used in this experiment.

In paper chromatography, a liquid sample flows down a vertical strip of adsorbent paper, on which
the components are deposited in specific locations. The stationary phase in a paper chromatography
consists of a sheet of paper of suitable texture and thickness. The paper should be made of highly purified
cellulose, which has a great affinity for water and other polar solvents. The cellulose paper will normally
have a thin coat of water adsorbed from the air since it is very hydrophilic. In this technique, the sample to
be separated is spotted onto a strip of the paper with a micropipette and the chromatogram is developed
by placing the bottom of the paper into the solvent phase. The solvent is drawn up the paper by capillary
action and the sample components move up the paper at different rates, depending on their solubility and
their degree of retention by the paper. If the paper used is wide, several samples and standards can be
spotted and developed simultaneously.

Thin-layer Chromatography is very similar to paper chromatography except that the stationary
phase is a uniform layer of a finely divided powder that has been coated on the surface of a glass, plastic or
aluminium sheet that is held in place by a binder. The capacity of the system is dependent on the thickness
of the layer, which may range from 0.1 to 2.0mm. Thinner layers are used primarily for analytical
separations while thicker layers are used for preparative work. The most commonly used stationary phases
are adsorbents such as silica gel, alumina and powdered cellulose. When such substances are used,
separations occur due to adsorption of the solutes from the mobile phase onto the surface of the thin-
layer.

APPARATUS USED

 Capillary tubes
 Measuring cylinder
 Beaker
 Whatman#1 filter paper
 A cover
 A rule and a pencil

CHEMICALS USED

 Distilled water
 Butanol
 Acetic acid

PROCEDURE
PAPER CHROMATOGRAPHY

A Solution of butanol, acetic acid and water was made in a beaker according to the ratio, 50:1:49, 60:1:39,
40:2:58 and 20:3:77.

A clean sheet of whatman#1 filter paper was taken, and a light pencil line parallel to the bottom of about
2.0cm away were made on each of them.

Using the capillary tubes, a small amount of each appropriate solution was placed along the line on the
filter paper. The paper was then made dried for some few minutes.

The paper was then placed gently into the beaker containing the solvent. The spots were above the level of
the solvent. It was then covered and left to stand for separation to take place.

After some time, the paper was removed and the level at which the separations were made was calculated
and the R f of each was calculated.

PRECAUTIONS TAKEN

 The addition of the sample to the stationary phase was in bits to obtain better separation.
 Protective clothing was worn.
 The spots were positioned in a way that it was above the solvent in the beaker when it was
lowered.

TABLE OF RESULTS

PAPER CHROMATOGRAPHY

 Blue Ink

Solvent Used – butanol (50ml) + H2O (10ml)

Distance moved by solvent – 11.1cm

Distance moved by compound – 11.1cm

distance moved by compound
R.F =
distance moved by solvent front
11.1cm
= =1
11.1cm

 Red Ink

Distance moved by solvent front = 11.6cm

Distance moved by solvent front = 11.6cm

distance moved by compound
R.F =
distance moved by solvent front
11.6 cm
= =1
11.6 cm
ERROR ANALYSIS

 Most of the beakers were not covered after the solvent phase was prepared and when the solution
was left for separation to occur. For this reason, the atmosphere in the beaker was not saturated
and this could have affected the separation.
 After the sample have been added to the stationary phase, it was not allowed to dry before placing
it into the beaker.
 The R f values were not marked immediately the paper was removed.

DISCUSSION

This experiment was an ascending chromatography, this is because the lower edge of the paper \plate was
dipped into the mobile phase to permit the mobile phase to rise on the chromatographic sheet. The
solvent was drawn up the paper\plate by capillary action and the sample components move up the paper
at different rates depending on their solubility and their degree of retention by the paper.

From this experiment, the samples separated into its components but the R f for each sample was
calculated to be 1, because distance moved by compound was equal to distance moved by solvent front.

The separations of the ink is based on the differences in their affinities. Also the amount of development
depended on the complexity of the mixtures of the solutes being separated. The paper adsorbs which
caused it to swell this is because a water immiscible solvent which consist of a mixture of organic solvent
and water was used. The thin-layer chromatography has some advantages over the paper
chromatography, this is because it is more sensitive and more versatile than the paper chromatography.
Despite these, none of its components separated, which means that the solvents given were not good
enough to adsorb the components.

In the experiment, some of the beakers were not covered during the separation, thus the atmosphere in
the beaker was not saturated and this could be attributed to the fact that none of the samples separated.
Also the solvents spotted onto the paper were not made to dry before immersing it in the solution.

For the paper chromatography, sample B had a gradual decrease in the distance travelled by the solvent
front as water level increases with decrease in butanol. for the thin-layer, there was also a gradual
decrease in solvent front level as volume of acetic acid decreases with an increase in toluene.

CONCLUSION

The R f values for the two inks were 1 because distance moved by compound was equal to distance moved
by solvent front.

REFERENCES

 Principles of organic chemistry 3rd edition, by T.A Geissman, University of Calofornia, Los Angelos,
publishers: W.H Freeman and company. pages 12-15
 Practical chemistry V, CHEM 369, laboratory manual K.N.U.S.T pages 27-33.
 CHEM 361 LECTURE NOTES BY MR. RAY BRIGHT VOEGBOLOR PAGES 13-22.
 Murray, Royce W. "Chemical Analysis." Microsoft® Student 2008 [DVD]. Redmond, WA:
Microsoft Corporation, 2007

POST LAB
1. distance travelled by the compound=0.24cm

Distance travelled by the solvent front=0.64cm

R distance travelled by the compound
f=
distance travelled by the solvent front

R 0.24 =0.375.
f =¿ ¿
0.64