DATED: September 16TH, 2009

EXPERIMENT#

OBJECT:
TO OBSERVE THE SEPARATION OF A MIXTURE BY THE METHOD OF PAPER
CHROMATOGRAPHY.

REQUIREMENTS:
Mixture obtained by column chromategrapghy by rose, filter paper of 8/3 size,
pencil, paper chromatography apparatus, solvent, uv chamber, capillary tubes,
scale

THEORY:
PAPER CHROMATOGRAPHY
DEFINITION
Paper chromatography is an analytical chemistry technique for separating and identifying
mixtures that are or can be coloured, especially pigments by the flow of solvent on a filter paper.

ADVANTAGES AND DISADVANTAGES

• This technique provides an easy way to separate the components of a
mixture.
• in analytical chemistry, technique for separating dissolved chemical
substances by taking advantage of their different rates of migration across
sheets of paper.
• It is an inexpensive but powerful analytical tool
• requires very small quantities of material
• Paper chromatography is a useful technique because it is relatively quick

PHASES
In paper chromatography, the stationary phase is a very uniform absorbent paper.
The mobile phase is a suitable liquid solvent or mixture of solvents.

THE ESSENTIAL STRUCTURE OF PAPER

Paper is made of cellulose fibres, and cellulose is a polymer of the

simple sugar, glucose.

The key point about cellulose is that the polymer chains have -OH groups sticking
out all around them. To that extent, it presents the same sort of surface as silica gel
or alumina in thin layer chromatography.

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It would be tempting to try to explain paper chromatography in terms of the way

that different compounds are adsorbed to different extents on to the paper surface.
In other words, it would be nice to be able to use the same explanation for both thin
layer and paper chromatography. Unfortunately, it is more complicated than that!

The complication arises because the cellulose fibres attract water vapour from the
atmosphere as well as any water that was present when the paper was made. You
can therefore think of paper as being cellulose fibres with a very thin layer of water
molecules bound to the surface.

It is the interaction with this water which is the most important effect during paper
chromatography.

PAPER CHROMATOGRAPHY USING A NON-POLAR SOLVENT

Suppose you use a non-polar solvent such as hexane to develop your
chromatogram.

Non-polar molecules in the mixture that you are trying to separate will have little
attraction for the water molecules attached to the cellulose, and so will spend most
of their time dissolved in the moving solvent. Molecules like this will therefore travel
a long way up the paper carried by the solvent. They will have relatively high Rf
values.

On the other hand, polar molecules will have a high attraction for the water
molecules and much less for the non-polar solvent. They will therefore tend to
dissolve in the thin layer of water around the cellulose fibres much more than in the
moving solvent.

Because they spend more time dissolved in the stationary phase and less time in
the mobile phase, they aren't going to travel very fast up the paper.

The tendency for a compound to divide its time between two immiscible solvents
(solvents such as hexane and water which won't mix) is known as partition. Paper
chromatography using a non-polar solvent is therefore a type of partition
chromatography.

PAPER CHROMATOGRAPHY USING A WATER AND OTHER

POLAR SOLVENTS
A moment's thought will tell you that partition can't be the explanation if you are
using water as the solvent for your mixture. If you have water as the mobile phase
and the water bound on to the cellulose as the stationary phase, there can't be any
meaningful difference between the amount of time a substance spends in solution
in either of them. All substances should be equally soluble (or equally insoluble) in
both.

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If water works as the mobile phase as well being the stationary phase, there has to
be some quite different mechanism at work - and that must be equally true for
other polar solvents like the alcohols, for example. Partition only happens between
solvents which don't mix with each other. Polar solvents like the small alcohols do
mix with water..

INTERPRETING THE DATA

The Rf value for each spot should be calculated. Rf stands for "ratio of fronts" and is
characteristic for any given compound. Hence, known Rf values can be compared to
those of unknown substances to aid in their identifications.

(Note: Rf values often depend on the temperature, solvent, and type of paper used
in the experiment; the most effective way to identify a compound is to spot known
substances next to unknown substances on the same chromatogram.)

In addition, the purity of a sample may be estimated from the chromatogram. An

impure sample will often develop as two or more spots, while a pure sample will
show only one spot.

PRINCIPLE
This technique proceeds by a partly partition (distribution) and partly adsorption mechanism.
The constituents of the mixture are distributed between the water held on the filter paper (water
acts as a stationary phase) and organic solvent (mobile phase).

TYPES
ASCENDING AND DESCENDING PAPER CHROMATOGRAPHY
Depending upon the direction of the solvent travel, paper chromatography is termed as
ascending or descending type. When the solvent travels upward on the paper, it is termed as
ascending paper chromatography and descending paper chromatography when the solvent
travels in the downward direction.

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An experimental setup for paper chromatography

APPLICATION
• Chemical separations
• Widely used in protein and nucleic acid blotting
• Separating and identifying mixtures
• Chromatography is an important tool of the forensic chemist in solving
crimes, DNA and RNA sequencing, among others.
• Paper chromatography is one method for testing the purity of compounds

PROCEDURE
THE PROCEDURE IS SASME AS THIN LAYER CHROMATOGRAPHY INCLUDING SO NOT
WRITING DETAILS IN FOLLOWING PROCEDURE

In this procedure, a drop of the test solution is applied as a small spot near one edge of the filter
paper and the spot is dried. Then, the end of the paper strip is dipped into a developing solvent.
The liquid is sucked up through the capillaries of the paper; it reaches the mixture and removes
its components at various speeds.
After the solvent front has reached a suitable height (15-20 cm), the paper is dried.
The separated components are made visible with a suitable reagent called
'visualization reagent'. HERE WE HAVE USED UV CHAMBER FOR VISUALIZATION

The movement of any compound relative to the solvent is determined by the measurement of RF
values, viz.,

Then,

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Pictorial representation of RF value

ROSE
PHARMACOGNOSTIC FEATURES:
BOTANICAL NAME: ROSA

FAMILY: ROSEACEAE

ENGLISH NAME: ROSE

LOCAL NAME: gulab

PARTS USE: FRUIT, BARK, leaves

CONSTITUENTS: Roses contain vitamin C , pectin, malic & citric acids, and
antioxidant flavonoids, TANNINS

ACTIONS: Mildly astringent, aperient, carminative, and refrigerant, cardiac tonic.

Antidepressant, Feminine scent, antiseptic essence.

Uses

• Roses are popular garden shrubs, as flowering shrubs. They are also
grown as cut flowers, as one of the most popular and commonly
sold florists' flowers.
• A roses are grown for scented foliage
• Roses are also of great value to the perfume industry. An attar of roses is
distilled from the flowers.
• The rose hip, the fruit of some species, is used as a minor source of
Vitamin C.
• Roses may also be planted as hedging, and for game cover.

MEDICINAL USES
Because of the medicinal properties of rose, it is widely used in Ayurveda medicine. One cup
of fresh rose hips has the equivalent amount of vitamin C in 60 oranges. Rose tea (tea made
with rose petals and hips) is not a new idea. Here are 10 extraordinary health benefits of rose
tea.

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Why roses are considered an ideal gift to express love? There is an interesting reason for this.
Roses soothe heart and emotions. They also balance the mind.
The reasons for the diuretic effects of the roses are they contain Vitamin C, pectin, malic and
citric acids.
1. It clears toxins and heat from the body. As a result it has a cooling effect on the body.
2. It can relieve from sore throat, runny nose and blocked bronchial tubes
3. It is useful to people those prone to chest problems by fighting against infections
4. Rose tea helps to fight the infection in the digestive tract and re-establish the normal
bacterial population of the intestines
5. It relieves fluid retention and hastens the elimination of wastes through kidneys
6. It is a wonderful remedy for dysentery, diarrhea and gastro enteritis
7. It is a laxative. It works as a remedy for all liver problems including sluggishness and
constipation.
8. It cleanses the liver and gall bladder and promotes bile flow
9. Rose petal tea can be used to relieve uterine congestion causing pain and heavy
periods. It is an excellent remedy for irregular periods and infertility.
10. It has an uplifting effect on the nervous system and can relieve insomnia, depression
and fatigue

OBSERVATION

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 DATED: September 16TH, 2009

RESULT

CONCLUSION

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