Paper chromatography is an
analytical method used to separate coloured chemicals or substances.
It is now primarily used as a teaching tool, having been replaced in the
laboratory
A laboratory (; ; colloquially lab) is a facility that provides controlled conditions in which scientific or technological research, experiments, and measurement may be performed. Laboratory services are provided in a variety of settings: physicia ...
by other chromatography methods such as
thin-layer chromatography
Thin-layer chromatography (TLC) is a chromatography technique used to separate non-volatile mixtures.
Thin-layer chromatography is performed on a sheet of an inert substrate such as glass, plastic, or aluminium foil, which is coated with a t ...
(TLC).
A paper chromatography variant,
two-dimensional chromatography
Two-dimensional chromatography is a type of chromatographic technique in which the injected sample is separated by passing through two different separation stages. Two different chromatographic columns are connected in sequence, and the effluent fr ...
, involves using two solvents and rotating the paper 90° in between. This is useful for separating complex mixtures of compounds having similar polarity, for example,
amino acids
Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha am ...
. The setup has three components. The mobile phase is a solution that travels up the stationary phase, due to
capillary action
Capillary action (sometimes called capillarity, capillary motion, capillary rise, capillary effect, or wicking) is the process of a liquid flowing in a narrow space without the assistance of, or even in opposition to, any external forces li ...
. The mobile phase is generally a mixture of non-polar organic solvent, while the stationary phase is polar inorganic solvent water. Here paper is used to support the stationary phase, water. Polar water molecules are held inside the void space of the cellulose network of the host paper. The difference between
TLC
TLC may refer to:
Arts and entertainment Television
* ''TLC'' (TV series), a 2002 British situational comedy television series that aired on BBC2
* TLC (TV network), formerly the Learning Channel, an American cable TV network
** TLC (Asia), an A ...
and paper chromatography is that the stationary phase in TLC is a layer of adsorbent (usually
silica gel
Silica gel is an amorphous and porous form of silicon dioxide (silica), consisting of an irregular tridimensional framework of alternating silicon and oxygen atoms with nanometer-scale voids and pores. The voids may contain water or some other l ...
, or
aluminium oxide), and the stationary phase in paper chromatography is less absorbent paper.
R''ƒ'' value, solutes, and solvents
The
retention factor In chromatography, the retardation factor (''R'') is the fraction of an analyte
An analyte, component (in clinical chemistry), or chemical species is a substance or chemical constituent that is of interest in an analytical procedure. The purest ...
(R
''ƒ'') may be defined as the ratio of the distance travelled by the solute to the distance travelled by the solvent. It is used in chromatography to quantify the amount of retardation of a sample in a stationary phase relative to a mobile phase.
R
''ƒ'' values are usually expressed as a fraction of two decimal places.
* If R
''ƒ'' value of a solution is zero, the solute remains in the stationary phase and thus it is immobile.
* If R
''ƒ'' value = 1 then the solute has no affinity for the stationary phase and travels with the solvent front.
For example, if a compound travels 9.9 cm and the solvent front travels 12.7 cm, the R
''ƒ'' value = (9.9/12.7) = 0.779 or 0.78. R
''ƒ'' value depends on temperature and the solvent used in experiment, so several solvents offer several R
''ƒ'' values for the same mixture of compound. A solvent in chromatography is the liquid the paper is placed in, and the solute is the ink which is being separated.
Pigments and polarity
Paper chromatography is one method for testing the
purity
Purity may refer to:
Books
* ''Pureza'' (novel), a 1937 Brazilian novel by José Lins do Rego
* ''Purity'' (novel), a 2015 novel by Jonathan Franzen
** ''Purity'' (TV series), a TV series based on the novel
*''Purity'', a 2012 novel by Jackson P ...
of compounds and identifying substances. Paper chromatography is a useful technique because it is relatively quick and requires only small quantities of material. Separations in paper chromatography involve the principle of partition. In paper chromatography, substances are distributed between a stationary phase and a mobile phase. The stationary phase is the water trapped between the cellulose fibers of the paper. The mobile phase is a developing solution that travels up the stationary phase, carrying the samples with it. Components of the sample will separate readily according to how strongly they adsorb onto the stationary phase versus how readily they dissolve in the mobile phase.
When a colored
chemical sample
In general, a sample is a limited quantity of something which is intended to be similar to and represent a larger amount of that thing(s). The things could be countable objects such as individual items available as units for sale, or an uncou ...
is placed on a filter paper, the colors separate from the sample by placing one end of the paper in a
solvent
A solvent (s) (from the Latin '' solvō'', "loosen, untie, solve") is a substance that dissolves a solute, resulting in a solution. A solvent is usually a liquid but can also be a solid, a gas, or a supercritical fluid. Water is a solvent for ...
. The solvent
diffuse
Diffusion is the net movement of anything (for example, atoms, ions, molecules, energy) generally from a region of higher concentration to a region of lower concentration. Diffusion is driven by a gradient in Gibbs free energy or chemical p ...
s up the paper,
dissolving the various molecules in the sample according to the
polarities of the molecules and the solvent. If the sample contains more than one color, that means it must have more than one kind of molecule. Because of the different chemical structures of each kind of molecule, the chances are very high that each molecule will have at least a slightly different polarity, giving each molecule a different
solubility
In chemistry, solubility is the ability of a substance, the solute, to form a solution with another substance, the solvent. Insolubility is the opposite property, the inability of the solute to form such a solution.
The extent of the solubil ...
in the solvent. The unequal solubility causes the various color molecules to leave solution at different places as the solvent continues to move up the paper. The more soluble a molecule is, the higher it will migrate up the paper. If a chemical is very non-polar it will not dissolve at all in a very polar solvent. This is the same for a very polar chemical and a very non-polar solvent.
It is very important to note that when using water (a very polar substance) as a solvent, the more polar the color, the higher it will rise on the papers.
Types
![T](https://upload.wikimedia.org/wikipedia/commons/3/3d/T.baccata-paper_chromatography.jpg)
Descending
Development of the chromatogram is done by allowing the solvent to travel down the paper. Here, mobile phase is placed in solvent holder at the top. The spot is kept at the top and solvent flows down the paper from above.
Ascending
Here the solvent travels up the chromatographic paper. Both descending and ascending paper chromatography are used for the separation of organic and inorganic substances.
The sample and solvent move upward.
Ascending-descending
This is the hybrid of both of the above techniques. The upper part of ascending chromatography can be folded over a rod in order to allow the paper to become descending after crossing the rod.
Circular chromatography
A circular filter paper is taken and the sample is deposited at the center of the paper. After drying the spot, the filter paper is tied horizontally on a
Petri dish
A Petri dish (alternatively known as a Petri plate or cell-culture dish) is a shallow transparent lidded dish that biologists use to hold growth medium in which cells can be cultured,R. C. Dubey (2014): ''A Textbook Of Biotechnology For Class- ...
containing solvent, so that the wick of the paper is dipped in the solvent. The solvent rises through the wick and the components are separated into concentric rings.
Two-dimensional
In this technique a square or rectangular paper is used. Here the sample is applied to one of the corners and development is performed at a right angle to the direction of the first run.
History of paper chromatography
The discovery of paper chromatography in 1943 by
Martin Martin may refer to:
Places
* Martin City (disambiguation)
* Martin County (disambiguation)
* Martin Township (disambiguation)
Antarctica
* Martin Peninsula, Marie Byrd Land
* Port Martin, Adelie Land
* Point Martin, South Orkney Islands
Austr ...
and
Synge provided, for the first time, the means of surveying constituents of plants and for their separation and identification.
[ ]Erwin Chargaff
Erwin Chargaff (11 August 1905 – 20 June 2002) was an Austro-Hungarian-born American biochemist, writer, Bucovinian Jew who emigrated to the United States during the Nazi Germany, Nazi era, and professor of biochemistry at Columbia University ...
credits in Weintraub's history of the man the 1944 article by Consden, Gordon and Martin. There was an explosion of activity in this field after 1945.
References
Bibliography
*
{{DEFAULTSORT:Paper Chromatography
Chromatography