Principle and Procedure of Paper chromatography

What is Paper Chromatography? Paper chromatography is one of the kinds of processes that operate on a piece of special paper. It is a method of planar chromatography in which a cellulose filter paper works as a stationary phase in which the compounds are separated. Paper chromatography is the most effective technique of separation, particularly in the analysis of chemical compounds and lipid samples. Several types of chromatography, such as liquid chromatography, ion-exchange chromatography, gas chromatography, thin-layer chromatography (TLC), column chromatography and affinity chromatography have the same basic principles.

Paper Chromatography Principle: In paper chromatography, the adsorption chromatography or partition chromatography is involved in which components are divided or distributed between liquid phases. In this, a special chromatography paper is used in which water is trapped which is the stationary phase. A sample solution is spotted on the base of the page. In the appropriate solvent, the paper strip is suspended and the mobile phase rises through capillary action to the top of the paper. The analytes are selectively retained on the stationary phase (cellulose paper). The spots of different analytes move to different heights along the mobile phase and each component in the sample mixture separates based on varying degrees of adhesion.

Experimental Procedure of Paper ChromatographyChoose an appropriate type of development: It’s decided based on the solvent, mixture, paper complexity, etc. As they are easy to execute, radial type or ascending paper chromatography is usually employed. It's also simple to handle, the chromatogram obtained is quicker.Selecting an appropriate filter paper: Based on a sample quality and pores size, the filter paper is selected.Sample preparation: Sample preparation involves dissolving the sample in an appropriate solvent that is used in the process of mobile phase making.Sample spot on paper: The sample mixture should be spotted in a proper position on the paper using a capillary tube.Development of chromatogram:The development of chromatograms is spotted by immersing the stationary phase (Paper) into the mobile phase. Because of the paper's capillary action, the mobile process passes over the paper sample.Paper drying and compound detection: Upon the development of the chromatogram, the paper is dried by an air dryer. Detecting solution also is sprayed on developed paper and dried to recognize the spots of the sample chromatogram. Paper with varying bands of analytes is observed under UV-light and further Rf values are calculated.

Paper Chromatography Applications

  • Paper chromatography is used for separating polar and non-polar compounds.
  • It is used for the detection and separation of color mixtures including pigments as a qualitative analytical chemistry technique.
  • Paper chromatography is used for identifying sugars, nucleic acids, lipids, amino acids, and other biomolecules.
  • It is used in DNA/RNA sequencing.
  • It is used to test pharmaceutical purity.
  • It is used for the detection of pollutants in food and beverages.
  • For investigations and criminal trials, paper chromatography is used in forensic research.
  • This is used for ripening and fermentation study.
  • It is used for the analysis of cosmetics.
  • Paper chromatography is used for component separation and purification.
  • Paper chromatography can be applied to monitor chemical synthesis reactions.

Types of paper chromatography

Ascending Paper Chromatography: As the name suggests the developing solvent or mobile phase travels in an upward direction

Descending Paper Chromatography: In such paper chromatography, due to the gravitational pull and capillary action the direction of the solvent flow is downward

Ascending-Descending Paper Chromatography: This type of paper chromatography the solvent first moves up on the paper folded over a rod and after crossing the rod it continues its path downwards.

Horizontal or Circular Paper Chromatography: In this method, the sample mixture is employed in the center of a round paper placed on a flat surface.

Two Dimensional Paper Chromatography: With the aid of two-dimensional paper chromatography, substances that have the same Rf values can be solved.

The Advantages of Paper Chromatography are as follows.

  • Compounds are separated within a short time.
  • Analysis of the paper chromatography required small sample quantities.
  • It is an inexpensive technique compared to other chromatographic methods.
  • It is possible to identify both organic and inorganic compounds.
  • Paper chromatography is easy to use and setup.

The Disadvantages of Paper Chromatography are as follows.

  • The use of paper chromatography techniques cannot separate volatile substances.
  • Paper chromatography cannot separate large sample volumes.
  • The paper chromatography has less accuracy compared to TLC, HPTLC, and HPLC.
  • It is unable to separate the complex mixture.
  • It cannot save results for long periods.

The questions commonly asked about Paper Chromatography are as follows.

What is the basic principle of paper chromatography? The principle involved is partition chromatography in which the compounds are distributed or partitioned into liquid phases.

What is paper chromatography? Paper chromatography is a method of separating and identifying mixtures, especially pigments.

What is the major advantage of paper chromatography? The main advantages of paper chromatography are simplicity, rapid separation, low cost and have good sensitivity.

What type of paper is used in paper chromatography? Filter paper, chemically modified papers, acetylated papers, and pure cellulose papers of different grades like No.1, 2, 3, 4, 7, 17, 20, 40, 42, etc are used as stationary phases in paper chromatography.

What is the major difference between thin layer chromatography and paper chromatography? The major difference between TLC and paper chromatography is that the stationary phase of the paper chromatography is a cellulose paper while TLC uses a thin layer of silica or alumina supported on a flat surface.

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