Column+Chromatography+-+Danielle+Hinckley

=Edited by Christie Cockayne 10/12/2014 = =Column Chromatography =

**Column Chromatography Background: **
Column chromatography is a technique used to separate proteins, in which a mixture of proteins in solution is passed through a column containing a porous solid matrix. There are two separate phases to the column chromatography, mobile and stationary. The stationary phase is usually a solid material that the samples interact with (ex. silica gel and alumina). The mobile phase flows over the stationary phase and carries along with it the sample to be separated.

Column chromatography can separate proteins based on charge, hydrophobicity, their size, or their ability to bind to particular molecules. An ion-exchange column is a column that is packed full of positively or negatively charged beads. Hydrophobic column chromatography consists of columns that are packed with beads that have hydrophobic side chains protruding to help slow down the hydrophobic regions. Gel-Filtration column chromatography

consists of columns that are packed with tiny porous beads to help separate the proteins by their size.

A visual example of separation via column chromatography

**Column Chromatography Technique: **
An example set up of lab equipment before a column chromatography purification. Zubrick, James W. The Organic Chem Lab Survival Manual. 7th ed. New York: Wiley, 2008. Print.

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Visual representation of a column chromatography set-up and results. http://becuo.com/column-chromatography-protein-purification

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The first true chromatography experiment is attributed to Mikhail Tsvet. The method was described by him on December 30, 1901. He extracted plant pigments from leaves with ether and alcohol and running the solution through a column containing calcium carbonate. =====

 Mikhail Tsvet, credited with the first chromatography experiment in 1901.

**Current Research Using Column Chromatography: **
As column chromatography is an established purification technique in cellular biology, as well as many other natural science fields, new research commonly uses this technique in recent publications.

The University of Olso in Norway has just completed cell biology research using this technique. Column chromatography was used to isolate different proteins from small cell samples from tumor tissues (Hustoft et al., 2014). These proteins were then isolated from the columns for further research. This research can benefit the general public as it can increase productivity in other tumor cell research studies.

Various separation methods have been used in amino acid analysis in past research. Column chromatography prevailed as the best separation technique in amino acid analysis (Rigas, 2013). This is an important discovery in cell biology as it can be used to improve future amino acid research and help speed up the separation process in current experiments.

Column chromatography can be used in the laboratory as a part of a separation using multiple techniques. While some research experiments may have conflicting data between separation methods (Zheng et al., 2014), an adjustment in column chromatography is simple and can eliminate these discrepancies. Since column chromatography is a simple but efficient technique, it can eliminate waste during other separation methods and optimize productivity.

**Bibliography:**

Hustoft, H., Vehus, T., Brandtzaeg, O., Krauss, S., Greibrokk, T., Wilson, S., & Lundanes, E. (2014). Open Tubular Lab-On-Column/Mass Spectrometry for Targeted Proteomics of Nanogram Sample Amounts. //Plos ONE//, //9//(9), 1-10. doi:10.1371/journal.pone.0106881

Rigas, P. (2013). Post-column labeling techniques in amino acid analysis by liquid chromatography. //Analytical & Bioanalytical Chemistry//, //405//(25), 7957-7992. doi:10.1007/s00216-013-7127-3

Zheng, S., Ma, Z., Han, H., Ye, J., Wang, R., Cai, S., & ... Jiang, H. (2014). Post-column mobile phase adjustment: A strategy to eliminate the contradiction between liquid chromatography and mass spectrometry in the determination of flavonoids in rat plasma. //Journal Of Pharmaceutical & Biomedical Analysis//, //95//176-183. doi:10.1016/j.jpba.2014.02.024