ion-exchange+chromatography

Ion-exchange chromatography is a type of column chromatography, in which molecules are separated by their net charge. An insoluble matrix (such as DEAE-cellulose, CM-cellulose, or phosphocellulose, depending on the desired charge of the matrix) is placed into a column. The protein sample is loaded at the top, and then a selected solvent is continuously applied above it with increasing pH to create a gradient. [1] 
 * Alison Taylor **
 * Description **

The speed at which the different types of molecules in the sample elute through the matrix depends on the magnitude of the association between the matrix and the dissolved molecules in the sample, which is dependent on their individual ionic strengths as well as the pH of the solvent being used to separate the molecules in solution. Since the matrix itself is charged, molecules with the opposite charge will bind to it while those with like charges will move through immediately.

Ion-exchange chromatography provides information about the proteins in a sample. A chromatogram is produced, which shows elution time in relation to absorbance data. The number of peaks on the chromatogram indicates how many proteins there are in the sample, and comparing the areas of the peaks can provide information about which proteins are present.
 * Purpose **

Modern ion chromatography was first developed by H. Small, T.S. Stevens, and W.C. Bauman in 1975. Their method involved the use of suppressed conductivity detection to separate positively- and negatively-charged ions. Non-suppressed conductivity detection was used for anion chromatography by Gjerde et al. in 1979, and for cation chromatography in 1980. However, the development of the technique began in 1947, when displacement ion-exchange chromatography was used in order to separate rare earth metals by Spedding and Powel at Iowa State University. [2]
 * Origin and History**

Grein et. al. (2012) used ion-exchange chromatography to process viral vectors used to deliver genes into mammalian cells. Ionic conductivity was used in order to control product purity, yield, and infectivity of a recombinant baculovirus of //Autographa californica//. These factors could be influenced in part by manipulating different factors during processing, including the membrane used in ion-exchange membrane chromatography. [3]
 * Recent Applications in Research**

Knoll and Seubert (2012) used this technique in environmental research, measuring traces of EDTA, DTPA, and CDTA in surface water by running a water sample with either Fe3+ or In3+ through an ion-exchange chromatography column. They tested the viability of the method for NTA as well, but the complexes formed with it after the addition of the metal did not last long enough for the accurate separation during the process. Detecting these aminopolycarboxylic acid-based chelating agents is useful, since these are chemicals that are not taken out of the water during wastewater treatment, but are used in various industrial processes. [4]

Palmeira-de-Oliveira et al. (2011) used ion-exchange chromatography to test yeasts that had been showing resistance to a potential antifungal medication. The surface charge densities in sixteen //Candida// species were analyzed and related to their sensitivity to chitosan, a biopolymer with antifungal properties that may be useful for the treatment of //Candida// infections in the future. The study found that the less sensitive species had a lower affinity for the anionic resin, while the more sensitive species had a very high affinity. This trend shows that chitosan activity is probably a result of an ionic reaction with the cell surface. [5]

Lee et. al (2014) used multiple techniques to analyze and extract recombinant measles virus nucleoproteins (rMeV N) proteins, and they also used ion exchange chromatography to purify the proteins produced, because it is efficient and reliable, and thus resulted in a high-purity rMeV N protein. When compared to naïve nucleoproteins, the rMeV N protein purified by ion exchange chromatography had similar feature. Therefore, the purification methods can be applied to the large-scale production of rMeV N proteins, which are essential for the development of new diagnostic tools and vaccines for acute and chronic measles virus (MeV) infections. [6]

Terefe et. al (2014) suggest a s new method to be utilized. Stimuli responsive polymers (SRPs) with ion exchange functional groups can be used to selectively capture and release charged molecules from a complex mixture using physical stimuli to trigger conformational transitions in the polymer. The structural change of the polymers in response to a stimulus may lead to reduced ligand target molecule interaction resulting in the release of the captured molecule without the use of chemical reagents. This could result in a smaller environmental impact. [7]

pH gradients in ion exchange columns is only rarely applied due to the difficulties to form controllable, linear pH gradients over a broad pH range. Recently, Kroner and Hubbuch (2013) developed a method for the systematic generation of buffer compositions with linear titration curves, resulting in well controllable pH gradients. An in silico method was successfully developed, and this tool, buffer compositions for pH gradient ion exchange chromatography with pH ranges spanning up to 7.5 pH units were established and successfully validated. [8]

> and Francis group, NY, pp. 481-484.
 * References**
 * 1) Alberts, B., Johnson, A., Lewis, J., Raff, M., Roberts, K., & Walter, P. 2002. //Molecular Biology of the Cell//, 4th ed. Garland Science, Taylor
 * 1) Fritz, J.S. (2004). Early milestones in the development of ion-exchange chromatography: a personal account. //Journal// //of Chromatography A//. 1039 (1-2), 3-12.
 * 2) Grein, T.A., Michalsky, R., Lopez, M.V., & Czermak, P. (2012). Purification of a recombinant baculovirus of Autographa californica M nucleopolyhedrovirus by ion exchange membrane chromatography. //Journal// //of Virological Methods//. 183(2): 117-124.
 * 3) Knoll, J. & Seubert, A. (2012). Indirect ultra trace determination of aminopolycarboxylic acids in surface water using ion-exchange chromatography coupled on-line to inductively coupled plasma mass spectrometry//. Journal of Chromatography A//. 1270: 219-224.
 * 4) Palmeira-de-Oliveira, A., Passarinha, L.A., Gaspar, C., Palmeira-de-Oliveira, R., Sarmento, B., Martinez-de-Oliveira, J., Pina-Vaz, C., Rodrigues, A.G., & Queiroz, J.A. (2011). The relationship between Candida species charge density and chitosan activity evaluated by ion-exchange chromatography. //Journal of Chromatography// //B//. 879(31): 3749-3751
 * 5) Lee, H. S., Kim, Y.-J., Yang, J., Yoon, H. S., Kim, S. T., & Kim, K. (2014). Alternative purification method for recombinant measles viral nucleoprotein expressed in insect cells by ion-exchange chromatography. //Journal of Virological Methods//, //197//, 55-62.
 * 6) Terefe, N. S., Glagovskaia, O., De Silva, K., & Stockmann, R. (2014). Application of stimuli responsive polymers for sustainable ion exchange chromatography. //Food and Bioproducts Processing//, (92), 208-225.
 * 7) Kroner, F., & Hubbuch, J. (2013). Systematic generation of buffer systems for pH gradient ion exchange chromatography and their application. Journal of Chromatography A, 78-87