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Ion exchange chromatography

Ion chromatography workstation
Acronym IC
Classification Chromatography
Manufacturers Dionex Corp., Lachat Instruments, Metrohm, Ltd.
Other Techniques
Related High performance liquid chromatography
Aqueous Normal Phase Chromatography
Size exclusion chromatography
Micellar liquid chromatography
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Ion-exchange chromatography (or ion chromatography) is a process that allows the separation of ions and polar molecules based on the charge properties of the molecules. Chromatography (from Greek χρώμα chroma, color and γραφειν"graphein" to write is the collective term for a family of Laboratory High-performance liquid chromatography (or High pressure liquid chromatography, HPLC) is a form of Column chromatography used frequently in Biochemistry Aqueous normal phase chromatography (ANP is a chromatographic technique which encompasses the Mobile phase region between reversed-phase chromatography (RP and organic Size exclusion chromatography (SEC is a chromatographic method in which particles are separated based on their size or in more technical terms their Hydrodynamic volume Micellar liquid chromatography (MLC is a form of reversed phase liquid chromatography that uses an Aqueous micellar solutions as the mobile phase An ion is an Atom or Molecule which has lost or gained one or more Valence electrons giving it a positive or negative electrical charge "Polar molecule" and "Non-polar" redirect here It can be used for almost any kind of charged molecule including large proteins, small nucleotides and amino acids. Proteins are large Organic compounds made of Amino acids arranged in a linear chain and joined together by Peptide bonds between the Carboxyl Nucleotides are Organic compounds that consist of three joined structures a nitrogenous base a Sugar, and a Phosphate group In Chemistry, an amino acid is a Molecule containing both Amine and Carboxyl Functional groups In Biochemistry, this The solution to be injected is usually called a sample, and the individually separated components are called analytes. It is often used in protein purification, water analysis, and quality control.

Contents

History

Ion methods have been in use since 1850, when H. Thompson and J. T. Way, researchers in England, treated various clays with ammonium sulfate or carbonate in solution to extract the ammonia and release calcium. In 1927, the first zeolite mineral column was used to remove interfering calcium and magnesium ions from solution to determine the sulfate content of water. The modern version of IEC was developed during the wartime Manhattan Project. A technique was required to separate and concentrate the radioactive elements needed to make the atom bomb. Researchers chose adsorbents that would latch onto charged transuranium elements, which could then be differentially eluted. Ultimately, once declassified, these techniques would use new IE resins to develop the systems that are often used today for specific purification of biologicals and inorganics. In the early 1970s, ion chromatography was developed by Hamish Small and co-workers at Dow Chemical Company as a novel method of IEC usable in automated analysis. IC uses weaker ionic resins for its stationary phase and an additional neutralizing stripper, or suppressor, column to remove background eluent ions. It is a powerful technique for determining low concentrations of ions and is especially useful in environmental and water quality studies, among other applications.

The Dow Chemical Company technology was acquired by Durrum Instrument Corp. The Dow Chemical Company () is an American Multinational corporation headquartered in Midland Michigan. (maker of the Durrum D-500), which later formed a separate business unit for its new IC products, naming it Dionex (Dow Ion Exchange). The D-500 Amino Acid Analyzer was designed and built by Durrum Instruments in the late 1960s Dionex Corporation was incorporated in Sunnyvale, California in 1980, and, led by A. Blaine Bowman, purchased the Dionex assets.

Principle

Ion Chromatogram
Ion Chromatogram

Ion exchange chromatography retains analyte molecules based on coulombic (ionic) interactions. ---- Bold text Coulomb's law', developed in the 1780s by French physicist Charles Augustin de Coulomb, may be stated in scalar form The stationary phase surface displays ionic functional groups (R-X) that interact with analyte ions of opposite charge. This type of chromatography is further subdivided into cation exchange chromatography and anion exchange chromatography. The ionic compound consisting of the cationic species M+ and the anionic species can be retained by the stationary phase.

Cation exchange chromatography retains positively charged cations because the stationary phase displays a negatively charged functional group:

\text{R-X}^-\text{C}^+\,+\, \text{M}^+ \, \text{B}^- \rightleftarrows \,\text{R-X}^-\text{M}^+ \,+\, \text{C}^+ \,+\, \text{B}^-

Anion exchange chromatography retains anions using positively charged functional group:

\text{R-X}^+\text{A}^-\,+\, \text{M}^+ \, \text{B}^- \rightleftarrows \,\text{R-X}^+\text{B}^- \,+\, \text{M}^+ \,+\, \text{A}^-

Note that the ion strength of either C+ or A- in the mobile phase can be adjusted to shift the equilibrium position and thus retention time. An ion is an Atom or Molecule which has lost or gained one or more Valence electrons giving it a positive or negative electrical charge

The ion chromatogram shows a typical chromatogram obtained with an anion exchange column.

Separating Proteins

Proteins have numerous functional groups that can have both positive and negative charges. Ion exchange chromatography separates proteins according to their net charge, which is dependent on the composition of the mobile phase. By adjusting the pH or the ionic concentration of the mobile phase, various protein molecules can be separated. For example, if a protein has a net positive charge at pH 7, then it will bind to a column of negatively-charged beads, whereas a negatively charged protein would not. By changing the pH so that the net charge on the protein is negative, it too will be eluted.

Elution by changing the ionic strength of the mobile phase is a more subtle effect - it works as ion from the mobile phase will interact with the immobilized ion in preference over those on the stationary phase. This "shields" the stationary phase from the protein, (and vice versa) and allows the protein to elute.

Typical Technique

Another Ion Chromatography Workstation
Another Ion Chromatography Workstation

A sample is introduced, either manually or with an autosampler, into a sample loop of known volume. A buffered aqueous solution known as the mobile phase carries the sample from the loop onto a column that contains some form of stationary phase material. For an individual weak acid or weak base component see Buffering agent. This is typically a resin or gel matrix consisting of agarose or cellulose beads with covalently bonded charged functional groups. Agar or agar agar is a Gelatinous substance derived from Seaweed. Cellulose is an Organic compound with the formula, a Polysaccharide consisting of a linear chain of several hundred to over ten thousand β(1→4 The target analytes (anions or cations) are retained on the stationary phase but can be eluted by increasing the concentration of a similarly charged species that will displace the analyte ions from the stationary phase. For example, in cation exchange chromatography, the positively charged analyte could be displaced by the addition of positively charged sodium ions. The analytes of interest must then be detected by some means, typically by conductivity or UV/Visible light absorbance.

In order to control an IC system, a Chromatography Data System (CDS) is usually needed. In addition to IC systems, some of these CDSs can also control Gas Chromatography (GC) and HPLC systems.

Manufacturers of Ion Chromatographs

Manufacturers of Ion Chromatography Accessories and Columns

References

See also

The isoelectric point (pI is the PH at which a particular Molecule or surface carries no net electrical charge. High-performance liquid chromatography (or High pressure liquid chromatography, HPLC) is a form of Column chromatography used frequently in Biochemistry
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