A BRIEF REVIEW ON ION EXCHANGE CHROMATOGRAPHY

 

APPLICATIONS: Ion exchange chromatography can be applied for the separation and purification of many charged or ionisable molecules such as proteins, peptides, enzymes, nucleotides, DNA, antibiotics, vitamins and etc. from natural sources or synthetic origin [14]. Some of its applications are as follows:

1. Separation of similar ions: The ion exchange chromatography is used for separation of similar ions as different ions undergo exchange reactions to different extent [13-16]. E.g.: A mixture of H+, Na+, and K+ can be separated by using cation exchange resin. Similarly Cl-, Br-, I- can be separated by passing through basic anion exchanger.

Graph-1: Separation of potassium from sodium on Dowex 50 x 12 by elution with 0.6 M HCl

2. Softening of hard water: Hardness of water is due to the presences of Ca2+,Mg2+ and other divalent ions may be removed by passing the hard water through the cation exchanger charged with Na+ions. Then the following exchange reaction takes place:

TheCa+, Mg+ ions from water are retained on the column while Na + ions pass into the solution. These Na+ ions are harmless for washing purpose. After using the ion exchange for a time, it becomes in active. Percolating through it a concentrated solution of NaCl when the following reverse reaction takes place can revive its activity [12-16].

3. Complete demineralization of water: This requires complete removal of ions i.e., both cations and anions. For this, water is passed through an acidic cation exchanger then metallic cations are exchanged with H+ ions. The water obtained is then passed through a basic anion exchanger then the anions present in the water are exchanged by OH- of the exchanger. The H+and OH- ions which pass into solution combine to form unionized water [5, 12, and 14]. E.g.: Cation exchanger – Sulphonic acid resin is commonly used  Anion exchanger – Strong basic resin is used

4.Purification of organic compounds: Many natural products extracted in water have been found to contain ions originally present in water. Those ions can be removed by using ion exchange process [17].

5. Separation of sugars: This method is developed by Khym and Zill in 1951 [17]. Sugars are first converted into borate complex and the separation of borate complexes have been achieved quantitatively on columns by using ion exchange chromatography. In this, disaccharide can be separated from monosaccharide’s and the individual compounds of hexose, pentose from the mixture can be resolved [6, 12].

6. Separation of amino acids: Ion exchange methods can be used to separate the complex mixture of 18 amino acids obtained by the acid hydrolysis of proteins. The mixture of amino acids is first introduced on a very short column at pH 2 and eluted with 0.35 N sodium citrate buffers at pH 5.25. Acidic and neutral amino acids at first leave the column unseparated and after that other amino acids are separated [17]. Similarly, a mixture of vitamins like vitamin B1, B2, B6, Niacin, Folic acid, B12 etc. can be separated using ion exchange technique [16].

7. Purification and recovery of pharmaceuticals:
The process is used for purification and recovery of antibiotics, vitamins, alkaloids, hormones and other chemicals of pharmaceutical importance during their manufacturing process [17].

8.  Medicinal importance:
Anionic resins are introduced in the treatment of ulcer while cation exchangers have been used to remove Na+ from body during the treatment of hypertension and edema. The resins are also used as a diagnostic aid in gastric acidity tests. The resins have been successfully used with other medicinal agents to achieve delayed action dosages [17].

9.  Biochemical separations:
Used for biochemical separations like some drugs or metabolites from blood, urine or other biological fluids [9, 13].

10.  Ion exchange column in HPLC:
For separation of compounds of mixed nature like acidic and basic substances, ion exchange is used in HPLC [14].

11.  Concentration of ionic solutions:
A cation or anion from a bulk of solution can be adsorbed onto ion exchange resins, after adsorption, it can be eluted by using small volume of eluent [9].

12.  Separation of lanthanides:
Solution having mixture of lanthanides is passed through a column packed with particles of a suitable ion exchange resin. Cations present in solution undergo exchange with hydrogen with hydrogen ions [12, 16, and 17].

13.Separation of actinides:
The IEC technique has played a unique role in the discovery on the Trans plutonium elements in the actinide series. The power of the method can be judged from the order of elution of lanthanides (4f) and actinide (5f) ions in the +3 oxidation state from a cation exchange resin column with an aqueous solution of ammonium hydroxyl isobutyrate. In the actinide series also the elution occur in the reverse order of the atomic number due to actinide contraction and this proved that IEC is only way for identifying these elements [1, 8, 11, 16].

14. Removal of interfering radicals: The estimation of Ca+ or Ba+ ions is carried out by the oxalate or sulphate method in which phosphate ion is found to interfere. Therefore, its removal becomes necessary which is achieved by passing a solution of Ca+ or Ba+ ions through suitable ion exchanger in the column. The process has to be repeated so that the phosphate ions are completely removed. Now, the calcium and Ba+ ions held by resin will be removed by using suitable eluent. Finally, these ions are estimated by the usual methods [12, 17, and 18].

15.  Other applications:
* For the measurement of various active ingredients in medicinal formulations.
* For the measurement of drugs and their metabolites in serum and urine, for residue analysis in food raw materials.
* For the measurement of additives such as vitamins and preservatives in food and beverages

LIMITATIONS:
1)Column efficiency is less.
2)It is difficult to achieve control over selectivity and resolution.
3)Stability and reproducibility of the columns become questionable after repeated use.
4
)Nature of exchanging ions is not known.
5)There are substance, such as organic matter Fe3+occurring in some water which can foul the resin

CONCLUSION:
Several different types of liquid chromatography techniques are utilized for isolation of bioactive molecules from different sources. Follow-up of the nonsolvent extractable natural products can be realized by this technique. Consequently ion exchange chromatography, which has been used in the separation of ionic molecules for more than half a century is still used as an useful and popular method for isolation of natural products in modern drug discovery and it continue to expand with development of new technologies.

REFERENCES:
1.    B.K. Sharma; Instrumental Methods of Chemical Analysis; page. No: 123-160.
2.    Willard Merritt Dean Settle; Instrumental Methods of Analysis; 7th edition; page. No: 633-639.
3.    Kenneth A. Connors; A Text Book of Pharmaceutical Analysis; 3rd edition; page. No: 398 - 408.
4.    Amersham pharmacia biotech; Ion Exchange Chromatography: Principles and Methods.
5.    P. Parimoo; Pharmaceutical Analysis; page. No: 284 - 286.
6.    Harvard apparatus; Guide to Ion Exchange Chromatography.
7.    Tosoh Bioscience; Ion Exchange Chromatography.
8.    Yasser M. Moustafa and Rania E. Morsi; Ion Exchange Chromatography - An Overview.
9.    Dr. S. Ravi Sankar; Text book of pharmaceutical analysis; 4th edition; page. No: 16-2 – 16-10.
10.    ROBERT D. Braun; Introduction to Instrumental Analysis; page. No: 850-859.
11.    R. A. Day, A. L. Underwood; Quantitative Analysis; 6th edition; page. No: 535-543.
12.    J. Mendham, R.C Denney, J.D Barnes, M. Thomas, B. Siva Sankar; Text Book of Quantitative Chemical Analysis, 6th edition; page. No: 213-222.
13.    G. Vidya Sagar; A Text Book of Pharmaceutical Analysis; volume-II; page. No: 13 – 18.
14.    Anees A. Siddiqui; Pharmaceutical Analysis; volume-I;page. No: 216 – 236.
15.    Gurdeep R. Chatwal, Sham K. Anand; Instrumental Methods of Chemical Analysis; page. No: 2.662-2.672.
16.    Dr. A. V. Kasture, Dr. K. R. Mahadik, Dr. S. G. Wadodkar, Dr. H. N. More; Pharmaceutical Analysis; volume-II; page. No: 39 – 47.
17.    G. DevalaRao; A Text Book of Pharmaceutical Analysis; volume-II; page. No: 54 – 57.
18.    Mianbin Wu, Xuewan Wang, Zhengyu Zhang and Rutao Wang; Isolation and Purification of Bioactive Proteins from Bovine Colostrum.

 

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