Ion Exchange chromatography-Modern Pharmaceutical Analytical Techniques.pptx

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1. Ion exchange chromatography Subject:- Modern Pharmaceutical Analytical Techniques Subject Code:- MPC 101T Presented to - Dr. Junmoni Kalita Associate Professor Department of Pharmaceutical Chemistry Presented by- Seemanta Nayan Das M.Pharm 1 st semester Roll No.- 2412023009 Department of Pharmaceutical Chemistry 1

2. Contents Introduction (Definition and Principle). Mechanism. Instrumentation. Selection of suitable ionic systems. Chromatographic parameters. Factors. Isolation procedure. Data interpretation. Applications. Bibliography.

3. Definition Ion exchange may be defined as a reversible reaction in which free mobile ions of a solid called ion exchange are exchanged for different ions of similar charge present in solution. Properties of ion exchangers are as follows: Insoluble in water and organic solvents like benzene, CCl 4 , ether. Complex in nature. Presence of active or counter ions which exchanged with other ions.

4. Principle Ion-exchange chromatography undergoes separation of differently charged or ionizable compounds comprises from mobile and stationary phases. Mobile phases Aqueous buffer system Stationary phase Inert organic matrix(positive and negative ions. The two phases produces anion and cation exchangers which are referred to as counter ion.

5. Mechanism Diffusion of the ion to the exchanger surface. Diffusion of the ion through the matrix structure of the exchanger to exchanger site. Exchange of ions at the exchange site. Cation exchanger: Resin:- Sulphonic , carboxylic, phenolic groups Cations :- Na + , Ca 2+ Trade name Functional groups Framework material Amberlite IR-120 -SO 3 H Styrene/ divinylbenzene Amberlite IRC-50 -COOH Methacrylic acid/ divinyl SE cellulose -C 2 H 4 , -SO 3 H Cellulose

6. Mechanism Anion exchanger: Resin:- Amine or quaternary ammonium groups. Anions:- Cl - , SO 4 2- , OH - Diffusion of the exchanged ion through the exchanger to the surface. Selective desorption by the eluent and diffusion of the molecule into the external solution. Trade name Functional groups Framework material Amberlite IRA-400 -CH 2 -N + (CH 3 ) 3 Styrene/ divinyl benzene Zerolit FF-IP -CH 2 -N + (CH 3 ) 3 Styrene/ divinyl benzene QAE Sephadex A-25 Dextran

7. Instrumentation The components of an ion-exchange chromatography are as follows :- A high pressure pump with pressure and flow indicator, to deliver the eluent . An injector for introducing the sample into the eluent stream and onto the column. A column is used to separate the sample mixture into the individual components . A detector, to measure the analyte peaks as eluent from the column. A data system for collecting and organizing the chromatograms and data.

8. Selection of suitable ionic systems Ion exchangers are classified according to the pH values of their ion groups:- pH study Dissociated form of material. One component retained by the resin. Other component does not interact with the ion exchanger. Specification of ion exchanger Ionic groups Suitable chromatographic column Strong acid -SO 3 - Acidic and alkaline Weak acid -COO - Only alkaline Strong base -N + R 3 Acidic and alkaline Weak base -NH R 2 Only acidic

9. Chromatographic parameters Ion exchange capacity:- It is defined as the number of ion active groups which is referred to as total ion exchange capacity. Expressed in milliequivalents per gram of ion exchange resin. Known concentration of NaCl Known weight of resin and acid eluting from column is collected Titrated with standard solution of alkali The capacity is given by:- Passed through a column V = Volume of alkali solution N = milliequivalents per litre W = weight of resin in gm

10. Factors affecting ion exchange chromatography Affects the charge state IS Elution of bound ions Affects kinetics and capacity Smaller resin better resolution. Higher concentration saturate resin quickly Intermediate

11. Isolation procedure Source:- https://images.app.goo.gl/7qQxnGTGEyB1JyXF9 Ricardo M. Silva, Sara Sousa Rosa, Raquel Cunha, Cláudia L. da Silva, Ana M. Azevedo , Ana Fernandes - Platzgummer , “Anion exchange chromatography-based platform for the scalable purification of extracellular vesicles derived from human mesenchymal stromal cells”, Separation and Purification Technology, Volume 310, 2023, 123238, ISSN 1383-5866, https://doi.org/10.1016/j.seppur.2023.123238 .

12. Data interpretation Source: https://chem.libretexts.org/Courses/University_of_Arkansas_Little_Rock/CHEM_4320_5320%3A_Biochemistry_1/03%3A_Methods_of_Protein_Purification_and_Characterization/3.4%3A_Chromatography/3.4.3._Ion_Exchange_Chromatography

13. Applications Separation of similar ions: A mixture of Li + , Na + and K + ion can be separated by passing their solution through cation exchanger. 0.1 N HCl is used as an eluent . Softening of hard water: These sodium ions are harmless for washing purposes.

14. Applications 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. Separation of sugars:

15. Applications 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. Separation of lanthanides: Lanthanides Column packed with ion exchange resin Lanthanides separation Ion exchange column in HPLC: For separation of compounds of mixed nature like acidic and basic substances, ion exchange is used in HPLC.

16. Bibliography Chatwal , G. R., & Anand , S. K. (2014). Instrumental Methods of Chemical Analysis (5th edition). Himalaya Publishing House Pvt. Ltd. Page No.- 2.662-2.672 Martin, Dean. (2013). Column Chromatography || Ion-Exchange Chromatography and Its Applications. , 10.5772/47823(Chapter 2), doi:10.5772/55744. Mohammed JS; A brief review on Ion Exchange Chromatography; PharmaTutor ; 2017; 5(2); 30-38

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