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Counter current chromatography
Thiviaraj Palanisamy
1107181018
Introduction
• Counter-current chromatography is a liquid chromatography that uses two immiscible liquid phases i.e.., one act as stationary liquid and mobile liquid phase and no solid support.
• The inventor Yochiro Ito(1978) ,named it after the counter current partition method.
Definition
• Counter-current chromatography is a method of distribution of two (or) more substances by repeated distribution between two immiscible liquid phases that flow in opposite directions.
Principle
Principle
• The principle of separation involves the partition of a solute between two immiscible solvents (mobile phase and stationary phase).
• The relative proportion of solute passing into each of the two solvents determined by partition coefficient.
• Here both phases are liquid. Partition of solute in two phases is based on differences in capacity factor, k and distribution coefficient ,kd of the analyte.
Stages
1. Mixing
2. Settling
3. Separation
Stages
Importance
1. Distribution constant D
2. Stationary phase retention volume
3. Elution time
4.Resolution
5.Effect of column length
Procedure
Content
• The best results are obtained by filtering the sample solution before introduction into the column.
• The whole ccc is a chromatographic column
• centrifugal fields imply rotating parts, rotors, gears, spools, motors and speed regulators.
• Mixing of phases is done so that the analyte can move between the phases.
• Mobile phase is mixing with then settling from stationary phase throughout the column.
• Maximum retention of stationary phase is obtained by using less viscous phase as mobile phase.
Instrumentation
INSTRUMENTATION
Modes of operation
1. Head to tail
2. Tail to head
3. Dual-mode
4. Gradient mode
5. pH zone refining
Types of
CCC
Hydrodynamic CCC counter
Hydrostatic CCC counter
Types of CCC
Comparison
Types
Types of CCC
1. Droplet counter current chromatography.
2. Elution Extrusion Counter Current Chromatography
3. Centrifugal Partition Chromatography
4. High Speed Counter Current Chromatography.
1.
Droplet counter current chromatography.
1. It uses only gravity to move the mobile phase through the stationary phase.
2. In descending mode, droplets of the denser mobile phase and sample are allowed to fall through a column of the lighter stationary phase using only gravity.
3. If a less dense mobile phase is used it will rise through the stationary phase, this is called ascending mode.
4. The eluent from one column is transferred to another; the more columns that are used, the more theoretical plates can be achieved.
2.
Elution Extrusion Counter Current Chromatography
1. When run reaches a certain point, mobile phase is stopped & stationary phase is pumped in to extrude the column content.
2. Elution extrusion makes use of the fact that compounds may be fully separated inside the column before eluting from it. Due to the fact we are using a liquid SP, we are able to recover the separated compounds without completing the full elution cycle.
3. In elution extrusion, the separation is started in the same manner as in single-mode CCC.
4. However, when the run reaches a certain point (e.g. D=1.0), MP will be stopped and stationary phase pumped in to extrude the column contents (i.e. the phase initially used as SP will be introduced in the instrument as a new MP).
Centrifugal Partition Chromatography
3.
1. CPC use CENTRIFUGAL FORCE to speed separation and achieves higher flow rates than DCCC.
2. The centrifugal partition chromatograph is constituted with a unique rotor (column). This rotor rotates on its central axis.
3. With less vibrations and noise, the CPC offers a wider rotation speed range (from 500 to 2000 rpm) than HSCCC.
4. That allows a better decantation and retention for unstable biphasic system (e.g., aqueous aqueous systems or Butanol/water systems). Basics of CPC: The CPC rotor is constituted by the superposition of disks engraved with small cells connected by head / tail ducts.
5. These are the cells, where actual separation takes place.
6. Rotor is filled with the stationary phase, which stays inside the rotor, while mobile phase is pumped through
High-Speed countercurrent chromatography
4
- 1. High-Speed Countercurrent Chromatography (HSCCC) instruments are a modern and popular type of Countercurrent Chromatography (CCC) apparatus. These machines consist of:
- Helically coiled inert tubing (Usually Teflon or sometimes stainless steel) that rotates on its own axis.
- A gear assembly arranged such that the helical coils revolve around a central axis to achieve planetary motion.
Advantages
+
1. It is simple, rapid, and reproducible.
2. High sensitivity.
3. High performance.
4. Rapid process and hence time saving.
5. It is having a high resolution and separation capacity
6. Accuracy and Precision.
7. Stationary phase was chemically innert.
8. Wide varities of stationary phase.
9. Mobile phase was chemically innert.
10. Less requirement of mobile phase in developing chamber.
11. Early recovery of separated component.
12. Easy visualization of separated components.
13. It is having Good reproducibility and repeatability.
14. It is analytical technique is important for validation of product, quality control studies of product.
15. It is important for qualitative and quantitative analysis.
Applications
1.Analysis of plant and different natural products.
2.Food analysis.
3.Environment analysis.
4.Inorganic ions
5.Lanthanides and actinides
6.Alkali and alkaline-earth elements
7.Platinum group metals
8.Various other elements
9.Purification of salt solutions by CCC
10. Recovery of PAHs from liquid sewage sludge and soil
References
1. Conway WD (1990) Countercurrent Chromatography, Apparatus Theory and Applications. New York: VCH
2. Conway WD and Petroski RJ (eds) (1995) Modern Countercurrent Chromatography. ACS Symposium Series 593. Washington, DC: American Chemical Society
3. Conway WD, Bachert EL, Sarlo AM and Chan CW (1998) Comparison of countercurrent chromatography with Sash chromatography. Journal of Liquid Chromatography and Related Technologies 21: 53}63.
4. Counter Current Chromatography: A Superior and An Advanced Analytical Technique by Naman B. Doshi, Avani H. Sheth, Dr Dhrubo Jyoti Sen and Dr J B Dave Pharma Times - Vol 42 - No. 01 - January 2010.
5. www.dynamicextractions.com
6. Berthod, Alain; Maryutina, Tatyana; Spivakov, Boris; Shpigun, Oleg; Sutherland, Ian A. (2009). “Countercurrent chromatography in analytical chemistry (IUPAC Technical Report)". Pure and Applied Chemistry 81 (2): 355–387. doi:10.1351/PAC-REP-08-06- 05. ISSN 0033- 4545.
7. Ito, Y.; Bowman, RL (1970). “Countercurrent Chromatography: Liquid-Liquid Partition Chromatography without Solid Support”. Science 167 (3916): 281–283. Bibcode:1970Sci...167..281I. doi:10.1126/science.167.3916.281. PMID 5409709.
8. Foucault, Alain P. (1994). Centrifugal Partition Chromatography. Chromatographic Science Series, Vol. 68.CRC Press. ISBN 978-0824792572.
9. Ito, Yoichiro (2005). “Golden rules and pitfalls in selecting optimum conditions for high-speed counter-current chromatography”. Journal of Chromatography A 1065 (2): 145–168. doi:10.1016/j.chroma.2004.12.044. ISSN 0021-9673.
10. Ian A. Sutherland (2007). “Recent progress on the industrial scale-up of counter-current chromatography” Journal of Chromatography A 1151: 6–13.