COUNTER CURRENT CHROMATOGRAPHY 1. BASIC PRINCIPLES 2. DROPLET CCC 3. CENTRIFUGAL PARTITION CCC 4. CHOICE OF SOLVENTS 5, MODE OF SEPARATION 6, APPLICATIONS

2. CONTENTS
• INTRODUCTION & DEFINITION
• PRINCIPLE
• PROCEDURE
• INSTRUMENTATION
• HYDROSTATIC CCC
• TYPES OF CCC (DCCC, CPCCC, HPCCC, EECCC)
• CHOICE OF SOLVENTS
• MODES OF SEPARATION
• APPLICATIONS

3. INTRODUCTION
& DEFINITION
• COIL PLANET CENTRIFUGE SUBJECTS
COILED TUBES TO PLANETARY MOTION
• THE COILED TUBES ARE FILLED WITH EQUAL
VOLUMES OF MUTUALLY IMMISCIBLE LIQUIDS
• THE PLANETARY MOTION PRODUCES
COUNTERCURRENTS OF THE 2 PHASES-
HYDROSTATIC COLUMN (STATIONERY PHASE),
HYDRODYNAMIC EQUILIBRIUM SYSTEMS
(MOBILE PHASE)

4. PRINCIPLE OF COUNTER COLUMN
CHROMATOGRAPHY

6. Stages of CCC-
1. Mixing
2. Settling
3. Separation
FACTORS AFFECTING CCC
1. Distribution constant- D
2. Stationery phase retention volume
3. Elution time
4. Resolution
5. Effect of column length

7. PROCEDURE Why low viscosity of mobile phase because-

8. INSTRUMENTATION
OF COUNTER
CURRENT
CHROMATOGRAPHY

9. WORKING PROCEDURE OF CCC

11. HYDRODYNAMIC V/S HYDROSTATIC CCC
• HYDRODYNAMIC CCC • HYDROSTATIC CCC

12. HYDRODYNAMIC V/S HYDROSTATIC CCC
HYDRODYNAMIC CCC/ CENTRIFUGAL PARTITION CHROMATOGRAPHY
• SOLUTE PARTITIONING IN A ROTATING COIL
• COIL IS ROTATED AROUND ITS HORIZONTAL AXIS.
• THIS MOTION CREATES AN ARCHIMEDEAN SCREW FORCEWHICHDRIVES
EVERY OBJECT WITH DIFFERENT DENSITIES TOWARDS THE END OF THE
COIL CALLED THE HEAD, THE OTHER END IS CALLED THE TAIL.
• UNDER THE SLOW ROTATION OF THE COIL, THE MOBILE PHASE INTRODUCED AT
THE HEAD OF THE COIL IS MIXED WITH THE STATIONERY PHASE TO ESTABLISH
A HYDRODYNAMIC EQUILIBRIUM WHERE A LARGE AMOUNT OF
THE STATIONERY PHASE IS PERMANENTLY RETAINED IN THE COIL WHILE IT
IS CONTINUOUSLY MIXED WITH THE MOBILE PHASE
HYDROSTATIC CCC/ DROPLET CCC
• USES A STATIONERY COILED TUBE
• THE COIL IS FIRST COMPLETELY FILLED WITH STATIONERY PHASE
WHICH CAN BE EITHER THE LIGHTER/ HEAVIER PHASE.
• THE OTHER PHASE IS THEN INTRODUCED INTO THE INLET OF THE COIL
WHERE IT PERCOLATES THROUGH THE STATIONERY PHASE
SEGMENTS ON ONE SIDE OF THE COIL.
• THE PROCESS CONTINUES UNTIL THE MOBILE PHASE ELUTES FROM
THE COIL.
• SOLUTES INTRODUCED LOCALLY AT THE INLET OF THE COIL
RE SUBJECTED.

13. HYDRODYNAMIC V/S HYDROSTATIC CCC

15. TYPES OF COUNTER
CURRENT
CHROMATOGRAPHY
Droplet CCC
Elution extrusion
CCC
Centrifugal
partition
chromatography
High speed CCC
DROPLET CCC
ELUTION
EXTRUSION CCC
CENTRIFUGAL
PARTITION CCC
HIGH SPEED
CCC

16. DROPLET CCC (DCCC)
• IT USES ONLY GRAVITY TO MOVE THE MOBILE PHASE THROUGH THE STATIONERY
PHASE
• IN DESCENDING MODE THE DROPLETS OF THE DENSER MOBILE PHASE & SAMPLE
ARE ALLOWED TO FALL THROUGH THE COLUMN OF THE LIGHTER STATIONER
PHASE USING ONLY GRAVITY.
• IF A LESS DENSE MOBILE PHASE IS USED IT WILL RISE THROUGH THE
STATIONERY PHASE, THEN IT IS CALLED ASCENDING MODE.
• THE ELUENT FROM ONE COLUMN IS TRANSFERRED FROM ONE COLUMN TO THE
OTHER, THE MORE THE COLUMNS ARE USED MORE THE NUMBER OF COLUMNS,
MORE THE THEORETICAL PLATES WILL BE FORMED.

17. CENTRIFUGAL PARTITION CCC (CPCCC)
• THIS METHOD USES CENTRIFUGAL FORCE TO SPEED SEPARATION &
ACHIEVE HIGHER FLOW RATES THAN DCCC.
• IT IS CONSTITUTED WITH THE A UNIQUE ROTOR COLUMN WHICH
ROTATES ON ITS CENTRAL AXIS.
• WITH LESS VIBRATIONS & NOISE, THE CPC OFFERS A WIDER RANGE
OF ROTATION SPEED RANGE (FROM 500 TO 2000 RPM)
• THIS ALLOWS A BETTER DECANTATION & RETENTION OF UNSTABLE &
BIPHASIC SYSTEM.
CPC ROTOR

18. HIGH SPEED CCC (HPCCC)
• HIGH-SPEED COUNTER-CURRENT CHROMATOGRAPHY (HSCCC) IS A TWO-PHASE SOLVENT SYSTEM,
WITHOUT SOLID PHASES, INSTEAD WITH LIQUID STATIONERY PHASE, TO RESOLVE TARGET COMPOUNDS
RELYING ON THE DIFFERENT PARTITIONING OF SOLUTES BETWEEN TWO IMMISCIBLE SOLVENTS WHICH
MAKES IT A VERY EFFECTIVE TOOL FOR THE PREPARATIVE SEPARATION AND PURIFICATION OF NATURAL
PRODUCTS
• MOST OF THE ACTIVE COMPOUNDS FROM CHINESE HERBS AND OTHER NATURAL PLANT PRODUCTS
EMPLOY THIS METHOD WIDELY

19. ELUTION- EXTRUSION CCC
• IN THIS METHOD, THE MOBILE PHASE IS EXTRUDED AFTER A CERTAIN POINT BY SWITCHING THE PHASE
BEING PUMPED INTO THE SYSTEM WHILE MAINTAINING ROTATION.
• FOR E.G.- IF THE SEPARATION HAS BEEN INITIATED WITH THE AQ. PHASE AS THE MOBILE PHASE, A CERTAIN
POINT THE ORGANIC PHASE IS PUMPED THROUGH THE COLUMN WHICH EFFECTIVELY PUSHES OUT BOTH
THE PHASES THAT ARE PRESENT IN THE COLUMN AT THE TIME OF SWITCHING.
• THE COMPLETE SAMPLE IS ELUTED IN THE ORDER OF POLARITY WITHOUT THE LOSS OF RESOLUTION BY
DIFFUSION.

20. CHOICE OF SOLVENTS

21. MODES OF SEPARATION

23. APPLICATIONS
OF CCC