The document discusses gel permeation chromatography (GPC), which separates molecules by size as they pass through a column of porous beads. It describes the basic components and working principles of GPC, including how larger molecules have shorter residence times in the column than smaller molecules. Applications include determining relative molecular weight and molecular weight distribution of polymer samples, and separating substances like sugars, polypeptides, proteins, and polymers.

1. GEL PERMEATION CHROMATOGRAPHY
BY : VISHNU PRAJAPATI
SUB : CHEMISTRY...

2. CONTENTS
• AIM
• INTRODUCTION
• WORKING PRINCIPLE OF INSTRUMENT
• MATERIALS AND METHOD FOR SAMPLE ANALYSIS
• APPLICATIONS

3. AIM :
TO STUDY ABOUT GEL PERMEATION CHROMATOGRAPHY.

4. INTRODUCTION
• CHROMATOGRAPHY IS A PHYSICAL METHOD OF SEPARATION IN WHICH
THE COMPONENTS TO BE SEPARATED ARE DISTRIBUTED BETWEEN TWO
PHASES, ONE OF WHICH IS STATIONARY (STATIONARY PHASE) WHILE
THE OTHER (THE MOBILE PHASE) MOVES IN A DEFINITE DIRECTION.
• TYPES OF CHROMATOGRAPHIC TECHNIQUES:

5. WORKING PRINCIPLE OF INSTRUMENT:
 THE SEPARATION IS BASED STRICTLY ON THE SIZE OF THE SAMPLE IN
SOLUTION.
 THERE SHOULD BE NO INTERACTION WITH THE COLUMN PACKING.
 THE MODE OF SEPARATION IS NOT BASED ON MOLECULAR WEIGHT, BUT
ON THE SIZE OF THE MATERIAL BEING ANALYZED (USUALLY A POLYMER)
IN SOLUTION.
 IN OTHER WORDS, TO DO GPC CORRECTLY, THE SAMPLE MUST BE
DISSOLVED IN A SUITABLE SOLVENT.

7. MATERIALS AND METHOD FOR
SAMPLE ANALYSIS
Gel
• Gels are used as stationary phase for GPC.
• The pore size of a gel must be carefully controlled in order to be able to apply the gel to a
given separation.
• Other desirable properties of the gel forming agent are the absence of ionizing groups and,
in a given solvent, low affinity for the substances to be separated.
• Commercial gels like PLgel,Sephadex,Bio-Gel (crosslinked polyacrylamide) agarose
gel and Styragel are often used based on different separation requirements.
Eluent
• The eluent (mobile phase) should be a good solvent for the polymer.
• It should permit high detector response from the polymer and should wet the packing
surface.
• The most common eluents in for polymers that dissolve at room temperature GPC
are tetrahydrofuran (THF)

8. SCHEMATIC OF A BASIC GEL PERMEATION CHROMATOGRAPH
This diagram illustrates how the sample is injected into the mobile phase and the path the sample takes
to the detector.

9. A. sample holder,
B. Column
C. Pump
D. Refractive Index Detector
E. UV-vis Detector
A typical Waters GPC instrument including

10. COMPONENTS
1. Pump
Pumps the polymer in solution through the system.
Different polymers produce solutions of different viscosities. To compare data from one analysis
to the next, the pump must deliver the same flow rates independent of viscosity differences
2. Injector
Introduces the polymer solution into the mobile phase.
The injector must be capable of small volume injections (for molecular weight determinations)
and large volume injections (if fraction collecting is desirable). The injector should not disturb
the continuous mobile phase flow. It should also be capable of automatic multiple sample
injection when the sample volume is large.
3. Column Set
Efficiently separates sample components from one another.
High efficiency columns give maximum separating capability and rapid analyses. Every
column must provide reproducible information over extended periods for both analytical and
fraction collecting purposes.

11. 4. Detector
Monitors the separation and responds to components as they elute from the
column.
Detectors must be nondestructive to eluting components if they are to be collected
for further analysis.
The detectors must be sensitive and have a wide linear range in order to respond to
both trace amounts and large quantities of material if necessary.
5. Automatic data processing equipment
Automatically calculates, records, and report numerical values for Mz, Mw,
Mv, Mn, and MWD.
Data systems can also provide complete control of GPC systems so that large
numbers of samples can be run unattended and raw data can be automatically
processed. Today's GPC software offerings need to be able to provide special
calculations for multi-detection processing, band broadening correction, special
calibration routines and polymer branching determination, just to name a few.

12. GPC SEPARATION MECHANISM
POLYMER IS PREPARED AS A DILUTE SOLUTION IN THE ELUENT
AND INJECTED INTO THE SYSTEM
THE GPC COLUMN IS PACKED WITH POROUS BEADS OF
CONTROLLED POROSITY AND PARTICLE SIZE
LARGE MOLECULES ARE NOT ABLE TO PERMEATE ALL OF THE
PORES AND HAVE A SHORTER RESIDENCE TIME IN THE COLUMN
SMALL MOLECULES PERMEATE DEEP INTO THE POROUS
MATRIX AND HAVE A LONG RESIDENCE TIME IN THE COLUMN
POLYMER MOLECULES ARE SEPARATED ACCORDING TO
MOLECULAR SIZE, ELUTING LARGEST FIRST, SMALLEST LAST

14. APPLICATIONS
• IT IS USED TO DETERMINE THE RELATIVE MOLECULAR WEIGHT OF POLYMER
SAMPLES.
• IT IS USED TO DETERMINE THE DISTRIBUTION OF MOLECULAR WEIGHTS OF
POLYMER SAMPLES.
• SEPARATION OF SUGARS POLYPEPTIDES, PROTEINS, LIQUIDS, BUTYL RUBBERS,
POLYSTYRENES, SILICON POLYMERS.
• SEPHADEX G-25 : FOR SEPARATION OF SALTS & AMINO ACIDS FROM PROTEINS .
• SEPHADEX G-75:PURIFICATION OF PROTEINS POLYSACCHARIDES & NUCLEIC
ACIDS.