For SEC analysis, sample concentration is very important. The following is the pretreatment procedure of sample.
Basically, the sample should be dissolved in the same solvent that is to be used as the eluent.
1) Dissolving the sample in such solvent will make the blank peaks as small as possible.
2) In case the sample has a molecular weight of 1,000,000 minimum, soak it in such solvent for 12 to 24 hours to let it swell. After the swelling, stir the solvent gently for dissolution.
Strong agitation or use of an ultrasonic bath for dissolution will degrade such sample.
3) In case the sample is a polymer, its concentration in the solution and the injection volume should be 0.05 to 0.5 % and 50 to 100 μL, respectively.
If the concentration is higher, the retention volume of the sample will increase. The optimum concentration changes with the molecular weight and the viscosity. The following table gives the molecular weight vs. the optimum concentration.
|Molecular weight||Sample concentration|
|Injection volume per column|
|KF-600 series||KF-800 series|
less than 1.0
|5 μL||50 to 100 μL|
|5,000 to 25,000||
less than 0.5
|25,000 to 200,000||
less than 0.25
|200,000 to 2,000,000||
less than 0.1
less than 0.05
4) In case the sample is organic, it is desirable that the concentration be 1 % maximum and the injection volume, 50 μL maximum.