Smaller molecules (smaller than the pores in the gel) must traverse a longer path through the gel than larger molecules, PC Fig. 10.18. Larger molecules are therefore eluted first. The method provides a relative molecular weight after the column is calibrated with standards of known molecular weight, PC Fig. 10.19. Monodisperse polystyrene is often employed as a standard. Often polystyrene standards are used to calibrate a column for use with other polymers, a common practice that is well-known to produce systematic errors, PC Fig. 10.21. The practice is perpetuated because it is simple, in spite of the fact that it is inaccurate.
The reliability of the measurement can be enhanced by the so-called universal calibration. It exploits the fact that the intrinsic viscosity for a spherical particle can be written as
[eta] = 2.5LVh/M
where Vh is the hydrodynamic volume and L is Avogadro's number. The universal calibration plots log [eta]M vs. elution volume, PC Fig. 10.22.
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