Resolution power and calibration in gel chromatography

The resolution power in gel chromatography is determined by both the efficiency and selectivity of the chromatographic system. 

The efficiency of a real GPC system depends above all on the rate of mass transfer between mobile phase and gel phase, as well as on the extent of secondary processes. Quantitatively, the efficiency can be expressed by the terms like width (w) or deviation (o) of the chromatographic peak, as well as by other terms of the theoretical plate concept. Since the diffusion rate of solute molecules decreases with an increase of their dimensions, one has to expect generally lower efficiency in gel chromatography of macromolecules in comparison with any other mode of liquid chromatographic separation of low molecular substances. 

The courses of calibration curves can be expressed mathematically by the appropriate polynomes. The central part of calibration curves that is most important for practical use can, however, often be quite well approximated by the straight line (cf.. Fig. 4.6.1). 

In analytical practice, the logarithm of sample molar masses, or molar volumes, is plotted versus retention volumes in calibration dependences of low molecular substances while values or effective hydrodynamic volumes, are used as size parameters in gel chromatography of macromolecules [12,13]. is often called universal calibration parameter because in ideal gel chromatography of randomly coiled macromolecules, it enables the transfer of data from one polymer to another regardless of both the physical (linearity, branching, tacticity, etc.) and the chemical (composition) structure of macromolecules [12]. The hydrodynamic volume of a particular polymer is proportional to the product of its molar mass and limiting viscosity number [ij], in the solvent that is used as mobile phase [ij]Mm. 

Globular proteins are uniform (monodisperse) as to their molar mass and usually exhibit approximately spherical shape. In this case, the corresponding Stokes radius can be used as the calibration size parameter. 

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