Band broadening in size-exclusion chromatography of polydisperse samples

Understanding and controlling the band broadening is essential to obtain accurate molar-mass distributions by size-exclusion chromatography (SEC). In this paper, band broadening in SEC is reviewed from a contemporary perspective. The observed band broadening is due to dispersion inside and outside the chromatographic column (undesirable band broadening) and to the polydispersity of the sample (desirable SEC selectivity). The various contributors to band broadening are discussed. Integrity plots are introduced as a tool to evaluate the performance of specific SEC columns at given experimental conditions. For narrow polymer standards on single SEC columns the observed peak width is dominated by the chromatographic dispersion. MALDI-ToF-MS is demonstrated as an alternative to determine the PDI of narrowly distributed samples. The plate heights encountered at very high reduced velocities are found to be lower than expected. This is advantageous for fast separations by SEC. (C) 2004 Elsevier B.V. All rights reserved.