Fractionation of Natural Organic Matter by Size Exclusion Chromatography−Properties and Stability of Fractions

An aquatic natural organic matter sample was concentrated by membrane filtration and fractionated using a size exclusion chromatography (SEC) system with a Superdex 75 column. Stability over time was investigated by reinjection of selected fractions. The reinjection yielded single peaks of a more or less Gaussian shape. From the peak shape of the individual fractions it was concluded that non-SEC mechanisms play an important role for the elution behavior. No significant change in the UV chromatograms could be detected over 5 weeks, suggesting a high stability of the fractions and only little or no alterations of the molecular weight distribution during that time period. The fractions were reexamined using a liquid chromatography system with online dissolved organic carbon detection (LCDOC system) with a TSK HW-50 (S) column. The average total recoveries of all fractions were 90 ± 4% for the Superdex system (calculated from UV-peak areas), and 102% and 105% for the spectral UV-absorbances (UVA) and dissolved organic carbon (DOC) contents measured with the LCDOC system. Peak molecular weights (M p) were determined using polystyrene sulfonate (PSS) standards. The M p ranged from M p = 17870 ± 113 Da to M p = 799 ± 28 Da for the original sample and from M p = 17998 ± 306 Da to M p = 737 ± 30 Da for the selected fractions.