Combination of asymmetric flow field-flow fractionation (AF⁴) and total-reflexion X-ray fluorescence analysis (TXRF) for determination of heavy metals associated with colloidal humic substances

To assess the structural variability of colloidal humic substances and the associated heavy metals an off-line coupling of asymmetric flow field-flow fractionation (AF⁴) with total-reflection X-ray fluorescence analysis (TXRF) is presented. AF⁴ allows a rather gentle separation of colloids with a minimum of interference and artifacts as no shear forces, drying, or interactions with a stationary phase are involved. After a calibration with suitable polymer particles of known molecular weight, the molecular weight distribution of colloidal humic substances between 1 and 10³ kDa can be assessed with AF⁴. The combination with TXRF permits a simultaneous multielement analysis after preconcentration of samples on the AF⁴ channel using an optimized buffer. The analysis of seepage and sewage water sample and a sewage sludge sample yielded continuous distributions of the molecular weight and the associated heavy metals. The potential of AF⁴-TXRF coupling for the study of metal ion exchange equilibria with colloids was demonstrated by spiking seepage water with various heavy metals and subsequent AF⁴-TXRF analysis of the heavy metals bound to the colloidal fraction (Cu, Cr, Zn, Ni, Co).