Measurement of diffusion coefficients of humic substances in a hydrogel and in water by fluorescence correlation spectroscopy

Diffusion coefficients of three different humic substances (HS) were measured in both water and in agarose hydrogels. The three HS were standard Suwannee River humic and fulvic acid [1] and a peat humic acid [2]. Measurements were made by fluorescence correlation spectroscopy (FCS). In general, values of diffusion coefficients were in the range of 1.9-2.5 x 10(-10) m(2) s(-1), for all of the HS under all conditions (except a single peat derived HS at low pH). The small degree of polydispersity of the HS (observed both in the autocorrelation functions of the FCS and by atomic force microscopy, AFM) did not preclude the use of a single diffusion coefficient to explain the data for a single HS at a particular pH. However, at low pH, the peat HS exhibited a high degree of polydispersity with the formation of aggregates of up to 30-50 nm in diameter, as measured by atomic force microscopy and ultrafiltration. In this case, FCS did not give reliable values of diffusion coefficients, most likely due to extensive aggregation. Excluding this extreme case, diffusion coefficients in the gel were approximately 10-20% lower than in the water, because of either interactions with the gel or increased path lengths due to an increased tortuosity of the gel compared to water. Large HS aggregates were excluded from the gel.