Macromolecular structure of a commercial humic acid sample

The molecular structure of a commercial sample of humic acids (HA) was investigated by membrane dialysis experiments (MD) and low-pressure size-exclusion chromatography (LP-SEC). MD showed that HA molecules were retained by dialysis membrane with a cut-off of 6–8 kDa, independently from HA concentration (15 or 150 mg L−1), NaHCO3 concentration (0.005–2.0 mol L−1), and from propan 2-ol (0–5 v/v %). SEC experiments at low pressure gave chromatograms with a broad peak, with an elution volume between those of the globular proteins bovine serum albumin (molecular weight = 66.5 kDa) and lysozyme from egg (molecular weight = 14.4 kDa). The pattern of the chromatogram did not vary with HA concentration, and second-run chromatograms of single eluted fractions showed relatively sharp peaks. From these data, we reveal that the commercial HA sample analysed has a macromolecular structure rather than being a supramolecular aggregate of relatively small molecules, as recently proposed for some samples of HA obtained from different sources.