Influence of ionic strength and organic modifier concentrations on characterization of aquatic fulvic and humic acids by high-performance size-exclusion chromatography

Aquatic fulvic acid (FA) and humic acid (HA) were characterized by an aqueous high-performance size-exclusion chromatography (HPSEC) using a hydrophilic polymeric stationary phase and an aqueous eluent at neutral pH and low-ionic strength (5 mM Na2HPO4; final ionic strength, 13 mM). Employed HPSEC showed low sensitivity of FA to variations in ionic strength (13 and 100 mM) and contents of organic modifier (0.1 or 40% methanol) in aqueous eluent. Under these analytical conditions, peak maxima of relative UV signals versus molecular mass (Mr) defined as Mp′ and peak maxima of relative mass concentrations versus Mr defined as Mp of FA were shown to be located at 548–690 and 500, respectively. Organic modifier concentrations of 40% methanol in aqueous eluent enabled not only analysis of FA, but also analysis of some aquatic HA by HPSEC. Analysis showed Mp′ and Mp values of aquatic HAs around 1000 and 600, respectively. Measured molecular mass data of FA were found to be consistent with the recently published data describing low molecular masses of FA. Results recommend the use of the described HPSEC as a simple, rapid, reproducible, low-cost method giving consistent molecular sizes/masses of FA and some aquatic HAs.