Separation of humic acids using capillary isoelectric focusing

Humic acids are natural polymers occurring in soils and sediments. These molecules modify the structure of soil and sediment and bind metal ions but because of structural diversity analysis of humic acids is difficult. Capillary isoelectric focusing (CIEF) was first applied in this field by Schmitt et al. but they obtained 3-4 fractions only. The aim of present study was to develop a new and reproducible CIEF method for the separation of humic acids with better resolution. Commercial humic acid samples purchased from Fluka and Aldrich and a peat humic acid were used for method development and comparative studies. Methyl cellulose and polyvinyl alcohol (PVA) were used to increase resolution. Reduced Triton X-100, SDS and urea were applied as anticoagulants. Effects of additives on resolution and reproducibility were compared in different concentrations and optimum concentrations were found to be as follows: 0.2% methyl cellulose, 0.5% PVA and 0.3% reduced Triton X-100. Thus, 30-50 stable fractions could be obtained depending on the sample.