Turkish Journal of Chemistry




An efficient and inexpensive monolithic stationary phase (PHEMA-HA) has been prepared through an easy process comprising addition of humic acid (HA) to a mixture of 2-hydroxyethyl methacrylate (HEMA) and N,N'-methylenebisacrylamide (MBAAm) and subsequent radical-polymerization at -20 $^{\circ}$C. The prepared monolithic material was characterized in terms of various techniques and methods such as elemental analysis, FTIR spectroscopy, scanning electron microscopy, mercury porosimetry, hydrolytic stability tests, pH$_{pzc}$ measurements, water-holding capacity, and water permeability. The amount of HA incorporated into the structure was calculated as 45 mg/g from the elemental analysis results. The study was conceptualized on the basis of protein ion-exchange chromatography, and some model proteins (i.e. $a$-chymotrypsinogen a, cytochrome c, lysozyme, human serum albumin, and myoglobin) were used in the chromatographic experiments. The effect of ionic strength and pH (5.0, 6.0, 7.0) on the retention behavior of proteins was investigated. The results revealed a typical ion-exchange behavior for the PHEMA-HA stationary phase, and with increasing gradient slope the model proteins were found to elute faster. Some model proteins could be separated by applying gradient elution where NaCl was used as the modifier. Dynamic adsorption capacity of PHEMA-HA was obtained by frontal analysis and was found as 4 mg/mL for Lys.


Cation exchange, cryogel, high-performance liquid chromatography, humic acid, separation

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