Investigations On Solid-Phase Extraction Of Biomacromolecules Using Anion Exchange Resin

Separation and purification of biomacromolecules is an important procedure for life-science studies. Thus, the employment of suitable method is the key to obtain purified biomacromolecules. Solid-phase extraction offers an excellent alternative to separate and purify biomacromolecules, due to its high separation and purification efficiency, ease of operation, speediness, minimized contamination by organic solvents and low running cost. In this work, hemoglobin and DNA were separated and purified from human whole-blood using solid-phase extraction technology with 330 anion exchange resin as sorbent.Hemoglobin was absorbed onto the resin via hydrophobic interactions, which was afterwards eluted with Tris-HCl (pH 8.9) solution. The experiments have demonstrated that retention and stripping efficiencies of 87% and 70% were achieved respectively by employing an absorption time of 20 min and an elution time of 20 min. The absorption capacity for hemoglobin was 5.0 mg/g. Some of the experimental parameters were optimized in order to improve the sorption and elution efficiencies, including the surface area of resin, the ionic strength, the acidity and the type of eluents. SDS-PAGE gel electrophoresis indicated that at the optimized experimental conditions, effective separation of hemoglobin from human whole blood could be achieved.By means of electrostatic interaction, DNA could be adsorbed onto the resin surface with a retention effificiency of of 96%. Afterwards,83% of the retained DNA could be recovered by elution with a Na2HPO4 solution of 40 mmol/L. The experimental parameters governing the sorption of DNA were investigated, including the resin surface area, the ionic strength, the acidity and the type of eluents. The isolation of DNA from human whole blood was achieved by using this technique, and the purified DNA sample was tested by using polymerasechain reaction (PCR), giving rise to satisfactory results.330 anion exchange resin could be used to extract hemoglobin and DNA without any modification, which simplified the operations. Conventional instrumentation suffices thedetection of the experimental process. The retained DNA could be directly eluted from the resin surface and no pre-washing procedure was required. In addition, the use of organic reagents and/or high ionic strength chaotropic reagents were avioded, which facilitating further biological investigations..