Manyetik Silika Partiküllerle Manyetik Alan Varlığında Boya Afinite Temelli Albümin Saflaştırılması

Abstract

Human serum albümin is the most abundant protein in the circulatory system. It performs many physiological functions such as the regulation, distribution and metabolism of a large number of endogenous and exogenous sunstances such as adjusting the osmotic pressure of the blood, bile acids, bilirubin, long cahin fatty acids, amino acids, steroids, metal ions and drugs. It is used for therapeutic purposes in case of excessive blood loss and kidney disease and protein losses and is one of the basic products derived from blood. Cohn fractionation method is generally used for purification from plasma. However, this method is not selective and can cause protein denaturation. Dye ligand adsorbents for purification of proteins are preferred beacuse they provide simple, fast and inexpensive separation and can provide high recovery from crude extract 20-80 times in one time without purifications and pretreatment. In this study, Cibacron Blue F3GA ligand, known to be associated with the albumin of magnetic silica particles, was ligated and albumin purificaiton was carried out in the magnetic system. The presence of magnetite in the silica particles with a 222 m2/g surface area was shown by ESR and VSM techniques and the spectroscopic splitting factor, g, was calculated as 2.3153. The amount of Cbacron Blue F3GA bound to the magnetic particles was increased with the increasing dye concentration and was determined by elemental analysis. The covalent binding of the dye was also shown by FTIR. The HSA adsorption studies were performed under different conditions. The maximum HSA adsorption was achieved at pH 5.5 as 27.82 mg/g and reached a plateu value at 3.0 mg/mL HSA concentration. The increase in the medium temperature and ionic strength was decreased the HSA adsorption. The mathematical calculations have shown that the adsoption of HSA onto the magnetic silica particles is consistent with the Langmuir isotherm and adsorption occured chemically. 1.0 M of NaCl solution was used as a desorption agent and after 0 adsorption-desorption cycle the adsorption capacity of the adsorbend decreased only 5%. The adsorption of albumin from artificial plasma occured as 48.6 mg/g and the 97% of total protein adsorption from plasma occured as albumin.