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    Catechin-Molecularly Imprinted Cryogel for Determination of Catechin in Red Wines by HPLC-DAD-Fluorescence Detector
    (Akademiai Kiado Zrt, 2018) Buyuktuncel, Ebru; Porgali, Esra; Ozkara, Serpil
    A new molecularly imprinted polymer (MIP) was prepared by using catechin (C) as the template molecule. The polymer was characterized by swelling tests, scanning electron microscopy (SEM), Brunauer-Emmett-Teller (BET), and Fourier transform infrared (FTIR) spectroscopy. The MIP with high recovery was selected as a solid-phase extraction (SPE) sorbent in this work. The standard solutions were directly applied onto the SPE cartridges following loading, washing, and elution procedures. A solution of the collected fractions was analyzed by high-performance liquid chromatography-diode-array detection (DAD) and fluorescence detector. The optimization of the method and validation was achieved on a C18 column (5 mu m, 250 x 4.6 mm) with methanol-water (35:65, v/v) mixture adjusted pH 2.5 as the mobile phase at a flow rate of 1 mL min(-1) at room temperature. The selectivity coefficient (k) of imprinted p(HEMA-MAH) cryogel was 5.1-fold that of non-imprinted cryogel. It showed good selectivity and affinity for C molecule. A comparison was made between the results obtained with the MIP cartridges and a traditional C18 reversed-phase cartridge. It was observed that 2.3 times higher recovery of C can be obtained on catechin-MIP cryogel. The results of the presented work showed that the prepared MIP can be used as SPE sorbent for extracting of C from red wines.
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    Evaluation of human interferon adsorption performance of Cibacron Blue F3GA attached cryogels and interferon purification by using FPLC system
    (Elsevier, 2012) Dogan, Ali; Ozkara, Serpil; Sari, Mufrettin Murat; Uzun, Lokman; Denizli, Adil
    In this study, we have focused our attention on preparing supermacroporous cryogels as a potential dye-affinity adsorbent for interferon purification. For this purpose, 2-hydroxyethyl methacrylate (HEMA) and Cibacron Blue F3GA (CB) were selected as main monomer and dye-ligand. Cibacron Blue F3GA attached supermacroporous poly(2-hydroxyethyl methacrylate) [poly( HEMA)/CB] cryogels were prepared and characterized by swelling test, scanning electron microscopy, elemental analysis, and FTIR. After that, the effecting factors such as pH, concentration, interaction time, and ionic strength on the interferon separation were evaluated. The maximum adsorption capacity of poly(HEMA)/CB cryogels was obtained as 38.2 mg/g at pH 6.0. Fast protein liquid chromatography (FPLC) system was used for interferon purification from human gingival fibroblast extract. The chromatography parameters, capacity and selectivity factors, resolution and theoretical plate number were found as 7.79, 9.62, 4.23 and 554, respectively. Although some decreases in total protein content, from 320 mu g to 18 mu g, and interferon activity, from 2.6 x 10(3) IU to 2.2 x 10(3) IU, were determined, specific antiviral activity increased from 7.19 IU/mu g to 122.2 IU/mu g. The purified interferon samples have 97.6% purity determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. After repeated ten adsorption-desorption cycles, no significant decrease was determined in adsorption capacity of cryogel. In result, poly(HEMA)/CB cryogels have an application potential for rapid, cheap and specific purification of interferon. (C) 2012 Elsevier B.V. All rights reserved.
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    Hydrophobic microbeads as an alternative pseudo-affinity adsorbent for recombinant human interferon-? via hydrophobic interactions
    (Elsevier, 2012) Saylan, Yeseren; Sari, Mufrettin Murat; Ozkara, Serpil; Uzun, Lokman; Denizli, Adil
    Hydrophobic interaction chromatography (HIC) is increasingly used for protein purification, separation and other biochemical applications. The aim of this study was to prepare hydrophobic microbeads and to investigate their recombinant human interferon-alpha (rHuIFN-alpha) adsorption capability. For this purpose, we had synthesized functional monomer, N-methacryloyl-L-phenylalanine (MAPA), to provide a hydrophobic functionality to the adsorbent. The poly(2-hydroxyethyl methacrylate-N-methacryloyl-L-phenylalanine) [poly(HEMA-MAPA)] microbeads were prepared by suspension copolymerization. microbeads were characterized using FTIR, swelling behavior, and SEM micrographs. Equilibrium swelling ratio of poly(HEMA-MAPA) and poly(HEMA) microbeads were 53.3% and 69.3%, respectively. The specific surface area and average pore diameters determined by BET apparatus were 17.4 m(2)/g and 47.3 angstrom for poly(HEMA) microbeads and 18.7 m(2)/g and 49.8 angstrom for poly(HEMA-MAPA) microbeads. Adsorption experiments were performed under different conditions. Maximum rHuIFN-alpha adsorption capacity was found to be 137.6 +/- 6.7 mg/g by using poly(HEMA-MAPA) microbeads with a size range of 150-250 mu m and containing 327 mu mol MAPA/g microbeads. Compared with poly(HEMA-MAPA) microbeads, nonspecific rHuIFN-alpha adsorption onto plain poly(HEMA) microbeads was very low, about 4.2 +/- 2.3 mg/g. To determine the effects of adsorption conditions on possible conformational changes of rHuIFN-alpha structure, fluorescence spectrophotometry was employed. Repeated adsorption-elution processes showed that these microbeads are suitable for repeatable rHuIFN-alpha adsorption. (c) 2012 Elsevier B.V. All rights reserved.