Nanospines incorporation into the structure of the hydrophobic cryogels via novel cryogelation method: An alternative sorbent for plasmid DNA purification

• Published in: Colloids and surfaces, B, Biointerfaces B, Biointerfaces, 2013-02, Vol.102, p.243-250
• Main Authors: Üzek, Recep, Uzun, Lokman, Şenel, Serap, Denizli, Adil
• Format: Article
• Language: English
• Subjects: acetates; Adsorption; agarose; aqueous solutions; chromatography; colloids; Cryogels - chemistry; desorption; Escherichia coli; Freeze Drying; gel electrophoresis; HIC; Hydrophobic and Hydrophilic Interactions; Hydrophobic cryogels; hydrophobicity; Modified cryogelation; Plasmid DNA; plasmids; Plasmids - isolation & purification; salmon; spermatozoa; temperature
• ISSN: 0927-7765; 1873-4367
• DOI: 10.1016/j.colsurfb.2012.08.020

. [Display omitted] ► Nanospines incorporation into the cryogel structure. ► Hydrophobic cryogels as an alternative sorbent for plasmid DNA purification. ► Surface area enhancement by modified cryogelation approach. ► Evaluation of chromatographic performance of the cryogel by means of FPLC. ► Plasmid DNA isolation from Escherichia coli lysate. In this study, it was aimed to prepare hydrophobic cryogels for plasmid DNA (pDNA) purification from Escherichia coli lysate. The hydrophobicity was achieved by incorporating a hydrophobic ligand, N-methacryloyl-(l)-phenylalanine (MAPA), into the cryogel backbone. In addition to the conventional cryogelation process, freeze-drying step was included to create nanospines. Three different cryogels {poly(2-hydoxyethyl methacrylate-N-methacryloyl-l-phenylalanine)-freeze dried, [P(HEMA-MAPA)-FD]; poly(2-hydoxyethyl methacrylate-N-methacryloyl-l-phenylalanine, [P(HEMA-MAPA)] and poly(2-hydoxyethyl methacrylate)-freeze dried, [P(HEMA)-FD]} were prepared, characterized, and used for DNA (salmon sperm DNA) adsorption studies from aqueous solution. The specific surface areas of cryogels were determined to be 21.4m2/g for P(HEMA)-FD, 17.65m2/g for P(HEMA-MAPA) and 36.0m2/g for P(HEMA-MAPA)-FD. The parameters affecting adsorption such as temperature, initial DNA concentration, salt type and concentration were examined in continuous mode. The maximum adsorption capacities were observed as 45.31mg DNA/g, 27.08mg DNA/g and 1.81mg DNA/g for P(HEMA-MAPA)-FD, P(HEMA-MAPA) and P(HEMA)-FD, respectively. Desorption process was performed using acetate buffer (pH 5.50) without salt. First, pDNA was isolated from E. coli lysate and the purity of pDNA was then determined by agarose gel electrophoresis. Finally, the chromatographic performance of P(HEMA-MAPA)-FD cryogel for pDNA purification was tested in FPLC. The resolution (Rs) was 2.84, and the specific selectivity for pDNA was 237.5-folds greater than all impurities.