Gonçalves Gabriel Ramos Ferreira, Gandolfi Olga Reinert Ramos, Santos Carilan Moreira Souza, Bonomo Renata Cristina Ferreira, Veloso Cristiane Martins, Fontan Rafael da Costa Ilhéu
Processes Engineering Laboratory, State University of Bahia Southwest, Itapetinga, BA, Brazil.
Processes Engineering Laboratory, State University of Bahia Southwest, Itapetinga, BA, Brazil.
J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Oct 15;1033-1034:406-412. doi: 10.1016/j.jchromb.2016.09.016. Epub 2016 Sep 14.
Affinity techniques are frequently used to purify biocompounds, because of specific interactions observed in many cases. One example are the lectins, proteins connected in a reversible manner and specific to carbohydrates or sugar-containing molecules. Four different methods were investigated (epoxy, Schiff base, glutaraldehyde and ethylenediamine) to immobilize the carbohydrate N-acetyl-d-glucosamine (d-GlcNAc) on the surface of supermacroporous cryogels made for lectin purification. The glutaraldehyde method presented the highest immobilization capacity of d-GlcNAc (147.77mg/g), while the ethylenediamine method presented the lowest capacity (32.47mg/g). FTIR spectra analysis confirmed the presence of the immobilized carbohydrate. The cryogels containing d-GlcNAc immobilized by the different methods were characterized in terms of swelling capacity, degree of expansion, porosity and constituent fractions. Results showed that the activation methods did not affect the macroporous structure. Images obtained from scanning electron microscopy evidenced the presence of interconnected macropores in the structure of the cryogels produced. The cryogels presented even lower flow resistance in the permeability analysis. Finally, the cryogel modified by the glutaraldehyde method was used in the Concanavalin A lectin adsorption process, presenting an adsorptive capacity of 44.49mg/g and high stability after five cycles of use.
由于在许多情况下观察到的特异性相互作用，亲和技术经常用于纯化生物化合物。一个例子是凝集素，它是与碳水化合物或含糖分子以可逆方式连接且具有特异性的蛋白质。研究了四种不同的方法（环氧法、席夫碱法、戊二醛法和乙二胺法），将碳水化合物N-乙酰-d-葡萄糖胺（d-GlcNAc）固定在用于凝集素纯化的超大孔冷冻凝胶表面。戊二醛法呈现出最高的d-GlcNAc固定容量（147.77mg/g），而乙二胺法呈现出最低的容量（32.47mg/g）。傅里叶变换红外光谱分析证实了固定化碳水化合物的存在。对通过不同方法固定d-GlcNAc的冷冻凝胶的溶胀能力、膨胀程度、孔隙率和组成部分进行了表征。结果表明，活化方法不影响大孔结构。扫描电子显微镜获得的图像证明了所制备的冷冻凝胶结构中存在相互连接的大孔。在渗透性分析中，冷冻凝胶表现出更低的流动阻力。最后，戊二醛法改性的冷冻凝胶用于伴刀豆球蛋白A凝集素的吸附过程，其吸附容量为44.49mg/g，在五个使用循环后具有高稳定性。
