Himachal Pradesh University, Department of Chemistry, Shimla, India.
Bioresour Technol. 2011 Feb;102(3):2177-84. doi: 10.1016/j.biortech.2010.11.062. Epub 2010 Nov 20.
Nanogels are promising materials as supports for enzyme immobilization. A new hydrogel comprising of methacrylic acid (MAAc) and N-vinyl pyrrolidone (N-VP) and ethyleneglycol dimethacrylate (EGDMA) was synthesized and converted to nanogel by an emulsification method. Nanogel was further functionalized by Curtius azide reaction for use as support for the covalent immobilization of invertase (Saccharomyces cerevisiae). As-prepared or invertase-immobilized nanogel was characterized by FTIR, XRD, TEM and nitrogen analysis. The characterization of both free and the immobilized-invertase were performed using a spectrophotometric method at 540 nm. The values of V(max), maximum reaction rate, (0.123 unit/mg), k(m), Michaelis constant (7.429 mol/L) and E(a), energy of activation (3.511 kj/mol) for the immobilized-invertase are comparable with those of the free invertase at optimum conditions (time 70 min, pH 6.0 and temperature 45°C). The covalent immobilization enhanced the pH and thermal stability of invertase. The immobilized biocatalyst was efficiently reused up to eight cycles.
纳米凝胶作为酶固定化的支撑材料具有很大的应用前景。本文合成了一种由甲基丙烯酸（MAAc）、N-乙烯基吡咯烷酮（N-VP）和 1,4-丁二醇二甲基丙烯酸酯（EGDMA）组成的新型水凝胶，并通过乳化法将其转化为纳米凝胶。然后通过 Curtius 叠氮反应对纳米凝胶进行功能化，将其用作固定化蔗糖酶（酿酒酵母）的载体。采用傅里叶变换红外光谱（FTIR）、X 射线衍射（XRD）、透射电子显微镜（TEM）和氮分析对纳米凝胶进行了表征。采用分光光度法在 540nm 处对游离酶和固定化酶进行了表征。在最佳条件下（70min 时间、6.0pH 和 45°C 温度），固定化酶的 Vmax、最大反应速率（0.123 单位/mg）、Km、米氏常数（7.429mol/L）和 Ea、活化能（3.511kJ/mol）与游离酶相当。共价固定化提高了蔗糖酶的 pH 值和热稳定性。固定化生物催化剂在八次循环中仍能有效地重复使用。
