辣根过氧化物酶在改性壳聚糖珠上的固定化。
Immobilization of horseradish peroxidase on modified chitosan beads.
机构信息
Chemistry Department, Drexel University, Philadelphia, PA, USA.
出版信息
Int J Biol Macromol. 2010 Apr 1;46(3):324-30. doi: 10.1016/j.ijbiomac.2009.12.018. Epub 2010 Jan 9.
A method has been developed to immobilize horseradish peroxidase (HRP) on modified chitosan beads by means of graft copolymerization of polyethylacrylate in presence of potassium persulphate and Mohr's salt redox initiator. The activity of free and immobilized HRP was studied. FTIR spectroscopy and scanning electron microscopy were used to characterize HRP immobilization. The efficiency of the immobilization was investigated by examining the relative enzymatic activity of free enzyme before and after the HRP immobilization. The obtained values were found to reach 98.4%. The results show that the optimum temperature of immobilized HRP was 45 degrees C, which was identical to that of free enzyme, and the immobilized HRP exhibited a higher relative activity than that of free HRP over 45 degrees C. The optimal pH for immobilized HRP was 10, which was higher than that of the free HRP (pH 9.0), and the immobilization resulted in stabilization of enzyme over a broader pH range. The apparent kinetic constant value (K(m)) of immobilized HRP was 3.784 mmol ml(-1), which was higher than that of free HRP. On the other hand, the activity of immobilized HRP decreased slowly against time when compared to that of the free HRP, and could retain 65.8% residual activity after 6 consecutive cycles.
已开发出一种通过在过硫酸钾和莫尔盐氧化还原引发剂存在下进行聚丙烯酸接枝共聚，将辣根过氧化物酶（HRP）固定在改性壳聚糖珠上的方法。研究了游离和固定化 HRP 的活性。傅里叶变换红外光谱和扫描电子显微镜用于表征 HRP 固定化。通过检查游离酶在 HRP 固定化前后的相对酶活性来研究固定化的效率。得到的值达到 98.4%。结果表明，固定化 HRP 的最适温度为 45°C，与游离酶相同，固定化 HRP 在 45°C 以上表现出比游离 HRP 更高的相对活性。固定化 HRP 的最适 pH 为 10，高于游离 HRP（pH 9.0），固定化导致酶在更宽的 pH 范围内稳定。固定化 HRP 的表观动力学常数（K(m)）值为 3.784 mmol ml(-1)，高于游离 HRP。另一方面，与游离 HRP 相比，固定化 HRP 的活性随时间缓慢下降，并且在连续 6 次循环后可保留 65.8%的残余活性。
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