Elçin A E, Elçin Y M
Department of Biology, Faculty of Science, Ankara University, Turkey.
Artif Cells Blood Substit Immobil Biotechnol. 2000 Jan;28(1):95-111. doi: 10.3109/10731190009119788.
Urease (EC 3.5.1.5) was immobilized within polyanionic carboxymethylcellulose/alginate (CMC/Alg) microspheres coated with a cationic polysaccharide, chitosan (C). Coating with chitosan improved the mechanically durability of the polyanionic microspheres, as well as increased enzyme immobilization yield [approximately 0.4 mg.mL-1 gel]. The effects of chitosan coating and CMC/Alg ratio on the water uptake and spherical morphology of the microspheres were investigated. The optimal pH of urease was not extensively affected by the immobilization procedure. However, the optimal temperature of urease activity increased upto 60 and 65 degrees C within CMC/Alg and C(CMC/Alg) microspheres, respectively, while the optimum for the free enzyme was 50 degrees C. The half life (t1/2) and deactivation rate constant (kd) of free urease were 79 min and 8.77 x 10(-3) min-1, respectively, whilst the t1/2 and kd values of urease within polyanion and polycation-coated polyanion microspheres were 142 min and 4.88 x 10(-3).min-1, and 179 min and 3.87 x 10(-3).min-1, at 80 degrees C, respectively. While the activation energy of the hydrolysis reaction of free urease was found to be 11.86 kJ.M-1.dm-3, it increased to 18.91 and 20.02 kJ.M-1.dm-3, for the immobilized urease within CMC/Alg and C(CMC/Alg) microspheres, respectively. The free enzyme exhibited K(m) and Vmax values of 2.85 mM.dm-3 and 31.9 mM.dm-3.s-1.g-1p-1, respectively, whilst the K(m) and Vmax for urease within polyanion and polycation-coated polyanion microspheres were 3.94 mM.dm-3 and 73.4 mM.dm-3.s-1.g-1.p-1, and 4.22 mM.dm-3 and 81.4 mM.dm-3.s-1.g-1.p-1, in the same order. C(CMC/Alg) microspheres showed a nearly stable urease activity of around 80-85% of the initial maximum activity, after the first 100 minutes.
脲酶（EC 3.5.1.5）固定在涂有阳离子多糖壳聚糖（C）的聚阴离子羧甲基纤维素/藻酸盐（CMC/Alg）微球内。壳聚糖涂层提高了聚阴离子微球的机械耐久性，并提高了酶固定化产率[约0.4 mg·mL-1凝胶]。研究了壳聚糖涂层和CMC/Alg比例对微球吸水率和球形形态的影响。固定化过程对脲酶的最佳pH影响不大。然而，在CMC/Alg和C(CMC/Alg)微球中，脲酶活性的最佳温度分别提高到60和65摄氏度，而游离酶的最佳温度为50摄氏度。游离脲酶的半衰期（t1/2）和失活速率常数（kd）分别为79分钟和8.77×10-3分钟-1，而在80摄氏度时，聚阴离子和聚阳离子涂层聚阴离子微球中脲酶的t1/2和kd值分别为142分钟和4.88×10-3分钟-1，以及179分钟和3.87×10-3分钟-1。虽然游离脲酶水解反应的活化能为11.86 kJ·M-1·dm-3，但对于CMC/Alg和C(CMC/Alg)微球内固定化的脲酶，活化能分别增加到18.91和20.02 kJ·M-1·dm-3。游离酶的K(m)和Vmax值分别为2.85 mM·dm-3和31.9 mM·dm-3·s-1·g-1·p-1，而聚阴离子和聚阳离子涂层聚阴离子微球中脲酶的K(m)和Vmax值依次为3.94 mM·dm-3和73.4 mM·dm-3·s-1·g-1·p-1，以及4.22 mM·dm-3和81.4 mM·dm-3·s-1·g-1·p-1。在最初的100分钟后，C(CMC/Alg)微球显示出接近稳定的脲酶活性，约为初始最大活性的80-85%。
