Construction of a flow-type of measuring system with functions for capturing and sensing heavy metal ions and its characteristics

○光度流动注射生物传感系统的构建及其特性嗜热菌蛋白酶是最典型的锌酶和耐热中性蛋白酶之一，溶解在含40%甘油的硼酸盐缓冲溶液中，并固定在具有可控孔径（24.2 nm）的多孔玻璃珠上。固定化率约为30%。将固定化制剂装入小聚合物柱中，然后连接到光度传感系统。锌（II）离子范围从1. OnM至1。0 mM，但由于接触螯合剂（0. 1 M EDTA，pH 4. 0），酶柱并不总是保持优异的操作稳定性。○量热流动注射生物传感系统的构建及其性能将溴化钠作为嗜热菌蛋白酶的良好溶剂添加到酶-硼酸盐缓冲溶液中，从而可以以令人满意的产率固定化酶。将该制剂填充在相同尺寸的柱中，应用于量热传感系统。酶柱对1.0 mL注射4.0 mM合成底物（N-（3-[2-呋喃基]丙烯酰基-甘氨酰-亮氨酸酰胺），pH 8.0）表现出高催化活性，温度响应约为4.0 mK。将2.5mL螯合剂引入柱中导致催化活性显著降低。一旦螯合剂暴露的柱通过注入锌（II）离子溶液容易地再活化。几乎无锌柱的再活化与加入的锌离子浓度有关。因此，基于脱辅基酶再活化方法，在0.01至1.0mM的浓度范围内肯定地实现了锌（II）离子的量热测定。

○ Construction of a photometric flow-injection biosensing system and its characteristics Thermolysin, one of the most typical zinc-enzymes and thermostable neutral protease, was dissolved in borate buffer solution containing 40 % glycerol and immobilized onto porous glass beads with controlled pore size(24.2 nm). The immobilization yield was about 30 %. The immobilized preparations were packed into a small polymer column and then, attached to a photometric sensing system. Zinc (II) ions ranging from 1. 0 nM to 1. 0 mM could be determined, but the enzyme column did not always retain excellent operational stability due to exposure to chelating agents (0.1 M EDTA, pH 4.0).○ Construction of a calorimetric flow-injection biosensing system and its performance Sodium bromide as good solvent for thermolysin was added to the enzyme-borate buffer solution, and thereby, the enzymes could be immobilized in a satisfactory yield. The preparations packed in the same size of the column was applied to a calorimetric sensing system. The enzyme column exhibited high catalytic activity for 1.0 mL-injections of 4.0 mM synthetic substrate (N- (3-[2-furyl]acryloyl-gly-leu amide), pH 8.0) and the temperature response was about 4.0 mK. Introduction of 2.5 mL of the chelating agent to the column caused significant decrease in the catalytic activity. The once chelator-exposed column was readily reactivated by injecting zinc)II) ions solution. Reactivation of the almost zinc-free column was related to concentrations of zinc)II) ions added. Thus, calorimetric detrmination of zinc(II) ions in a concentration range of 0.01 to 1.0 mM was positively achieved based on.the apoenzyme reactivation method.