Capacitive approach to determine phospholipase A2 activity toward artificial and natural substrates
Capacitive approach to determine phospholipase A2 activity toward artificial and natural substrates
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DOI：
10.1021/ac980102w
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发表时间：
1998-09-01
影响因子：
7.4
通讯作者：
Wolfbeis, OS
中科院分区：
文献类型：
--
作者：
Mirsky, VM;Mass, M;Wolfbeis, OS
电容法已被用来开发一种测定磷脂酶活性的新方法。传感电极具有类似Au/S(CH2)(17)CH3/基板/电解质的结构。由磷脂酶 A(2) 介导的底物水解导致不溶性底物形成水溶性产物。这导致这些产物解吸到水相中并相应增加电极电容。反应产物高水溶性的要求可以通过两种方式实现。第一种方法使用短链磷脂作为底物，在这种情况下，形成水溶性产物，并且不需要额外的试剂来促进这些产物的解吸。通过该策略制备的传感器提供了磷脂酶的灵敏定性检测。第二种方法是基于使用水溶性受体（例如β-环糊精）来溶解水解产物，它允许半定量检测磷脂酶对长链天然底物的活性。发现这种情况的反应动力学是单指数的并且线性依赖于磷脂酶浓度。以蜂毒磷脂酶A(2)和大豆卵磷脂为底物测试，该方法的检出限约为0.5 ng/mL（500 mu 单位/mL）。
A capacitive approach has been employed to develop a novel method to determine phospholipase activity. The sensing electrodes have a structure like Au/S(CH2)(17)CH3/substrate/electrolyte. Hydrolysis of the substrate, mediated by phospholipase A(2), leads to the formation of watersoluble products from the insoluble substrate. This results in desorption of these products into aqueous phase and corresponding increase of the electrode capacitance. The requirement of high water solubility of the reaction products can be achieved in two ways. In the first, short-chain phospholipids are used as the substrate, in which case, water-soluble products are formed and no additional reagents are required to promote desorption of these products. The sensors prepared by this strategy provide sensitive qualitative detection of phospholipases. The second way is based on the use of a water-soluble acceptor (for example, beta-cyclodextrin) to solubilize the products of hydrolysis, It allows semiquantitative detection of phospholipase activity toward long-chain natural substrates, The reaction kinetics for this case was found to be monoexponential and linearly dependent on the phospholipase concentration. The detection limit of this method, as tested with phospholipase A(2) from bee venom and soy bean lecithin as the substrate, is similar to 0.5 ng/mL (500 mu units/mL).