de Melo J V, Cosnier S, Mousty C, Martelet C, Jaffrezic-Renault N
Anal Chem. 2002 Aug 15;74(16):4037-43. doi: 10.1021/ac025627+.
Enzyme-based field effect transistors (ENFETs) for urea determination were developed based on the immobilization of urease within two different clay matrixes, one cationic (Laponite) and the other anionic (layered double hydroxide (LDH)), cross-linked with glutaraldehyde. The biosensor based on the enzyme immobilized in Laponite shows a greater sensitivity and smaller dynamic linear range, because the enzymatic reaction is protected from the effect of the buffer capacity of the outer medium. The apparent Michaelis-Menten constant, Km(app), is quite similar for both biosensors. Inhibition of the enzyme by sodium tetraborate was investigated. Tetraborate acts as a competitive inhibitor for urease in the two different types of clay, the inhibitor effect being stronger for the LDH/urease biosensor. In particular, the maximum limit of the dynamic linear range extends from 1.4 mM in the absence of the inhibitor to 12 mM in the presence of 0.5 mM tetraborate. The Km(app) values in the presence of 0.5 mM tetraborate for Laponite and LDH biomembranes were 10 and 62 mM, respectively. Comparison of the inhibition constant values, Ki 0.16 and 0.05 mM for Laponite and LDH biosensors, respectively, clearly indicates a stronger enzyme-inhibitor interaction in the LDH/urease biomembrane.
基于尿素酶固定在两种不同粘土基质（一种阳离子型（锂皂石），另一种阴离子型（层状双氢氧化物（LDH）））中并与戊二醛交联，开发了用于尿素测定的基于酶的场效应晶体管（ENFET）。基于固定在锂皂石中的酶的生物传感器显示出更高的灵敏度和更小的动态线性范围，因为酶促反应免受外部介质缓冲容量的影响。两种生物传感器的表观米氏常数Km(app)非常相似。研究了四硼酸钠对酶的抑制作用。在两种不同类型的粘土中，四硼酸盐对尿素酶起竞争性抑制剂的作用，对LDH/尿素酶生物传感器的抑制作用更强。特别是，动态线性范围的最大极限从不存在抑制剂时的1.4 mM扩展到存在0.5 mM四硼酸盐时的12 mM。在存在0.5 mM四硼酸盐的情况下，锂皂石和LDH生物膜的Km(app)值分别为10和62 mM。锂皂石和LDH生物传感器的抑制常数分别为0.16和0.05 mM，比较表明LDH/尿素酶生物膜中酶-抑制剂相互作用更强。
