Langmuir, Langmuir-Blodgett and layer-by-layer ultra-thin film properties of amphiphilic molecules and organophosphorus hydrolase (OPH), and the OPH thin film bioassay of an organophosphorus compound (paraoxon).

摘要

Organophosphorus hydrolase is an enzyme of high efficiency in catalyzing the hydrolysis of organophosphorus compounds. This process is of great importance because the toxicities of hydrolysis products are significantly lower than the original organophosphorus compounds. The current research emphasizes the immobilization of ultra thin OPH film onto different substrates as biosensors for the detection of organophosphorus compounds. The thin film immobilization overcomes such disadvantages as delayed signal transfer procedure, outer layer barrier, and poor reproducibility encountered in thick layer immobilized biosensors. Therefore, thin film immobilization offers great potential for producing high performance biosensors and reducing the system size.; Langmuir-Blodgett film deposition methodology makes it possible to transfer properly organized and oriented molecules from the air-water interface to solid substrates, therefore producing films of one layer thickness that have controllable molecular conformation. The properties of OPH macromolecules as Langmuir film and Langmuir-Blodgett film were investigated systematically. The surface chemistry studies indicated a well-organized and closely packed monolayer. This monolayer had oriented molecular conformation, high stability and one layer thickness. When employed as a biosensor for the detection of paraoxon, the hydrolysis effect of OPH on paraoxon can be detected and monitored by the aid of spectrophotometers as well as the in situ microscopic observations. The biosensor characterization showed detection of paraoxon as low as 10−9 M and a linear relationship response over the range of 1.0 × 10−7 to 1.0 × 10 −5 M (0.27–27 ppm). The biosensor prepared by L-B film immobilization technique demonstrated high sensitivity, rapid response time and good reproducibility.; The layer-by-layer self-assembly multilayer film fabrication was investigated using OPH (pH 7.3) as polycation and poly (3-thiophene acetic acid) (PTAA at pH 8.8) as polyanion. This method has an extremely simple procedure and can be used as a general approach for the fabrication of multi-composite molecular assemblies. Spectroscopic and microscopic studies of the film showed a consecutive deposition of each layer and this system was employed for the detection of paraoxon.