Acetylcholinesterase Biosensor Based On Ionic Liquid Functionalized Graphene For Organophosphates Detection

Electrochemical biosensors based on the inhibition of acetylcholinesterase(AChE) are regarded as highly promising candidates for organophosphorus pesticides(OPs) detection because of their advantages with rapidity, simplicity and economy. However, effective immobilization of AChE to solid electrode surface still remains as a great challenge for the fabrication of biosensor. A key consideration for immobilizing enzyme is how to retain its bioactivity. Therefore, the selection of materials and immobilization methods becomes critical. Ionic liquids functionalized graphene sheets(IL-GR) have attracted tremendous attention because of their many excellent properties such as high electrical conductivity, large surface area, good dispersibility and long-term stability in various solvents. In this paper, three different ionic liquid functionalized graphene were synthesized. Using Gelatin(Gel), polyvinyl alcohol(PVA) and sodium alginate-chitosan(SA-CHI) as film-forming material, a series of highly sensitive and stable acetylcholinesterase(AChE) biosensors have been successfully developed. The main contents of this research paper include:1. Ionic liquid(1-(3-aminopropyl)-3-methylimidazolium tetrafluoroborate) functionalized graphene(IL-GR) was synthesized through epoxide ring-opening reaction. A highly sensitive acetylcholinesterase(AChE) biosensor based on IL-GR and Gelatin(Gel) modified glassy carbon electrode(GCE) has been successfully developed. IL-GR was homogeneously dispersed in Gel and dropped on the surface of GCE. Glutaraldehyde(GA) was used as cross-linker to immobilize the AChE on IL-GR–Gel modified GCE. The IL-GR–Gel nanocomposite with excellent conductivity and biocompatibility offered an extremely hydrophilic surface for AChE adhesion. It effectively promoted the electron transfer rate of electrode interface and facilitated the access of substrates to the active centers. Under optimum conditions, the biosensor detected carbaryl and monocrotophos ranging from 1.0×10-14 to 1.0×10-8 mol/L and from 1.0×10-13 to 5.0×10-8 mol/L. The detection limits for carbaryl and monocrotophos were 5.3×10-15 mol/L and 4.6×10-14 mol/L(S/N=3), respectively.2. Ionic liquid modified graphene(IL-GR) was first dispersed in PVA and form IL-GR-PVA dispersion, then IL-GR-PVA and AChE was mixed evenly and was coated on the glass carbon electrode via PVA film-forming, a novel biosensor was developed and used for detecting organophosphorus pesticide(OPs). The IL-GR-PVA nanocomposites with excellent conductivity and biocompatibility offered an extremely hydrophilic surface for AChE adhesion. Linear relationship between the inhibition percentage(I%) and logarithm of concentration of phorate was found in the range from 1.0×10-14 to1.0×10-9 mol/L and from 1.0×10-9 to 1.0×10-6 mol/L, with a detection limit of 6.4×10-15 mol/L(S/N=3).3. Ionic liquid(1-hydroxyethyl-3-bromoethylamine chloride) functionalized graphene and sodium-chitosan alginate modified electrode was fabricated and used for OPs detection. The proposed AChE modified electrode displayed excellent electrochemical catalytic activities toward the detection of monocrotophos and methamidophos. The ionic liquid here could improve the dispersibility of graphene and provide a favorable conductive microenvironment for the immobilized AChE, thus promote the electron transfer rate at the electrade surface. Under optimum conditions, the biosensor detected monocrotophos in the linear range from 5.0×10-12 to 5.0×10-7 mol/Lwith a detection limit of 2.3×10-12 mol/L(S/N=3), and detected methamidophos in the range from 1.0×10-12 to 5.0×10-8 mol/Lwith a detection limint of 5.6×10-13 mol/L(S/N=3).