Carbofuran Modulating Functions of Acetylcholinesterase from Rat Brain In Vitro

Carbofuran, a potential environmental xenobiotic, has the ability to cross blood brain barrier and to adversely influence brain functions. In the present study, the impact of carbofuran on the biophysical and biochemical properties of rat brain AChE has been evaluated in vitro. This enzyme was membrane-bound which could be solubilised using Triton-X100 (0.2%, v/v), a nonionic detergent, in the extraction buffer (50 mM phosphate, pH 7.4). The enzyme was highly stable up to one month when stored at - 20°C and exhibited optimum activity at pH 7.4 and 37°C. AChE displayed a direct relationship between activity and varying substrate concentrations (acetylthiocholine iodide (ATI)) by following Michaelis-Menten curve. The K m and V m a x values as computed from the Lineweaver-Burk double reciprocal plot of the data were found to be 0.07 mM and 0.066 µmole/mL/min, respectively. The enzyme exhibited IC50 value for carbofuran equal to 6.0 nM. The steady-state kinetic studies to determine mode of action of carbofuran on rat brain AChE displayed it to be noncompetitive in nature with K i value equal to 5 nm. These experiments suggested that rat brain AChE was very sensitive to carbofuran and this enzyme might serve as a significant biomarker of carbofuran induced neurotoxicity.