Cyclovoltammetric acetylcholinesterase activity assay after inhibition and subsequent reactivation by using a glassy carbon electrode modified with palladium nanorods composited with functionalized C60 fullerene

A glassy carbon electrode (GCE) was modified with a nanocomposite consisting of tetraoctylammonium bromide (TOAB), C-60 fullerene, and palladium nanorods (PdNRs). The PdNRs were hydrothermally prepared and had a typical width of 20 +/- 2 nm. The nanocomposite forms stable films on the GCE and exhibits a reversible redox pair for the C-60/C-60 (-) system while rendering the surface to be positively charged. The modified GCE was applied to fabricate an electrochemical biosensor for detecting acetylcholinesterase (AChE) by measurement of the amount of thiocholine formed from acetylthiocholine, best at a working voltage of -0.19 V (vs. SCE). The detection scheme is based on (a) measurement of the activity of ethyl paraoxon-inhibited AChE, and (b) measurement of AChE activity after reactivation with pralidoxime (2-PAM). Compared to the conventional methods using acetylthiocholine as a substrate, the dual method presented here provides data on the AChE activity after inhibition and subsequent reactivation, thereby yielding credible data on reactivated enzyme activity. The linear analytical range for AChE activity extends from 2.5 U L-1 to 250 kU center dot L-1, and the detection limit is 0.83 U L-1.