Ayaz_2023_Talanta_266_124962

A novel colorimetric biosensor for the sensitive and selective detection of an organophosphate pesticide, paraoxon ethyl (POE), was developed based on its inhibitory effect on the acetylcholine esterase (AChE) enzyme. The bis-neocuproine copper (II) complex ([Cu(Nc)(2)](2+)) known as the CUPRAC reagent, was used as a chromogenic oxidant in the AChE inhibition-based biosensors for the first time. To initiate the biosensor, an enzymatic reaction takes place between AChE and its substrate acetylthiocholine (ATCh). Then, enzymatically produced thiocholine (TCh) reacts with the light blue [Cu(Nc)(2)](2+) complex, resulting in the oxidation of TCh to its disulfide form. On the other hand, [Cu(Nc)(2)](2+) reduces to a yellow-orange cuprous complex ([Cu(Nc)(2)](+)) which gives maximum absorbance at 450 nm. However, the absorbance of [Cu(Nc)(2)](+) proportionally decreased with the addition of POE because the inhibition of AChE by the organophosphate pesticide reduced the amount of TCh that would give a colorimetric reaction with the CUPRAC reagent. Based on this strategy, the linear response range of a colorimetric biosensor was found to be between 0.15 and 1.25 microM with a detection limit of 0.045 microM. The fabricated biosensor enabled the selective determination of POE in the presence of some other pesticides and metal ions. The recovery results between 92% and 104% were obtained from water and soil samples spiked with POE, indicating that the determination of POE in real water and soil samples can be performed with this simple, accurate, sensitive, and low-cost colorimetric biosensor.