Azure A Mediated Polypyrrole‐Based Amperometric Nitrate Biosensor

The co‐entrapment of Aspergillus niger‐nitrate reductase (NaR) and azure A (AzA), as a redox mediator has been investigated for fabrication of a polypyrrole (PPy)‐based electron mediated amperometric nitrate biosensor. Optimum formation of PPy‐NaR‐AzA film was achieved in a monomer solution which contained 0.3 M Py, 0.1 M KCl, 1U/mL NaR, 50 µM AzA with an applied current density of 0.5 mA/cm2 for 240 s. The optimum applied potential of −0.25 V for amperometric detection of nitrate with PPy‐NaR‐AzA biosensor was 50, 100 and 150 mV more positive than those achieved with the use of safranin O (Saf), methyl viologen (MV) and bromophenol blue (BrB) or red (BrR), respectively, as mediators. Similar optimum applied of −0.25 V was achieved with thionin acetate (ThAc) which was in the same phenothiazine group of mediators as AzA. The electron mediated amperometric detection of nitrate with the PPy‐NaR‐AzA biosensor also gave the best performance with a sensitivity of 40.8 nA µM−1 cm−2, detection limit of 0.5 µM, a response time of 18–20 s and a linear concentration range of 20–500 µM.