A Nitrite Sensor Based on a Highly Sensitive Nitrite Reductase Mediator-Coupled Amperometric Detection

Abstract

Highly sensitive nitrite sensors have been developed for the first time based on mediator-modified electrodes. Tetraheme cytochrome c nitrite reductase from Sulfurospirillum deleyianum and cytochrome cd(1) nitrite reductase from Paracoccus denitrificans are able to accept electrons from artificial electron donors, which simultaneously act as electron mediators between the enzyme and an amperometric electrode. In addition to methyl viologen, redox-active compounds such as phenazines (phenosafranin, safranin T, N-methylphenazinium, 1-methoxy-N-methylphenazinium) and triarylmethane redox dyes (bromphenol blue and red) were selected from a range of redox compounds exhibiting the most efficient performance for nitrite detection. After precipitation, the electron mediators were incorporated in a graphite electrode material. Enzyme immobilization is performed by entrapment in a poly(carbamoyl sulfonate) (PCS) hydrogel. Diffusion coefficients and apparent heterogeneous rate constants of the mediators as well as homogeneous rate constants of nitrite sensors were determined by chronoamperometry and cyclic voltammetry. The phenosafranin-modified electrode layered with the PCS hydrogel immobilization of tetraheme cytochrome c nitrite reductase yielded linear current responses up to 250 μM nitrite with a sensitivity of 446.5 mA M(-)(1) cm(-)(2). The detection limit of the enzymatic nitrite sensor was found to be 1 μM nitrite.

Sensing Nitrite through a Pseudoazurin–Nitrite Reductase Electron Transfer Relay

“…Though numerous organic mediators of NiR Paracoccus denitrificans have been reported in the past, [41] to our knowledge, this report constitutes the first use of an inorganic mediator and the first voltammetric study of this (green copper) NiR species. Enzyme mediation with [Ru(NH 3 ) 6 ] 3 + leads to well-behaved Michaelis-Menten kinetics over a 1-100 mm range, with faradaic current levelling off at nitrite levels above 500 mm (Figure 8 …”

“…These values are consistent with those reported previously from spectroscopic and voltammetric analyses. [41,44] At slow scan rates convective contributions to diffusive current become significant and plots of I p /I d versus 1/v 1/2 become nonlinear.…”

“…34 , a value that is in good agreement with previous determinations. [41] Interactions with Nitrite Reductase A significant perturbation of the pseudoazurin diffusive voltammetric signal is observed when nitrite reductase is added, in molar excess (0.5 mm), to the solution. Specifically, the quasi-reversible response becomes sigmoidal in appearance ( Figure 5 c)-an observation fully consistent with the occurrence of an electrocatalytic reaction with the enzyme (though the diffusive half wave potential is perturbed by interactions with the reductase, the accompanying changes in wave shape make this difficult to reliably quantify).…”

A Nitrite Biosensor Based on Coimmobilization of Nitrite Reductase and Viologen‐Modified Polysiloxane on Glassy Carbon Electrode

“…Strehlitz et al screened out a list of redox mediators for NiRs immobilized electrode [8], Abrǔ na et al reported a biosensor based on a fusion protein of NiR and maltose binding protein immobilized on electropolymerized film [9].…”

Electrochemical Determination of Nitrite Using a Gold Nanoparticles-modified Glassy Carbon Electrode Prepared by the Seed-mediated Growth Technique

“…The operational stability of Au/GCE in nitrite determination is much better than that of a metallophthalocyanines-modified electrode, 11 a carbon nanotube-modified electrode 35 or an enzyme-modified electrode. 19 Moreover, the long-term stability of Au/GCE was also studied by storing the electrode for 7 days at room temperature. Compared with the response for nitrite in the real wastewater sample measured at the first day, the current response was only decreased by 2.3% after 7 days, further revealing that this modified electrode has good long-term stability and repeatability for nitrite determination.…”