A sandwich structured SiO2/cytochrome c/SiO2 on a boron-doped diamond film electrode as an electrochemical nitrite biosensor

A novel sandwich structured SiO2 gel/cytochrome c (Cyt c)/SiO2 gel was designed and constructed on conductive boron-doped diamond (BDD) film substrate. A SiO2 gel membrane was first in situ deposited on the pretreated positive charged H-terminated BDD electrode with a simple and artful surface vapor sol–gel method. Cyt c was subsequently immobilized onto the SiO2 membranes by electrostatic attraction, followed by another SiO2 gel layer in situ depositing on it. The SiO2 interlayer was conceived to play an important role in the resultant sandwich structured SiO2/Cyt c/SiO2/BDD electrode as a selective “semi-open” medium, which guaranteed the immobilized Cyt c to maintain high stability and perform good electrochemistry and biocatalysis responses. The bioactivity of Cyt c was well protected and the immobilized biomolecule even didnʹt denature at extremely high or low pH condition. More attractively, Cyt c in the sandwich structured electrode could be further oxidized into highly reactive Cyt c π-cation by two-step electrochemical oxidation, which could oxidize NO2− into NO3− in the solution. A sensitive determination approach of nitrite was accordingly built up based on this biocatalytic oxidative interaction for the first time and a possible mechanism of the interaction was herein proposed.