A nitrite biosensor based on co-immobilization of nitrite reductase and viologen-modified chitosan on a glassy carbon electrode.
An electrochemical nitrite biosensor based on co-immobilization of copper-containing nitrite reductase (Cu-NiR, from Rhodopseudomonas sphaeroides forma sp. denitrificans) and viologen-modified chitosan (CHIT-V) on a glassy carbon electrode (GCE) is presented. Electron transfer (ET) between a conventional GCE and immobilized Cu-NiR was mediated by the co-immobilized CHIT-V. Redox-active viologen was covalently linked to a chitosan backbone, and the thus produced CHIT-V was co-immobilized with Cu-NiR on the GCE surface by drop-coating of hydrophilic polyurethane (HPU). The electrode responded to nitrite with a limit of detection (LOD) of 40 nM (S/N = 3). The sensitivity, linear response range, and response time (t(90%)) were 14.9 nA/μM, 0.04-11 μM (r(2) = 0.999) and 15 s, respectively. The corresponding Lineweaver-Burk plot showed that the apparent Michaelis-Menten constant (K(M) (app)) was 65 μM. Storage stability of the biosensor (retaining 80% of initial activity) was 65 days under ambient air and room temperature storage conditions. Reproducibility of the sensor showed a relative standard deviation (RSD) of 2.8% (n = 5) for detection of 1 μM of nitrite. An interference study showed that anions commonly found in water samples such as chlorate, chloride, sulfate and sulfite did not interfere with the nitrite detection. However, nitrate interfered with a relative sensitivity of 64% and this interference effect was due to the intrinsic character of the NiR employed in this study.
Department of Chemistry and Interdisciplinary Program of Integrated Biotechnology, Sogang University, Seoul, 121-742, Korea. quande@sogang.ac.kr
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D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D002244:Carbon / Q000737:chemistry; D002384:Catalysis; D048271:Chitosan / Q000737:chemistry; D055664:Electrochemical Techniques / Q000295:instrumentation / Q000379:methods; D004566:Electrodes; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism; D005898:Glass / Q000737:chemistry; D008954:Models, Biological; D009572:Nitrite Reductases / Q000737:chemistry / Q000378:metabolism*; D009573:Nitrites / Q000032:analysis* / Q000378:metabolism; D012242:Rhodobacter sphaeroides / Q000201:enzymology; D014755:Viologens / Q000032:analysis / Q000737:chemistry* / Q000494:pharmacology
D004800:Enzymes, Immobilized; D009573:Nitrites; D014755:Viologens; D002244:Carbon; D048271:Chitosan; D009572:Nitrite Reductases; C052695:nitrite reductase, copper-containing