Third-generation electrochemical biosensor based on nitric oxide reductase immobilized in a multiwalled carbon nanotubes/1-n-butyl-3-methylimidazolium tetrafluoroborate nanocomposite for nitric oxide detection

Abstract

Nitric oxide (NO) has a crucial role in signaling and cellular physiology in humans. Herein, a novel third-generation biosensor based on the Marinobacter hydrocarbonoclasticus metalloenzyme (nitric oxide reductase (NOR)), responsible for the NO reduction in the denitrifying processes, was developed through the direct adsorption of a new nanocomposite (multiwalled carbon nanotubes (MWCNTs)/1-nbutyl-3-methylimidazolium tetrafluoroborate (BMIMBF4)/NOR) onto a pyrolytic graphite electrode (PGE) surface. The NOR direct electron transfer behavior (formal potential of-0.255 ± 0.003 V vs. Ag/AgCl) and electrocatalysis towards NO reduction (−0.68 ± 0.03 V vs. Ag/AgCl) of the PGE/[MWCNTs/BMIMBF4/NOR] biosensor were investigated in phosphate buffer at pH 6.0. Large enzyme loading (2.04 × 10 −10 mol/cm 2), acceptable electron transfer rate between NOR and the PGE surface (ks = 0.35 s-1), and high affinity for NO (Km = 2.17 μmol L-1) were observed with this biosensor composition. A linear response to NO concentration (0.23-4.76 μmol L-1) was perceived with high sensitivity (0.429 μA/μmolL-1), a detection limit of 0.07 μmol L-1 , appropriate repeatability (9.1% relative standard deviations (RSD)), reproducibility (6.0-11% RSD) and 80-102% recoveries. The biosensor was stable during 1 month retaining 79-116% of its initial response. These data confirmed that NOR incorporated in the MWCNTs/BMIMBF4 nanocomposite can efficiently maintain its bioactivity paving a new and effective way for NO biosensing.

S-Layer Protein Coated Carbon Nanotubes

“…We will make use of this knowledge and specifically bind biomolecules on native and genetically functionalized S-layer fusion protein coated MWNTs (Figure 7a). CNTs and graphene have already shown great potential in the development of biosensors due to their huge surface area, great electron transfer rate, good electrical conductivity, and ability to immobilize biologically functional molecules, such as enzymes, aptamers, or receptors [12][13][14][15][16][17][18][19]. Concerning S-layers, a considerable amount of knowledge has accumulated concerning the fabrication of amperometric sensors for glucose [64] or sucrose [65] or of optical sensors for glucose too [66].…”

“…Proteins bound to the surface of CNTs are preferred in life-sciences because they provide a better biocompatibility and offer functional groups that may either be used for binding additional molecules in biosensor applications [12][13][14][15][16][17][18][19] or enable further chemical modifications, e.g., for the delivery of drugs, DNA and genes [20,21]. Nevertheless, although several proteins, such as bovine serum albumin (BSA), have already been successfully attached to CNTs through various physical or chemical methods, high resolution microscopical studies have demonstrated that their arrangement and density on the CNT surface and consequently the availability of functional groups varies considerably [5].…”

Application of Nanostructured Carbon-Based Electrochemical (Bio)Sensors for Screening of Emerging Pharmaceutical Pollutants in Waters and Aquatic Species: A Review

“…Additionally, CNT surfaces are easily modified with different functional groups which facilitates the attachment of biological entities to develop biosensors. Therefore, all these characteristics have been extensively used in the development of electrochemical (bio)sensors [ 46 , 99 , 100 , 101 , 102 , 103 ].…”

Progress in electrochemical detection of neurotransmitters using carbon nanotubes/nanocomposite based materials: A chronological review

“…Going a step further in the detection of NO, a third‐generation electrochemical biosensor based on NOR immobilized on a MWCNT‐BMIMBF4 composite was designed by Gomes et al. [ 252 ] in 2019. BMIMBF4 represents 1‐n‐butyl‐3‐methylimidazolium tetrafluoroborate which is an IL which combined with MWCNT and NOR to obtain the MWCNT/BMIMBF4 composite before subsequent immobilization of the MWCNT/BMIMBF4/NOR on PGE to get the PGE/MWCNT/BMIMBF4/NOR.…”

“…Nafion/MWCNTs-CS-AuNPs AP 7.6 0.019-54 [244] CAS/SOD/MP/MWCNT-PTTCA/ AuNPs/GCE AP 4.3 1-40 [246] 2011 GCE/MWCNT-CS-NiO AP 20 0.08-117 [247] MWCNT-CoTPPS/GCE CA 6600 6.6-13000 [ [252] current response in the presence of malathion using CV gave a LOD of 0.6 ng mL −1 . Another AChE modified MWCNT electrode was fabricated by Khan and Ghani (2012) for the determination of ACh and its metabolite simultaneously.…”