| Abstract | Nickel oxide (NiO) thin films prepared by glancing angle deposition (GLAD) were investigated as electrodes for enzymatic electrochemical quantification of xanthine, a noted indicator of meat freshness. The large surface area of the macroporous GLAD NiO electrodes provided a suitable scaffold to successfully immobilize the enzyme xanthine oxidase (XO), and this XO immobilization was characterized by cyclic voltammetry, electrochemical impedance spectroscopy, and X-ray photoelectron spectroscopy. The XO-modified GLAD NiO electrodes electrochemically oxidize xanthine, with electron transfer from this adsorbed XO to the (Ni2+/Ni3+) redox species resulting in a strong amperometric response to xanthine in a reagent-free alkaline medium. Under optimal conditions, the fabricated xanthine biosensor exhibited a rapid response (∼7 s), wide dynamic range (0.1–650 μM), good reproducibility (relative standard deviation of ∼4%, n = 18), superior limit of detection (37 nM), and very high sensitivity (1.1 μA·μM⁻¹·cm⁻² in the low concentration range from 0.1–5 μM and 0.3 μA·μM⁻¹·cm⁻² in the higher concentration range from 5–650 μM). The biosensor was also evaluated against a selection of potential interferents commonly found in fish samples (hypoxanthine, uric acid, glucose, and sodium benzoate), demonstrating good selectivity toward xanthine. A low Michaelis–Menten constant (Km) of 0.4 mM for xanthine signifies the high affinity of the enzymatic sensor toward the target analyte. Measurements in real fish samples were also successfully performed, revealing strongly increased xanthine sensitivity in the presence of fish matrices (from 0.085 μA μM⁻¹ without fish extract to as much as 0.27 μA μM⁻¹). |
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