Engineered Hierarchical CuO Nanoleaves Based Electrochemical Nonenzymatic Biosensor for Glucose Detection
Authors: Ahmad Rafiq , Khan Marya 
Published: July 2021

The blood glucose concentration is the basis of diabetes mellitus “chronic and metabolic disease” diagnosis. The elevated levels of blood sugar “glucose” lead to many health problems, such as chronic kidney failure, stroke, cardiovascular disease, eyes retina damage, and foot ulcers. Hence, early diagnosis is crucial to prevent and avoid life-threatening complications caused at abnormal glucose levels. In recent, different methods (such as electrochemical, colorimetric, piezoelectric, and thermoelectric based biosensors) have been utilized for glucose concentration detection. Among these methods, the electrochemical based biosensors were extensively employed; however, most of the biosensors were enzyme-based. The enzyme-modified electrodes suffered from some drawbacks, for instance, complicated immobilization procedures, high cost, instability, and low sensitivity. Therefore, it becomes crucial to develop novel electrode nanomaterials that work as electrocatalyst for glucose oxidation and also result in high sensitivity and stability. The performance of nonenzymatic glucose biosensors depends on the morphology of the electrode material. Hence, a variety of nanostructured nanomaterials are utilized to fabricate nonenzymatic biosensors with a high working electrode surface area.
To fabricate nonenzymatic biosensors for glucose detection, significant efforts have been made to synthesize nanomaterials and use them as an electrocatalyst, for example, metals, metal oxides, and their hybrid nanostructures16-18. Among different catalysts, nanostructures of copper oxide (CuO) have received considerable interest19, 20. CuO is the best candidate to fabricate electrochemical based nonenzymatic glucose biosensors, as CuO based biosensors directly oxidizes glucose on the working electrode surface. Moreover, CuO nanostructures possess advanced properties, which are beneficial for designing biosensors.
Research has been concentrated on the shape/dimensions controlling during synthesis of (nano)materials, which have better structural properties that results in enhanced electrochemical performance due to high specific surface area. Previously, a variety of morphologies of CuO nanomaterials have been produced (i.e. nanoparticles, nanowires, nanowhiskers, nanoneedles, nanorods, nanoshuttles, nanoribbons, and nanotubes) using solution-based approach, sonochemical deposition, vapor phase growth, high temperature synthesis, double-jet precipitation, micro-emulsion synthesis, etc. Among these synthesis methods, hydrothermal method presents an environmentally friendly, simple, cost-effective, high-yield, and low-temperature method to synthesize various CuO nanostructures.
Herein, we report hierarchical CuO nanoleaves synthesis in large-quantity by a low-temperature hydrothermal method. Hierarchical CuO nanoleaves were synthesized using a low-temperature (95 °C) hydrothermal method (Fig. 1a). For synthesis, 0.25g Cu(CO2CH3)2·H2O and 0.77g CTAB were added in 40 mL deionized (DI) water. Next, 4 mL of 50 mM NaOH was added in the above solution while stirring. Then, the above solution was poured into a refluxing pot on a heating mantle and refluxed at 95 °C for 5h. Finally, black colored precipitates were washed using methanol and DI water to remove impurities and dried at room temperature. FESEM images and EDS analysis of CuO nanoleaves are shown in Fig. 1b-d. The low- and high-magnification images confirm that the CuO nanostructures prepared bear nanoleaves like morphology, and the nanoleaves are uniformly grown in large quantity. EDS analysis of CuO nanoleaves shows that nanoleaves are made of copper (Cu) and oxide (O) elements only (Fig. 1d, inset). EDS spectra shows an additional peak of Si, which is originating from Si substrate used to spread CuO nanoleaves sample for FESEM and EDS analysis. To fabricate hierarchical CuO nanoleaves based nonenzymatic glucose biosensor; first, the slurry of engineered CuO nanoleaves was prepared after mixing with conducting butyl carbitol acetate binder (8:2 v/v ratio) (Fig. 1a). The prepared slurry (2-6 μL) was cast on cleaned glassy carbon electrode (GCE, 0.071 cm2) and kept for drying. Finally, Nafion (5 μL) was coated on the electrode (CuO nanoleaves/GCE) surface and kept for overnight at 4 °C.
The electrochemical behavior of fabricated biosensor towards glucose was analzed with cyclic voltammetry (CV) and amperometry (i-t) techniques. Owing to the high electroactive surface area, hierarchical CuO nanoleaves based nonenzymatic biosensor electrode shows enhanced electrochemical catalytic behavior for glucose electro-oxidation in 100 mM sodium hydroxide (NaOH) electrolyte. The nonenzymatic biosensor displays a high sensitivity (1467.32 mA/(mM cm2)), linear range (0.005-5.89 mM), and detection limit of 12 nM (S/N = 3). Moreover, biosensor displayed good selectivity, reproducibility, repeatability, and stability at room temperature over three-week storage period. Further, as-fabricated nonenzymatic glucose biosensors were employed for practical applications in human serum sample measurements. The obtained data were compared to the commercial biosensor, which demonstrates the practical usability of nonenzymatic glucose biosensors in real sample analysis. Therefore, we believe our engineered CuO nanoleaves based nonenzymatic biosensor can pave the way to detect glucose in low glucose level samples (i.e., saliva, tear, sweat).

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Figure 1. (a) Schematic representation of CuO nanoleaves synthesis, (b-d) FESEM images and EDS spectra (inset d) of engineered hierarchical CuO nanoleaves.

Figure 1