NiCo2O4 Nano-/Microstructures as High-Performance Biosensors: A Review

Highlights Various synthetic methods for the synthesis of NiCo2O4 nano-/microstructures in bare, doped, and composite/hybrid forms are reviewed. Currents status and development prospects of NiCo2O4 nano-/microstructure-based electrochemical biosensors for bioanalytes such as glucose, urea, and H2O2, along with condition governing the electrochemical biosensor parameters, are summarized. Also provide an insight into the key challenges and future perspectives about point-of-care monitoring of bioanalytes using NiCo2O4 nano-/microstructure-based biosensors.

Facile, low-cost and eco-friendly synthetic methods lead to varieties of low dimensional nano-/micro-structured morphologies with excellent porosity and specifically large surface area, opportunities to synthesize composite/hybrid and ease of electrode fabrications for end-user applications. Spinel NiCo 2 O 4 is a p-type semiconductor in which Ni occupies octahedral sites while Co is distributed in both octahedral and tetrahedral sites [13] (Fig. 1a, b). It shows a face-centered cubic arrangement and belongs to Fd3m space group with lattice constant a o = 8.269 Å [40].
Electrochemical sensing through miniaturized sensors based on nano-/micro-structured materials has taken over the conventional, expensive, laborious sensing techniques like lateral flow immunoassay, liquid chromatography, capillary electrophoresis, enzyme-linked immunosorbent assay, chemiluminescence, sequential injection analysis, gas chromatography-mass spectrometry and fluorescent methods [43][44][45][46][47][48]. Electrochemical biosensors can be categorized into amperometric and potentiometric sensors [49]. The amperometric biosensing involves a change in current response due to electrochemical redox reactions of the analytes when a potential is applied between the working and reference electrodes while the potentiometric biosensing makes use of ion-selective electrodes to transduce the biological reactions into a measurable electrical signal [43,50].
Among the main classes of biosensors, the non-enzymatic biosensor is considered to be better, faster, and more convenient as compared to an enzymatic biosensor that involves complicated and multi-step enzyme immobilization processes and high specificity of the enzymes. Also, due to pH and temperature sensitiveness, the enzyme-based biosensors are highly unstable as enzymes undergo denaturation leading to biological inactivity beyond physiological conditions [51][52][53]. Nanomaterials not only provide high-density catalytic sites for the electro-oxidation or electro-reduction in the biomarkers but also provide large surface area for adsorption of biomarkers and facilitate an appropriate path for electron transport for electrochemical activity [54][55][56]. Since the crucial part in electrochemical biosensors is the modified electrode, much attention has been devoted to modulate the electrocatalytic behavior of the NiCo 2 O 4 as electron mediator by engineering its composition, structure, specific surface area, and redox properties.
To date, many reviews have been reported for the applications of NiCo 2 O 4 nano-/micro-structured materials including Li-ion batteries [10], supercapacitors [11,57], fuel cells [58], and electro-catalyst for oxygen reduction, oxygen and hydrogen evolution reactions [59,60]. The applications of the NiCo 2 O 4 -based non-enzymatic biosensors are aimed not only at the extension of the spectrum of target bioanalytes but also at the improvement in the biosensor performance in terms of sensitivity, selectivity, detection limits, long-term stability as well as reusability. Many new synthetic strategies and techniques have been developed for the fabrication of NiCo 2 O 4 -based non-enzymatic biosensors, but they are rarely summarized. Hence, it is an appropriate time to go through the periodical progress of NiCo 2 O 4 -based non-enzymatic biosensors. This review covers the crystal structure of the spinel NiCo 2 O 4 , various synthetic strategies employed for the synthesis of nano-/micro-structured NiCo 2 O 4 , electrochemical biosensing toward biomarkers such as glucose, H 2 O 2 , and urea, through the fabrication of modified electrodes. Various factors affecting the morphologies and biosensing parameters of the nano-/micro-structured NiCo 2 O 4 are also reviewed.

General Biosensing Mechanism
Two types of strategies are generally involved in the electrochemical biosensing of biomarkers, i.e., enzyme based and enzyme-free [61,62]. An enzymatic biosensor operates on three main components which include sensitive recognition element, signal transducer element, and data evaluation component [63][64][65][66]. Enzymes, antibodies, and nucleic acid are generally used as recognition components. Glucose oxidase and glucose dehydrogenase for glucose [67,68], horseradish peroxidase for H 2 O 2 [69], urease for urea [70], laccase and polyphenol oxidase for rutin [71], tryptophan oxidase for tryptophan [72], etc. act as sensitive recognition elements. The function of the signal transducer is to convert chemical changes into detectable and readable electronic signals which are finally transferred to the data evaluation component. Recent developments in the field of nanotechnology and nanoscience reveal the excellent efficiencies of the nanostructured materials as signal transducers. Biosensors based on nanostructured materials as artificial bioreceptors are used for early detection and diagnosis of diseases through the estimation of the levels of biomarkers [73][74][75]. The signal transducer behavior of the nanomaterials mainly depends upon the electrochemical redox properties, surfaceto-volume ratio, crystal structure and phase, morphology, and the presence of some other conducting matrices along with the nanostructured materials [76][77][78]. In contrast, Reproduced with permission from Ref. [42]. Copyright © 2013 American Chemical Society in enzyme-free biosensors, nanostructured materials are used as signal transducers as well as sensitive recognition elements.
Electrochemical biosensors are mainly based on the output electrical signals changes incurred from either the oxidation or the reduction of the target bioanalyte on the surface of the transducer (Fig. 2) [79][80][81]. These redox reactions are catalyzed by signal transducer enzymes and nanostructured materials in enzyme-based and enzyme-free biosensors, respectively. The strength of the electrical signals is significantly affected by the concentrations of target bioanalytes, temperature, pH, and the presence of the interfering species [82][83][84][85].

Hydrothermal/Solvothermal Method
Hydrothermal synthesis involves heterogeneous reactions in an aqueous medium within a temperature range of 100-200 °C and high pressure. To achieve these conditions, the reaction is usually carried out in Teflon-lined sealed steel autoclaves. Alkali metal hydroxide or NH 3 is added to convert the precursor metal salts into their respective hydroxides at basic pH conditions [86,87]. An initial nucleation phase is followed by the directed crystal growth along appropriate crystal planes. The morphology, surface, and the structural features of the materials synthesized through hydrothermal method depend upon the conditions like temperature, pH of the solution, concentration of the precursor, nature of the solvent, and the presence of the templates [88]. NiCo 2 O 4 nano-/microstructures of various shapes and morphologies have been prepared hydrothermally. Nano-/micro-structured NiCo 2 O 4 of morphologies such as urchin shaped [89], corallike [90], core-ring-structured nanoplatelets [91], porous coral-like nanospheres [36], hollow nanospheres [92], nanospheres [93], urchin-like spheres [94], mesoporous nanoparticles [95], mesoporous nanoneedles [96,97], 3D network-like mesoporous nanostructures [98], 3D hierarchical tremella-like, flower-like, urchin-like and pine needle-like [99], nanoflakes [100], nanowalls [101], etc. are reported. Ni and Co precursor salt solutions with molar atomic ratio of 1:2 are taken during hydrothermal growth since Ni and
In the hydrothermal growth, the temperature is also a key factor in controlling the morphology of the nanostructures. Urchin-and sheaf-like NiCo 2 O 4 nanostructures were synthesized by Umeshbabu et al. [104] using CTAB as a surfactant under hydrothermal conditions at 120 °C and 200 °C temperatures, respectively. Different morphologies were attributed to different degrees of crystal splitting and anisotropic crystal growth at different growth temperatures [110]. Further, the temperature also affects the magnitude of the van der Waals forces, hydrogen bonding, hydrophobic attraction, crystal field attraction, and intrinsic crystal contraction which subsequently control the Ostwald ripening process [111,112].  (1) Even ethanol as the solvent can also initiate the formation of metal carbonate hydroxides. Two-dimensional porous NiCo 2 O 4 nanodisks were synthesized by a low-temperature hydrothermal method by Jain et al. [114] (Eqs. 8,9). Figure 4 proposes the initial formation of Ni 2 (CO 3 )(OH) 2 and Co 2 (CO 3 )(OH) 2 . Subsequent hydrothermal treatment in basic medium followed by calcination at 500 o C formed two-dimensional porous NiCo 2 O 4 nanodisks.
The nature of alkali source, capping agent, and other additives significantly affects the morphology of the NiCo 2 O 4 nanostructures. Wang et al. [99] reported tremella-like NiCo 2 O 4 nanostructures in the presence of HMTA, which transformed into flower-like nanostructures when NH 4 F was also added along with HMTA. However, when HMTA was replaced with urea, urchin-like and pine needle-like NiCo 2 O 4 nanostructures were formed, respectively, in the absence and presence of NH 4 F additive [99]. HMTA is hydrolyzed to produce NH 3 which finally produces OH − ions as stated earlier in this section (Eq. 10). (8) It was suggested that the F − ions released from NH 4 F stimulate the initially formed nanosheets and nanoneedles to produce more active sites to further activate nucleation, more mass loading of active material per unit area, firm binding between the active material, and hence more crystal growth [115][116][117]. The possible set of reactions elaborating the role of F − ions released from NH 4 F is shown as follows [118] (Eqs. [11][12][13].
Further, different concentrations of the NH 4 F also stimulated the initially formed nanostructures to acquire more versatile morphologies. For 3, 9, and 12 mmol concentrations of NH 4 F, various morphologies of the NiCo 2 O 4 nanostructures are shown in Fig. 5. With an increase in concentration from 3 to 9 mmol, aggregation of the neighboring nanosheets occured. Further increase in concentration to 12 mmol, rhombus-shaped architectures were formed [117].
Deng et al. [119] prepared novel urchin-like peapoded NiCo 2 O 4 @C nanostructures as a bifunctional catalyst for the water-splitting reaction. A three-phase process was proposed which included the initial hydrothermal synthesis of nanoneedles self-assembled microsphere followed by coating with polymerized glucose as green carbon source onto NiCo 2 O 4 microsphere. The final stage was the calcination   Fig. 6.
Still another way of engineering the morphology, porosity, and growth of the crystals along the particular oriented crystal planes of the nanomaterials, is the use of non-aqueous solvents. The modified method is named as solvothermal instead of hydrothermal. Solvents with different solubilities and polarities can significantly affect the degree of supersaturation, the diffusion rates of the chemical species to the surface of the growing crystals, the interfacial surface energy, etc. [120,121]. Fu et al. [122] synthesized 1D porous NiCo 2 O 4 microrods (using metal acetate salts) (Fig. 7a) and microspheres (using metal nitrate salts) (Fig. 7b) in aqueous and isopropanol media, respectively, under similar conditions of temperature and reaction time. of closely packed microplates aligned along one direction with sizes of 3-5 μm were formed (Fig. 7c). In pure ethanol microspheres composed of nanosheets, interweave together with an average diameter of 8 μm were formed (Fig. 7d). However, in diethylene glycol, irregular aggregates with sheet-like structures were synthesized (Fig. 7e). Wang et al. [123] in an interesting stepwise hydrothermal growth synthesized layers of NiCo 2 O 4 nanosheets on the surface of NiCo 2 O 4 nanocones precursor to give highly ordered 3D hierarchical NiCo 2 O 4 @NiCo 2 O 4 core-shell nanocone arrays on nickel foams (Fig. 8). Different morphologies were engineered by controlling the reaction time and the temperature during stepwise hydrothermal growth. Further, NiCo 2 O 4 nanocones arrays on Ni foam were synthesized in the absence of HMTA while the NiCo 2 O 4 nanosheets growth on NiCo 2 O 4 nanocones was guided by the presence of HMTA.

Templated Solution Growth Method
The morphology, size, shape, and surface area of nanostructures can be designed through template-based synthesis to produce nanostructures with controlled physical, chemical, electrical, and electronic properties essential in notable applications and are also quite different from those of the bulk materials. Generally, three stages, viz., template preparation, directed synthesis of the desired material using the template, and the template removal, are described in the overall growth process of nanostructures [124]. The chemical nature, structure, concentration, and growth temperature are some of the important environmental factors affecting the growth of nanomaterials. Template-based methodologies are reported in the literature which govern the synthesis of NiCo 2 O 4 nanomaterials with versatile morphologies including nanospheres, hollow spheres, nanocages, hollow submicron spheres, hollow irregular octahedra-like cages, flower-like nanostructure, microspheres with highly ordered mesoporous structures, nanowires, etc. With the development of new methods for synthesizing mesoporous binary NiCo 2 O 4 metal oxides, the combination of template method with other methods such as hydrothermal/solvothermal, sol-gel has been widely used. In one such study, Ren et al. [125] prepared mesoporous NiCo 2 O 4 microspheres using a mesoporous silica (KIT-6) template. The KIT-6 template was added into the metal nitrate precursor solution prepared in ethanol. The schematic illustration of the formation of mesoporous NiCo 2 O 4 microspheres is shown in Fig. 9a. The high porosity of the synthesized mesospheres was ascertained by FESEM and TEM images (Fig. 9b, c). The template was finally removed by etching with 2 M NaOH solution [125].
Yuan et al. [126] utilized silica spheres as hard templates prepared by the modified Stöber method [127], for Further, the alkalinity of the medium results in the formation of -OH-Ni-OH-Co-OH-complex networks. This property has been explored for the synthesis of NiCo 2 O 4 nanostructures by Veeramani et al. [128]. FESEM images shown in Fig. 10 are demonstrating the effect of dopamine on the morphology of the NiCo 2 O 4 nanostructures.
Flower-like dopamine derived NiCo 2 O 4 nanostructures were formed. In another significant strategy, Xiong et al. [129] used mollusk shell-based macroporous carbon material (MSBPC), as a template to grow NiCo 2 O 4 nanowires hydrothermally (Fig. 11a, b). The MSBPC was obtained from mollusc shells   [130] reported the synthesis of composite C@NiCo 2 O 4 hollow microspheres via a two-step strategy of hard template-induced hydrothermal synthesis followed by calcination. SiO 2 @RF (resorcinol-formaldehyde resin, RF) sphere was used as a hard template, whereas HMTA was used as precipitant. The template SiO 2 @RF was synthesized via a one-pot sol-gel process under alkaline condition using an alcohol-water mixed solvent [131]. The SiO 2 core was removed by treating the prepared material with 2 M NaOH at room temperature for 12 h. The SiO 2 @RF template was having a core-shell structure with an average diameter of 350 nm (Fig. 11c). The NiCo 2 O 4 nanoflakes were grown and assembled on the carbon surface of the SiO 2 @RF spheres (Fig. 11d). Recently, novel micron-sized NiCo 2 O 4 pompon was prepared by templated growth using polyvinylpyrrolidone (PVP) non-ionic polymer and cationic surfactant CTAB as co-template [132]. Columbic and coordinative forces between template, co-template, and the metal ions help to form a stable "hairball" structure which finally was converted into a micron-sized pompon-like product on annealing (Fig. 11e). In contrast, in the absence of co-template CTAB, mesoporous NiCo 2 O 4 hollow submicron spheres with a uniform diameter of 400-500 nm were obtained through a soft template method assisted by PVP ( Fig. 11f). Further, in the absence of even PVP, solid submicron spheres were obtained [133].
Qi et al. [131] also used RF microspheres as templates for the synthesis of NiCo 2 O 4 hollow microspheres with tunable shell numbers and shell thickness. The shell numbers were controlled by adjusting the solvent ratio (DI water: ethylene glycol) and heating ramp rate, whereas the shell thickness and porosity were controlled by adjusting the metal ion concentrations (Fig. 12). For total molar concentrations of Ni 2+ and Co 2+ of 0.05 and 0.1 M, thin and thick shells, respectively, were formed. NiCo 2 O 4 hollow microspheres with double and triple shells were formed at a heating ramp rate of 2 and 5 °C min −1 , respectively, in EG as a solvent. With the increase in the ramp rate, the increased temperature gradient of the infused RF microspheres along the radial direction favors the separation of adjacent NiCo 2 O 4 layers and the infused RF cores, thereby transforming double shell to triple shells [134]. Furthermore, EG prevents the formation of the metal aqua ions, and thus, the penetration of the metal ions into RF microspheres is accelerated which is essential for the formation of multi-shell NiCo 2 O 4 hollow microspheres [135,136]. Additionally, the final calcination process also results in some adhesion force in the outward direction and the contraction force by decomposition of the inner core which segregates the outer NiCo 2 O 4 shell and the inner infused RF [131].
In addition to templates of organic origin, inorganic metal oxides have also been reported as template materials for the synthesis of NiCo 2 O 4 nano-/microarchitectures [137] Yang et al. [142] reported NiCo 2 O 4 hollow nanorods prepared by the sacrificial template-accelerated acid hydrolysis of ZnO (Eq. 16).

Sol-gel Method
The sol-gel process represents the chemical conversion of the liquid "sol" to the network "gel" phase, subsequently post-treatment into solid metal oxides with microcrystalline ultrafine particles. It is superior to other methods because it can better control the texture and surface properties of synthesized nanomaterials. The sol-gel method for the synthesis of nanomaterials is affected by numerous factors including pH, temperature, nature of solvent, growth time, agitations time, presence of capping agents, template, etc. With the consideration of these factors and potential applications, many protocols have been used to design materials of different sizes and features, including nano-, micro-, meso-, and macro-materials. To get excellent porosity and conductivity for potential electrochemical applications, the addition of polymers stuffs such as PVP [143], organic solvents/ additives like propionic acid [144], citric acid [145,146], N,N-dimethylformamide (DMF) [147], and epoxides like propylene oxide [148,149], during the post-annealing process is suggested. Significantly the additive/metal ion molar ratio is very important in controlling the pore size and pore volume. Traditional use of SiO 2 is avoided as its addition decreases the conductivity and limits the connection of the film with conducting substrate in thin film forms of NiCo 2 O 4 [143]. In a typical sol-gel method, the NiCo 2 O 4 spinel oxide was prepared by mixing appropriate amounts of metal salt precursors along with citric acid. The resulting solution was magnetically stirred at 80 °C for 2 h to get a gelatinous matrix. Finally, the matrix was calcined at 550 °C for 5 h to get the desired product [146]. Citric acid was also used as a chelating ligand for the synthesis of highly porous coral-like crystalline NiCo 2 O 4 nanoparticles with submicron sizes via a facile sol-gel method in H 2 O-DMF mixture as solvent [147]. Liu et al. prepared nanoporous NiCo 2 O 4 thin films deposited on ITO glass. The precursor solutions for NiCo 2 O 4 nanospheres were prepared via a sol-gel method in glacial acetic acid and ethanol as solvents, and ethylene glycol and CTAB were used as a viscosity modifier template, respectively [150]. Thus, the sol-gel process is a proven and important method for preparing NiCo 2 O 4 nanoparticles.

Co-precipitation Method
Better stoichiometric control and high purity of the metal oxide nanomaterials can be easily achieved through the coprecipitation method which involves simultaneous precipitation from a homogeneous solution of two or more cations. Simultaneous occurrence of nucleation, growth, coarsening, Ostwald ripening, and aggregation dramatically affect the size, morphology, and properties of the metal oxide nanoparticles. The technique has been applied for the synthesis of NiCo 2 O 4 nanomaterials. NiCo 2 O 4 hexagonal nanostructures were prepared by Bhagwan et al. [151] using Ni and Co chlorides and 6 M KOH as the precipitating agent. The schematic illustration for the formation of NiCo 2 O 4 hexagonal is shown in Fig. 13a. It was suggested that the strong alkaline environment in the growth solution caused nickel and cobalt ions to precipitate and nucleate together, forming nickel-cobalt hydroxide which was subsequently converted into NiCo 2 O 4 hexagonal after calcination at 300 °C. Liang et al. [152] reported hierarchical NiCo 2 O 4 nanosheets@halloysite nanotubes (Fig. 13b). The initial formation of NiCo precursor@halloysite nanotubes was assisted by HMTA and dehydrated citric acid trisodium salt.
A stepwise co-precipitation template free method was designed by Chen et al. [153] for the synthesis of hierarchical urchin-like NiCo 2 O 4 hollow nanospheres. Ureaassisted mesoporous urchin-like NiCo 2 O 4 nanostructures were prepared by Jadhav et al. [154] by an easy, viable, and cost-effective co-precipitation method. Yu et al. [155] explored the structure-stabilizing properties of PVP, which can bind the metal ions through electrostatic interaction with the -N and/or C=O functional groups, for the formation of Ni-Co precursor particles with tetragonal prismlike shapes by a modified coprecipitation method. The yolk-shell Ni-Co oxide nanoprisms with a highly porous interior core structure consisting of numerous polycrystalline primary particles were obtained finally after annealing. Other stabilizing and precipitating agents like ethylene glycol (EG) [156], urea [157], NaOH, NH 4 OH, NH 4 HCO 3 , H 2 C 2 O 4 [158,159] and NaHCO 3 [153] are reported in the literature. Organic stabilizers such as EG are supposed to form a protective layer around the particle surface through interactions with hydroxyl groups preventing the aggregation. Moreover, EG also acts as a bidentate chelating ligand for solvated metal ions [160]. Another important factor that controls the morphology, shape, and size of the nanoparticles is the pH of the reaction medium during   Fig. 13 a Schematic representation for the synthesis of hexagonal NiCo 2 O 4 nanosheets, Reproduced with permission from Ref. [151]. Copyright © 2019 Elsevier Ltd. and b hierarchical NiCo 2 O 4 nanosheets@halloysite nanotubes via co-precipitation method. Reproduced with permission from Ref. [152]. Copyright © 2014 American Chemical Society coprecipitation. Wan et al. [159] observed the change in morphology of the NiCo 2 O 4 precursors from the cubic to the fibrous along the axial direction. The fibrous morphology was maintained at a still higher pH value of 8.4; however, the aspect ratio was increased ( Fig. 14a-d). A dynamic equilibrium was suggested to exist between metal ammoniated complexes and the coprecipitation of Ni 2+ and Co 2+ as their oxalates.
The post-annealing temperature is also an important factor for controlling the morphology of the NiCo 2 O 4 spinel structures. The homogeneous dark blue-colored suspension which was obtained by mixing the metal nitrates and NaOH solution was initially evaporated under rotation and reduced pressure conditions by a cost-effective rotary evaporation method. Hexagonal column-like mesoporous loose architectures and hexagonal dense blocks were obtained at 200 and 400 °C calcination temperatures, respectively (Fig. 15) [161].

Electro-Deposition
Electro-deposition is considered a very useful, versatile, and flexible tool for the deposition of dendritic hierarchical structures, thin and thick films, nanosheet, nanofoil, nanotubes, nanowires, and many well-ordered transition metal oxides on conducting surfaces. Potentiostatic, galvanostatic, and pulse plating are the three main techniques employed for electro-deposition [162,163]. The basic principle of electrodeposition involves three steps, viz. preparation of a metal ions precursor solution, co-electro-deposition, and final thermal decomposition [164]. Recently, this technique has also been used for the preparation of NiCo 2 O 4 spinel structures for various applications, including supercapacitors, anode materials for Li-ion batteries, gas sensors, biosensors, etc. Wu et al. [165] deposited nanostructured cauliflower-like NiCo 2 O 4 film through galvanostatic electro-deposition combined with annealing treatment (Fig. 16). Galvanostatic electro-deposition was performed using a three-electrode compartment comprising a stainless steel disk as a working electrode. An Ag/AgCl saturated with KCl and a platinum plate were used as the reference and counter electrodes, respectively. Hydroxide-SiO 2 template transformed nanoflakes to cauliflower-like NiCo 2 O 4 nanoparticles. Under cathodic potential, the generated OH − ions catalyzed the sol-gel process for the formation of SiO 2 . The generated OH − ions facilitated the formation of Ni(OH) 2 and Co(OH) 2 .
Heat treatment of the deposited at 250 °C in air for 2 h converts the metal hydroxides into NiCo 2 O 4 films.
Wang et al. [166] reported the electro-deposition of the nickel/cobalt/zinc ternary alloy layer on ultrafine nickel wire. Removal of the zinc by dealloying with NaOH solution followed by oxidation at the atmospheric environment resulted in mesoporous NiCo 2 O 4 film on the surface of ultrafine nickel wire. Zhao et al. [167]  ions from the metal salts were reduced to NO − 2 and NH + 4 ions at the cathode. This reduction also resulted in the formation of OH − ions which combined with the Ni 2+ and Co 2+ to form amorphous binary metal hydroxide NiCo 2 (OH) 6 nanosheet networks [168]. Post-annealing transforms the NiCo 2 (OH) 6 into NiCo 2 O 4 nanosheet networks [57,169] (Eqs. [17][18][19][20]. The dissolution of the ions decreases near the electrode due to the formation of the OH − ions and an increase in pH (17) near the electrode is observed. Since the solubility constants of Ni(OH) 2 (8.2 × 10 −16 ) and Co(OH) 2/3 (2.5 × 10 −16 ) are very low and comparable, their simultaneous precipitations occur which finally gives NiCO 2 (OH) 6 [170,171]. Ramadoss et al. [169] electrodeposited highly porous and binderfree 3D flower-like NiCo 2 O 4 /Ni nanostructures on Ni wire and explored their supercapacitor applications (Fig. 17a) [181], and many more.

Combustion Method
Combustion synthesis, also referred to as self-propagating high-temperature synthesis is one of the most versatile, convinced, convenient, cost-effective, and fast method for the synthesis of nanomaterials. It involves a thermally induced redox reaction between precursor salt as oxidizers and an organic fuel [182][183][184]. Glucose, fructose, tartaric acid, sucrose, glycine, citric acid, hydrazine, urea, and oxalic acid are generally used as organic fuels. However, if metal oxalate or acetate salts are used, the combustion process can be directly conducted in the absence of fuel [185]. Byproduct gases like CO 2 , H 2 O, N 2 , oxides of N (NO x ) and S (SO x ), etc. are evolved during the combustion process [186]. The release of these gases promotes the expansion of the product and rapid fall in temperature after the reaction ceases. This provides a solid product with a high degree of porosity and good dispersibility [187]. As compared to solid-state combustion, liquid phase combustion synthesis has proved to be the most suitable one as oxidizers and fuel are well dissolved in aqueous or alcoholic solutions [188]. Ni(NO 3 ) 2 ·6H 2 O, Co(NO 3 ) 2 ·6H 2 O (in 1:2 molar ratio) as oxidizers and tartaric acid as fuel were dissolved in acidified 2-methoxy ethanol solution. The resulting solution was combusted at 250 °C for 1 h to prepare NiCo 2 O 4 nanoparticles [189]. Sucrose assisted combustion of the Ni and Co nitrates also resulted in NiCo 2 O 4 nanoparticles when the combustion process was carried out at 350 °C for 6 h [190]. The oxalate precursors were directly decomposed into NiCo 2 O 4 powders by heating in an air ambient atmosphere at 320 °C for 10 h [185]. Citric acid assisted combustion at 400 °C for 4 h resulted in highly porous NiCo 2 O 4 nanomaterials [191]. Urea-assisted combustion was processed at 400 °C for 2 h in ethyl acetate as a solvent [192]. In each case, a viscous gel is obtained initially by heating the reaction solution at low temperature followed by auto-ignition resulting in the formation of highly fluffy mass which is finally calcined at high temperature. Direct calcination of the metal nitrate salts in the presence of alkalis without any fuel has also been reported for the synthesis of the NiCo 2 O 4 nanorods [41].
Though it is a fast and low-cost method for the synthesis of NiCo 2 O 4 powders, it suffers from some major drawbacks including less control over morphological uniformity and particle size, the simultaneous formation of a variety of crystal phases, the formation of highly agglomerated structures, complex and uncertain growth mechanism, and critically very low possibilities of formation of a versatile and wide range of morphological structures as those of in hydrothermal and other solution methods.

Electro-Spinning Method
Many electrospun carbonaceous materials such as carbon nanofibers, single-walled carbon nanotubes, multi-walled carbon nanotubes, etc. prepared from oxidation and carbonization of polymers like PVP, PAN, PVA have been used as templates for the growth and deposition of NiCo 2 O 4 nanostructures with versatile morphologies. In one synthetic way, there is simultaneous growth of NiCo 2 O 4 nanostructures and electro-spinning of template material [193,194]. In another strategy, NiCo 2 O 4 nanostructures are grown through other synthetic methods like hydrothermal, sol-gel, coprecipitation, etc. on pre-electrospun carbonaceous templates [39]. Electro-spinning setup comprises a high-voltage system, spinneret, and collector which results in the formation of continuous nanofibers with diameters ranging from nanometer to micrometer [195][196][197]. The deposition of NiCo 2 O 4 nanostructures on these carbonaceous materials not only improves the electrical and electronic properties but also enhances the thermal, mechanical and chemical stabilities which are the important prerequisite characteristics for the biosensing and other applications. The composition of the precursor solution, presence of additives like templates and capping agents, modification in the electro-spinning setup, post-annealing, electrospun voltage are some of the major factors which control the thickness, porosity, and morphology of the deposited NiCo 2 O 4 films. Lai et al. [198] through electro-spinning, co-deposition, redox deposition fabricated NiCo 2 O 4 -doped carbon nanofiber@MnO 2 nanosheet and nanorod hybrid membranes. Busacca et al. [199] prepared NiCo 2 O 4 /carbon nanofibers composites and investigated their oxygen evolution reaction in alkaline electrolyte. Metal acetate salt precursor in a molar ratio 1:2 was mixed in PAN (as carbon source) and DMF. The electrospun layer was thermally oxidized at 270 °C in air for 30 min followed by subsequent carbonization at 900 °C for 1 h under a helium gas flow. Li et al. [193] [194] synthesized spinel NiCo 2 O 4 nanofibers with diameters of 50-100 nm through electro-spinning of the PVA/cobalt acetate/nickel acetate composite precursor followed by annealing at high temperatures ranging from 400 to 800 °C. Liu et al. [39] demonstrated the surfactant-assisted hydrothermal uniform growth NiCo 2 O 4 nanoneedle on electrospun carbon nanofiber (ECF) and explored their glucose sensing properties non-enzymatically. ECF film was prepared through initial electro-spinning and subsequent oxidation and carbonization of PAN (Fig. 18a-c). Xu et al. [200] instead of PAN used PVP as a carbon source to produce NiCo 2 O 4 nanotubes. These nanotubes were used as scaffolds for hydrothermal growth of MnO 2 nanosheets for the additional improvement in electronic conductivity and electrochemical activity for supercapacitor applications (Fig. 18d-f). Copolymers like poly (acrylonitrile-co-methylhydrogen itaconate) [201] and biobased polymer composites such as PAN/lignin [202] are also reported in the literature for the formation of flexible carbon nanofibers. The hollow carbon nanofibers were used as a template for the hydrothermal growth of NiCo 2 O 4 with uniform dandelion-like morphology consisting of densely grown nanoneedle (Fig. 18g, h) [203]. The above discussion thus reveals that the proper combination and the composition of the polymers can result in the formation of carbonaceous materials with versatile structural features with high surface area necessary for potential applications.

Microwave-Assisted Method
Microwaves are the electromagnetic radiations having a frequency range between 300 MHz and 300 GHz and a wavelength range of 1 m-1 mm. Microwave-assisted synthesis of nano-/microstructures is superior to the conventional methods described above because it requires a very short reaction duration, is energy efficiency, cost-effectiveness, and gives an excellent yield of highly porous materials. Microwaves result in volumetric heating as they can penetrate throughout the volume of reactants [204]. This volumetric heating is caused by various types of polarization in the medium, including electron polarization, atomic polarization, directional polarization, and space charge polarization [205]. To obtain better morphological results, microwave-assisted synthesis of nanomaterials is usually combined with other synthetic methods such as sol-gel, co-precipitation, and hydro/solvothermal, etc. Recently, the improvement in the hydrothermal method in harmony with microwave assistance has been studied to synthesize NiCo 2 O 4 nano-/microstructures. Other ways of engineering the structural aspects of the NiCo 2 O 4 are the use of a template, capping agents, organic solvents, ionic solvents, and addition of other growth additives. The microwave-assisted hydrothermal method was applied by Zhang et al. [206] to prepare NiCo 2 O 4 doubleshelled hollow spheres with an outer and inner shell thickness of ~ 20 and ~ 70 nm, respectively. A mixture of isopropanol and glycerol was used to prepare a reaction solution (Fig. 19a). Glycerol molecules were supposed to form a selfassembled quasi-emulsions in isopropanol that serve as a soft template for the growth of Ni-Co double hydroxides. In the absence of glycerol, solid microspheres with diameters of ~ 1 µm were formed, demonstrating the templated role of glycerol in the synthesis of a double-shelled hollow nanostructure (Fig. 19b-d). In the presence of microwaves, the reaction mixture is heated due to dielectric loss, which significantly accelerates the reaction kinetics. Additionally, the presence of microwaves improves uniformity in terms of dispersion and size distributions.
Shanmugavani et al. [207] analyzed the effect of reaction times on the morphology of the NiCo 2 O 4 /NiO nanocomposites. The reaction was carried out in the presence of oxalic acid at an operating frequency of 2.45 GHz and 800 W output power. It was proposed that the initially formed nanoparticles are converted into bundled-like structures as the reaction time was increased. Recently, Sun et al. [103] reported novel porous nanoscale NiO/NiCo 2 O 4 heterostructure through two-stage calcination of nickel-cobalt bimetallic hydroxide precursors (NiCo precursors) which were

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initially synthesized using a microwave-assisted hydrothermal method in the presence of HMTA and NH 4 F. Notably, F − ions were supposed to act as functional template agents. Prolonged irradiation significantly affects the morphology of NiCo 2 O 4 materials. When the irradiation time was increased from 5 to 40 min, the incompletely self-assembled and non-uniform 2D nanosheets are converted into more optimized and thickened 3D frameworks with large open spaces (Fig. 20a-i). Nakate et al. [208] prepared nanocrystalline NiCo 2 O 4 nanoplates in the surfactant-free environment using metal chloride salts precursors through microwave irradiation. Gu et al. [209] reported 3D nanosphere-like NiCo 2 O 4 nanostructure composed of intertwined 2D ultrathin mesoporous nanosheets having large specific surface area 146.5 m 2 g −1 .
The reaction solution was exposed to microwaves (power 560 W) for 6 min. Su et al. [210] reported highly crystalline NiCo 2 O 4 supported on carbon black via a simple, one step intermittent microwave heating method avoiding the calcination process. However, in a contrary study, Tao et al. [211] analyzed the effect of post-annealing temperature on the morphologies of the NiCo 2 O 4 . Ni-Co double hydroxide was initially prepared through a microwave-assisted method using a tertbutanol solution (98%). Flower-shaped morphology of the Ni-Co double hydroxide was completely converted into unique coral-like morphology on calcination. As the post-annealing temperature was increased from 400 to 700 °C, individual ultrathin nanosheets shrink to smaller nano-sized crystal grains which finally self-assembled to form coral-like NiCo 2 [212], and non-ionic glucose-based polymeric surfactant, β-C 10 Alkyl Poly Glucoside [213] are also reported in the literature for the synthesis of NiCo 2 O 4 architectures with versatile morphologies.

Spray Pyrolysis Method
In spray pyrolysis technique, an aerosol of various precursor components is prepared in suitable solvent and is sprayed on the substrate. After that, sequential evaporation of the solvent from the surface of the substrate, heating to precipitate out the solute, high-temperature annealing, formation of microporous particles, and finally, sintering of solid particles is carried out [214]. NiCo 2 O 4 nanostructures with morphologies hollow nanosphere [215], hollow microspheres [216], dried plum-like particles [217], yolk-shell microspheres [218], nanoaggregates [219], thin films with uniform particle distribution size 20-30 nm [220], etc. are reported (Fig. 21a-e).
Similar to the electro-spinning method, carbonaceous materials such as reduced graphene oxide, carbon nanotubes, carbon nanofibers are also mixed in the precursor solution to improve the electrochemical properties of NiCo 2 O 4 . Park et al. [221] synthesized three-dimensional macroporous multi-walled carbon nanotubes microspheres densely loaded with NiCo 2 O 4 hollow nanospheres via spray pyrolysis process. The schematic illustration depicting the formation mechanism is shown in Fig. 22a. The polystyrene nanobeads added in the solution improved the structural uniformity and the dispersion of CNT microspheres. The similarity in the atomic radii of the Ni and Co ions resulted in the Kirkendall diffusion into the outer surface of the where they were oxidized to form NiCo 2 O 4 (Fig. 22b).

Glucose Biosensors
Non-enzymatic glucose sensing is considered to be a better, fast, and convenient way as compared to the enzymatic method since the later is a complicated and multi-step process involving immobilization of enzyme bioreceptor such as glucose oxidase, glucose dehydrogenase, and quinoprotein glucose dehydrogenase onto the electrode surface [51,52] [216]. Copyright © 2019 Elsevier Ltd and Techna Group S.r.l. c yolk-shell microspheres. Reproduced with permission from Ref. [218]. Copyright © 2017 Elsevier Ltd. d nanoaggregates. Reproduced with permission from Ref. [219]. Copyright © 2015 Elsevier Inc. and e thin films with uniform particle distribution size 20-30 nm. Reproduced with permission from Ref. [220]. Copyright © 2016 Elsevier Ltd.
Since the rates of oxidation of Ni 2+ and Co 2+ ions on the electrode surface during anodic scan determine the rate of sensing of glucose, NiCo 2 O 4 nano-/microstructures with versatile morphologies having large specific surface area, permeability, and most importantly short electron and ion diffusion pathways are synthesized. Ni 3+ and Co 3+ ions are reduced back to Ni 2+ and Co 2+ ions by the electrons lost by the oxidation of glucose to gluconolactone. According to Glucose C 6 H 12 O 6 → Gluconolactone C 6 H 10 O 6 + 2H + + 2e − Hussain et al. [225], H 2 O 2 is formed as one of the products along with gluconolactone if the electrochemical sensing is performed in the presence of oxygen. Glucose undergoes a spontaneous reaction with water and O 2 to form gluconolactone which is further oxidized into gluconic acid (Eqs. 27, 28). In a slightly basic medium (pH = 7.4), gluconic acid ionizes to gluconate ions which act as mobile charge carriers on the surface of the NiCo 2 O 4 nanostructures producing a strong electrical signal (Eq. 29). Elakkiya et al. [226] reported highly porous flower-like NiCo 2 O 4 nanostructures synthesized via a facile hydrothermal method for excellent electrocatalytic activity in alkaline electrolyte for the oxidation of glucose and lactic acid.

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The binary spinel NiCo 2 O 4 architecture exhibits better intrinsic electronic conductivity as compared to pure NiO and Co 3 O 4 which is attributed to the doping of Ni 3+ ions in the octahedral sites of the Co 3 O 4 crystal lattice which accelerates the electron hopping process [227]. Huang [229] (Fig. 24a, b). The fabricated glucose biosensor exhibited a high sensitivity of 1748.58 μA mM −1 cm −2 and a very low detection limit of 0.152 μM. MoS 2 nanosheets have also been reported as support material for the fabrication of NiCo 2 O 4 /MoS 2 nanocomposites through a simple ionothermal method in deep eutectic solvent (choline chloride (ChCl)-urea mixture) [230]. Deep eutectic solvents consist of simple eutecticbased ionic liquids prepared by eutectic mixing of ChCl and some hydrogen bond donors like acids, amides, alcohols, etc. [231]. These solvents have excellent thermal stability, high surface tensions, negligible vapor pressure, and most importantly biodegradability [232][233][234][235][236]. The NiCo 2 O 4 -MoS 2 /chitosan/GCE-modified electrode was used as an electrochemical sensor for glucose in red wine and honey [230].
Analysis of non-enzymatic glucose sensing properties of NiCo 2 O 4 nanosheets showed linear response with respect to the change in glucose concentration varying from 5 to 65 μM. The high sensitivity of 6.69 μA μM −1 cm −2 with a LOD value of 0.38 μM and liquid of quantification of 1.27 μM was observed. During CV measurements, scan rates increased the oxidation and reduction peak currents as well as peak-to-peak separations [224]. The electrochemical kinetics of the NiCo 2 O 4 hollow nanorods grown on stainless steel via a sacrificial template showed similar trends during glucose sensing in 0.1 M NaOH solution with scan rates ranging from 5 to 100 mV s −1 (Fig. 25a). Amperometric studies revealed a steady-state current optimization within 2 s of glucose addition. Calculated sensitivity, linear detection range, and detection limit were 1685.1 µA mM −1 cm −2 , 0.0003-1.0 mM, and 0.16 µM (S/N = 3), respectively (Fig. 25b) [142]. Cui et al. [237] prepared rectangular flake-like mesoporous NiCo 2 O 4 via a facile hydrothermal method and observed glucose  [238]. The microwave treatment completely removed the water and made the material highly porous for exhibiting excellent biosensing applications. One-dimensional porous NiCo 2 O 4 nanowires array grown on nickel foam (NiCo 2 O 4 NWs/NF) via a facile hydrothermal method exhibit highly efficient glucose sensitivity of 5916 μA mM −1 cm −2 , a detection limit of 1 μM-3.987 mM and LOD of 0.94 μM (S/N = 3) [239]. As conducting substrate, nickel foam not only provides the large electrochemically active surface area due to three-dimensional interconnected features, but also directs the growth of one-dimensional NiCo 2 O 4 porous nanowires [240]. Besides, the one-dimensional porous NiCo 2 O 4 nanowires array provided sufficient transport channels for ions and abundant active sites for redox reactions. Carbon cloth has also been used as a potential conducting surface for the growth of porous NiCo 2 O 4 nanowires. As fabricated enzyme-free NiCo 2 O 4 porous nanowire arrays supported on carbon cloth-based electrode for glucose sensing exhibited a linear dynamic range of 1 μM-0.63 mM, the sensitivity of 4.12 mA mM −1 cm −2 , and low detection limit of 0.5 μM [241].
One of the main disadvantages of using bare NiCo 2 O 4 is its poor electrical conductivity. However, this limitation can be overcome by forming its composite/hybrid materials. It has been reported that the electrical conductivity and hence the electrochemical biosensing performance of NiCo 2 O 4 can be improved by making its composites with conducting carbonaceous materials like graphene, reduced graphene oxide, carbon nanotubes (single and multi-walled), carbon nanofibers; conducting polymers like polypyrrole (PPy), polyaniline (PANI); metal oxides NiO, Co 3 O 4 , SnO 2 , MnO 2 ; and metals like Au, Pd, etc. Among these, the carbonaceous materials are considered to be potential candidates as compared to others due to their excellent electrical conductivities, good mechanical strength, thermal and chemical stabilities, and resistance to oxidation-reduction reactions. Besides, these carbonaceous materials provide a large specific surface area for better adsorption of analytes, which ultimately results in very high sensitivity and very low detection limits.
The two-dimensional one-atom-thick layered structure of graphene has been extensively used for making composites with NiCo 2 O 4 due to its high specific surface area of 2670 m 2 g −1 and excellent conductivity [242,243]. Studies have revealed a higher specific surface area for the NiCo 2 O 4 /reduced graphene oxide composites as compared to bare NiCo 2 O 4 nanoparticles (Fig. 26a) [244]. Even the pore width was less in the case of NiCo 2 O 4 /reduced graphene oxide composites. Various glucose-sensing scans are given in Fig. 26b-d. The enhanced redox peak current density for NiCo 2 O 4 /reduced graphene oxide composites as compared to pure NiCo 2 O 4 was attributed to the lesser extent of aggregation of graphene sheets due to the interception of the NiCo 2 O 4 nanoparticles on graphene surface causing weakening of π-π interaction between individual graphene sheets, faster diffusion rates and electron transfer between the glucose molecules and the electrode surface [245]. Ma et al. [246] developed NiCo 2 O 4 nanowrinkles/reduced graphene oxide hybrid-based modified GCE for non-enzymatic glucose detection at the physiological level. As far as the concentration of the glucose is concerned, the oxidation potential of glucose decreased while oxidation peak current increased proportionally to a greater extent for NiCo 2 O 4 nanowrinkles/reduced graphene oxide hybrid-based modified GCE as compared to single component Co 3 O 4 , NiO and bare NiCo 2 O 4 at a scan rate of 100 mV s −1 in 0.1 M NaOH (Fig. 27a-d). The results confirmed the crucial role of reduced graphene oxide in improving the electrocatalytic biosensing performance of the NiCo 2 O 4 spinel for different concentrations of glucose.
In addition to two-dimensional graphene, Wu et al. [245] reported the synthesis of three-dimensional graphene foam (3DGF) through a chemical vapor deposition technique. The 3DGF provides additional stability and large porous surface as well as high conductivity to the hierarchical NiCo 2 O 4 composites. NiCo 2 O 4 hierarchical nanoneedles were deposited onto the surface of 3DGF via a hydrothermal method. The synergism between hierarchical NiCo 2 O 4 nanoneedles and 3DGF exhibited a high sensitivity of 2524 μA mM −1 cm −2 and a limit of detection 0.38 μM (S/N = 3). Further, as fabricated electrode showed excellent selectivity for glucose even in the presence of interfering compounds like dopamine, ascorbic acid, lactose, d-Fructose, and urea as negligible current responses were observed on their additions as compared to glucose. NiCo 2 O 4

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nanospheres/reduced graphene oxide composite prepared by a template-based method using the Cu 2 O/GO template achieved a high sensitivity of 2082.57 μA mM −1 cm −2 , the detection range of 0.04-1.28 mM, and low detection limit of 0.7 μM [137]. Ni et al. [247] reported a reduced graphene oxide supported NiCo 2 O 4 nanorods composite prepared via an ionothermal method using deep eutectic solvents. The modified GCE exhibited superior electrocatalytic biosensing of glucose with a wide double-linear range from 1 μM to 25 mM and a very low detection limit of 0.35 μM (S/N = 3). The presence of a large number of small interconnected nanoparticles on the surface of the NiCo 2 O 4 nanorods provided the dense electrocatalytic active site in coordination with reduced graphene oxide which provided large surface area and excellent electrical conductivity (Fig. 28a).
Another way of preventing the aggregation of graphene sheets, which reduces the specific surface area and inhibits the fast mass transfer, is the nitrogen doping. This nitrogen doping is not only supposed to facilitates the charge transfers between adjacent carbon atoms but also suppresses the electrons and holes recombination necessary for better electrical conductivity and electrocatalytic oxidation of glucose [248,249]. Detailed characterization revealed that in the course of hydrothermal reactions, the graphene was reduced to nitrogen-doped reduced graphene oxide when glycine acted as a source of nitrogen. Further, the nitrogen-doped reduced graphene was self-assembled into hydrogels with interconnected 3D porous network structure resulted from an increased extent of π-π stacking interactions. This 3D form provides a sufficiently large surface area and active sites for the better adsorption of the analyte species. To ascertain this, Lu et al. [38] explored the interactions of flower-like NiCo 2 O 4 and 3D nitrogen-doped holey graphene hydrogel (NHGH)-modified GCE for electrochemical biosensing of glucose (Fig. 28b).  Similar to graphene, carbon nanofibers also possess excellent dimensional, thermal and chemical stability as well as good electrical conductivity. Recently, these fibers have attracted wide attention and have been widely explored in fields such as electrochemical cells, catalysis, adsorption, structure enhancement, biosensors, gas sensors, and nanodevices [250,251]. Among various synthetic methods, electro-spinning is considered to be the most suitable low-cost and simple method for synthesizing carbon nanofibers [252,253]. Liu et al. [39] explored the glucose-sensing behavior of NiCo 2 O 4 nanoneedledecorated electrospun carbon nanofiber nanohybrids. Faster electrocatalytic oxidation of glucose was reported for nanohybrids as compared to bare NiCo 2 O 4 nanoneedle and electrospun carbon nanofiber-modified GCEs. The fact was supported by a large increase in the anode peak current and a positive shift in the anode peak potential.
Novel metals such as Au, Ag, and Pd, have also been used to prepare NiCo 2 O 4 composites to improve the biosensing capabilities. Recently, dealloying has been used as a convenient method for preparing nanoporous metals with a 3D bicontinuous structure, which is characterized by open nanopores with adjustable sizes [254][255][256]. These 3D nanoporous metals act as conductive surfaces for the deposition of biosensors electrocatalytic materials such as NiCo 2 O 4 since they provide high conductivity and large surface area. Disposable needle-type hybrid electrode comprising a stainless steel core modified with a 3D nanoporous Au/NiCo 2 O 4 nanowall hybrid structuremodified electrochemical non-enzymatic glucose sensor showed a linear response of 0.01-21 mM glucose, high  Fig. 28 a Proposed mechanism of glucose sensing using NiCo 2 O 4 nanorods/rGO/Nafion composite-modified GCE. Reproduced with permission from Ref. [247]. Copyright © 2018 Elsevier Ltd. b Schematic presentation of the synthesis of the NHGH/NiCo 2 O 4 electro-catalyst for nonenzymatic glucose sensing, Reproduced with permission from Ref. [38]. Copyright © 2019 Elsevier B.V. sensitivity of 0.3871 μA μM −1 cm −2 , detection limit of 1 μM within a response time of < 1 s [257]. Naik et al.  [259].
Similar to metals, conducting polymers also possess the electronic, electrical, and optical properties, easy synthesis, excellent mechanical stabilities and most importantly the low toxicity and biodegradability, the issues which are generally associated with metals. Moreover, the noble metals are easily poisoned by some intermediates produced during the oxidation of glucose. Among various conducting polymers, polyaniline and polypyrrole have gained much attention due to their superior thermal and oxidative stabilities [260,261]. Constructing a core-shell nanostructure comprising conductive polymer coating as the outer walls of metal oxides is the most important strategy for enhancing the conductivities [262]. NiCo 2 O 4 @PANI nanoparticles with an average particle size 25 nm shortened the ion transport pathway and the modified GCE exhibited a sensitivity of 4.55 mA mM −1 cm −2 , a detection limit of 0.3833 μM and linear dynamic range of 0.0150-4.7350 mM (Fig. 29a, [263]. The PANI core-shell provided more effective electrical contact between redox-active centers and the electrolyte resulting in good contact and small diffusion distances for electron transports which subsequently improved the sensor activity. NiCo 2 O 4 @Ppy nanowires grown on Ni foam were synthesized via hydrothermal growth and oxidant-induced polymerization process (Fig. 29c-e). The fabricated glucose sensor showed high sensitivity 3059 μA mM −1 cm −2 , low detection limit 0.22 μM, and wide linear dynamic range 0.001-20 mM. The excellent electrocatalytic behavior was attributed to the synergism due to bimetallic oxide, the significant role of Ppy in transmitting charges among electrode material due to its excellent conductivity, non-collapsing and non-agglomeration of the NiCo 2 O 4 due to Ppy coating, and absence of any adhesive or conductive agent during electrode fabrication [264]. NiCo 2 O 4 nano-/microstructures combined with smarter nano-architectured metal oxides (Co 3 O 4 , SnO 2 , NiO, and MnO 2 , etc.) have many synergistic multifunctional properties of nanostructured components including dense and easily accessible electroactive sites, altered bandgap energies, faster charge transfer processes, and reduced internal resistance. The p-type semiconductor nanostructured NiCo 2 O 4 [32], Co 3 O 4 [265], and n-type materials SnO 2 [266] and MnO 2 [267] have the bandgap energies of 2.1, 2.2, 3.6, and 1.3 eV, respectively. Due to slightly different bandgap energies, the semiconductor metal oxides introduce in situ impurity bands in NiCo 2 O 4 which increase the electron conductivity to extract excellent electrocatalytic efficiencies [268]. Chen et al. [269] [272]. Therefore, for practical applications, these components should not affect the amperometric parameters and the positive results have been reported [230,237,238,241,247,272,273]. Additionally, reproducibility and stability of the modified NiCo 2 O 4 -based electrodes have been analyzed. The results have shown acceptable reproducibility with a very low relative standard deviations in many studies [123,142,161,193,199,200,227,274]. The electrochemical sensing parameters such as sensitivity, linear dynamic range, and detection limits for various NiCo 2 O 4 -based modified electrodes toward glucose are compared in Table 1.
In addition to electrochemical sensing of glucose using NiCo 2 O 4 nano-/microstructure-modified electrodes, colorimetric sensing has also been reported by Huang et al. [274]. They explored the peroxidase-like activity of the hierarchical NiCo 2 O 4 hollow sphere which was directly dependent on the concentration of H 2 O 2 produced by the oxidation of glucose to gluconic acid in the presence of glucose oxidase (Go x ). Hence, a colorimetric method for the detection of glucose can be designed using NiCo 2 O 4 . The higher the concentration of the glucose, the more was the production of H 2 O 2 and hence the greater was the oxidation of the 3,3,5,5-tetramethylbenzidine (TMB) to oxidized TMB. Absorbance at λ max = 652 nm for oxidized TMB was increased linearly with the concentration of glucose. The linear range was observed between 0.1 and 4.5 mM with a low detection limit of 5.31 μM (Fig. 30a, b). The corresponding reaction mechanism is shown in Fig. 30c. Intrinsic peroxidase and oxidase-like activities of the NiCo 2 O 4 architectures were also confirmed by Su et al. [275] by analyzing the electron spin resonance spectra for the oxidation of TMB by NiCo 2 O 4 mesoporous spheres. The oxidation was accompanied without the production of 1 O 2 and OH · radicals. Additionally,  86 10-10480 - [277] these peroxidase-like activities were feasible even over a broad temperature range.

H 2 O 2 Biosensors
H 2 O 2 is the most important byproduct produced from some enzyme-catalyzed biochemical reactions. In addition to its importance as a regulator of immune cell activation, vascular remodeling, and stomatal closure during metabolic processes, it also has pharmaceutical, clinical, environmental, textile, and food manufacturing applications [278].  [284], nickel phosphide (Ni 2 P) nanosheets array on titanium mesh [285], cobalt nitride (Co 3 N) nanowire array on Ti mesh [286], nanoporous carbon nanofibers/Pt nanoparticles [287], Ag decorated hierarchical Sn 3 O 4 [288] and many more for the electrochemical sensing of H 2 O 2 amperometrically.

Urea Biosensors
Urea (carbamide or carbonyl diamide) is one of the main end products of protein metabolism in humans and animals. Urea is exclusively formed in the liver, and is transported by the bloodstream to the kidneys for excretion in the human body. The normal level of urea in human blood serum is 2.5-7.5 mM [292][293][294][295]. Amount of urea above or below the permissible level in the serum results in chronic renal and hepatic failure, gastrointestinal bleeding, and nephritic syndrome [296]. Similar to other metabolically important biomolecules, the literature reports enzymatic as well as non-enzymatic biosensors for the selective and highly sensitive urea sensors. Enzyme-based urea biosensors explore the use of urease enzyme which facilitates the hydrolysis of urea into ammonium ( NH + 4 ) and bicarbonate ( HCO − 3 ) ions (Eq. 38) [297].
The overall reaction can be written as: Recently, Amin et al. [298] explored the urea sensing behavior of NiCo 2 O 4 nanoneedle-modified GCE which showed high sensitivity with a linear response concentration range of 0.01-5 mM and low detection limit of 1.0 µM. It was proposed that initially Ni 2+ ions are oxidized to Ni 3+ ions to form NiOOH in an alkaline medium which is reduced back to give Ni(OH) 2 at the time of urea electro-oxidation [299]. Mesoporous spinel NiCo 2 O 4 nanostructures prepared via facile chemical deposition method showed higher catalytic activities, lower overpotential, and more tolerance toward urea electro-oxidation as compared to Co 3 O 4 [227]. NiCo 2 O 4 /3D graphene/ITO exhibited high sensitivity of 166 μA mM −1 cm −2 , a linear range of 0.06-0.30 mM, and a low detection limit of 5.0 µM with a very small response time of 1 s through non-enzymatic detection method [300]. Further, a higher oxidation peak for NiCo 2 O 4 /3D graphene in the CV as compared to NiCo 2 O 4 /CNT-modified electrode confirmed the superiority of 3D graphene as a matrix material for electrode fabrication. The higher oxidation current potential was attributed to the highly porous nature and excellent conductivity of the interconnected 3D graphene matrix [301]. Since oxidation of urea is in alkaline medium, higher electrocatalytic oxidation of urea is recorded at higher pH conditions. However, beyond an optimum pH the electrooxidation decreases due to blockage of the active sites by OH − ions [302].

Electrochemical Determination of Some Other Bioanalytes
Some other bioanalytes such as rutin, trypsin, ascorbic acid, dopamine, uric acid, and tryptophan have also been electrochemically analyzed using nano-/micro-structured hybrid NiCo 2 O 4 -modified electrodes. Rutin, a flavonoid substance, is used as anti-bacterial, anti-viral, antiproliferative, antioxidants, antagonists, and anti-inflammatory. It also controls the blood pressure and vascular fragility including hereditary hemorrhagic telangiectasia, haemangiomas, vitamin C deficiency, etc. [303,304]. Cui et al. [305] reported the fabrication of GCE modified with mesoporous NiCo 2 O 4 -decorated reduced graphene oxide for the electrochemical sensing of rutin using differential pulse voltammetric (DPV) technique.
Flake-like NiCo 2 O 4 sheets anchored on the wrinkled reduced graphene oxide sheets through electrostatic interaction prevented the self-agglomerations. The wide linear range of 0.1-150 μM and a low detection limit of 0.01 μM were observed along with excellent anti-interference capabilities. The strong synergism between reduced graphene sheets and mesoporous NiCo 2 O 4 resulted in increased redox peak current and decreased potential difference. During electrooxidation, rutin is converted into 3′,4′-diquinone with the release of two H + ions and two electrons (Eq. 42) [306,307].
Trypsin, a serine protease secreted from the pancreas, has also been widely studied recently as it used for identifying and determining the amino acid sequence in protein and peptide, particularly at the C-terminus and as a specific biomarker for diseases like chronic cystic fibrosis, chronic pancreatitis, cancer, and many pathological changes [308,309]. Lin et al. [310] reported a large and prompt rise in electrochemical signal in the presence of trypsin by NiCo 2 O 4 -poly(amidoamine)/peptide@g-C 3 N 4 nanocomposite-modified GCE. 3,4,9,10-perylene tetracarboxylic acid (PTCA) was used to connect the peptides and g-C 3 N 4 . The modified GCE exhibited increased DPVs peak currents when the trypsin concentration was increased from 10 −10 to 10 −4 mg mL −1 . Kaur electrochemical sensing of ascorbic acid, dopamine, uric acid, and tryptophan using NiCo 2 O 4 /Nano-ZSM-5 nanocomposite-modified GCE. Wide linear ranges were 1-1200, 0.6-900, 0.9-1000, and 0.9-1000 μM, while the corresponding detection limits were 0.8, 0.5, 0.7, and 0.7 μM for ascorbic acid, dopamine, uric acid, and tryptophan, respectively. Simultaneous detection was possible as the anodic oxidative peak currents were observed at different applied potentials, i.e., 0.158, 0.394, 0.561, and 0.820 V, respectively, for ascorbic acid, dopamine, uric acid, and tryptophan in DVP plots at a scan rate of 20 mV s −1 .
Detailed comparison from Tables 1 and 2 indicates that the morphology of the NiCo 2 O 4 nano-/microstructures and the presence of any other component along with NiCo 2 O 4 significantly affect the biosensing efficiency. Comparative analysis revealed better electrochemical sensing of glucose by one-dimensional nanofibres and nanorods and twodimensional nanosheets like morphologies of NiCo 2 O 4 than other morphologies. Doped and composites/hybrid NiCo 2 O 4 nano-/microstructures exhibit superior sensing parameters that bare NiCo 2 O 4 . In particular, graphenic nanomaterials due to their excellent conductivity and large surface area significantly elevate the I-V characteristics to many folds. These materials also accelerate the rate of heterogeneous electron transfer, i.e., the transfer of electrons from/to electrode to/from bioanalyte molecules [312].

Conclusion
Herein, various strategies for the synthesis of spinel NiCo 2 O 4 nano-/microstructures with versatile morphologies and their subsequent use for the development of biosensors for efficient non-enzymatic sensing and detection of biomolecules such as glucose, H 2 O 2 and urea are comprehensively reviewed. As compared to NiO and Co 3 O 4 , the NiCo 2 O 4 nanomaterials showed better electrochemical sensing as adjudged by broader redox peaks with larger area coverage in the CV curves. The biosensing efficiency of the NiCo 2 O 4 nano-/microstructures can be improved by engineering the morphology, specific surface area, porosity, doping and by making composite/hybrids with various carbonaceous materials, conducting polymers, metal oxides, non-metals and metals. These materials not only improve the mechanical, thermal, and chemical stability but also modulate the bandgap energies, electronic and ionic conductivities, dispersion behavior, avoid aggregation of the NiCo 2 O 4 nanomaterials and provide short electron and ion diffusion pathways. All these factors contribute to better electrocatalytic behavior with excellent sensitivity, selectivity, and long-term stability of the spinel NiCo 2 O 4 nano-/microstructure-based biosensors. It is hoped that this review will provide basic ideas as well as new insights for future research and progress in this field.

Challenges and Future Perspectives
Despite extensive research in this area, many issues that impede the practical application of NiCo 2 O 4 nano-/microstructures need to be addressed for further improvement. Some of these issues have been identified herein. Structural features of the NiCo 2 O 4 nanomaterials are controlled by factors like temperature, pH of the reaction solution, precursor concentration, solvent nature and quantity, presence of the growth directing agents and templates, etc. It is, therefore, one of the major challenges to design largescale and low-cost morphology controlled synthesis of the NiCo 2 O 4 nanomaterials for next-generation biosensors.
Rational combination of NiCo 2 O 4 nano-/microstructures with other hybrid materials or conductive substrates to designing NiCo 2 O 4 composite/hybrids is found to improve the intrinsic characteristics like low electronic conductivity and wide bandgap and hence the biosensing behavior of NiCo 2 O 4 . However, still, more in-depth understanding is required to correlate the synergism between the components of the composite/hybrid materials.
Cost-effectiveness, easy to manufacture, recyclability, sensor disposal, and biocompatibility of NiCo 2 O 4 nano-/ microstructure-based biosensors are other issues that need to be addressed and solved. The high cost of electrochemical work stations restricts the practical applications of these sensors. In this regard, portable and wearable sensing devices will be promising. The toxicity issues of the NiCo 2 O 4 nano-/ microstructures and other components are very rarely discussed in the literature. Future research thus should also focus on studying this important issue.
The biosensing behavior of the NiCo 2 O 4 nano-/microstructure-based sensors is affected by factors like working temperature, pH of the medium, scan rates, and applied potential. The optimization of these parameters is rarely addressed. Studying the specificity of NiCo 2 O 4 -based sensors from competitive assays requires an appropriate protocol because it is reported that the sensor can detect glucose, H 2 O 2 , urea, trypsin, etc.
Due to complex structures of the biomolecules, the interface and mechanistic studies at the surface of the nanomaterials are still undefined. The redox transformation of the molecules during electrocatalytic biosensing is also a debatable issue. Therefore, future work should focus on elucidating the interaction mechanism between nanomaterials and biomolecules on the electrode surface, to fabricate a new generation of biosensors. licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creat iveco mmons .org/ licen ses/by/4.0/.