Highly sensitive non-enzymatic lactate biosensor driven by porous nanostructured nickel oxide
Introduction
In recent years, biotechnology has evolved to allow the detection and continuous monitoring of various metabolites in bodily fluids, such as blood and perspiration, using wearable devices [[1], [2], [3], [4], [5]]. Among various metabolites, lactate concentration is one of important key parameters often targeted for continuous monitoring in sports medicine, and clinical diagnostics in contexts including shock/trauma treatment and hypoxia [[6], [7], [8]]. During intensive physical exercise, as oxygen supply becomes insufficient, pyruvate in the body converts to lactic acid, thus increasing the level of lactate in blood and perspiration. This is referred to as lactic acidosis [[9], [10], [11]]. The normal concentration of lactate in human blood (0.5–1.5 mM) and perspiration (∼5 mM) can increase to over 10 and 25 mM, respectively in lactic acidosis [12]. This abnormal increase of lactate concentration induces an increase of acidity inside cells that have switched to anaerobic metabolism, causing muscular fatigue as well as reducing training efficiency [13]. In addition to its applications in training, monitoring the level of lactate in the body can have clinical relevance as an indicator of whether the acid-base (pH) balance is normal. It has been reported that certain diseases such as cancer, diabetes, sepsis, and cardiovascular disease also give rise to lactic acidosis because of increased lactate level [[14], [15], [16], [17]]. Therefore, reliable detection of the level of lactate in bodily fluids has great relevance for monitoring the physiological condition of humans.
Most lactate biosensors described in the literature are based on enzymes such as lactate oxidase (LOx) or lactate dehydrogenase (LDH), which are known to biologically react selectively with lactate [[18], [19], [20]]. These approaches are attractive for their biological selectivity and direct measurement of lactate, which are owing to inherent characteristics of the enzymes employed. Enabled by the development of novel fabrication methods, enzyme-immobilized lactate sensors have demonstrated the advantages of good selectivity, rapid response, and high sensitivity [[21], [22], [23]]. Enzyme-based lactate sensors, however, have some limitations including low reliability because of biological degradation that prevents measurement of a constant signal over time, and high production cost resulting from the price of enzymes [24]. In order to address these issues, non-enzymatic detection of lactate based on electrocatalytic reaction of non-biological materials such as metal oxide, metal organic framework and conducting polymer have been proposed [[25], [26], [27], [28], [29]]. This can bring high efficiency, reliability, and recyclability to lactate sensors. Among various candidates, metal oxide nanostructures such as CuO, Co3O4, and NiO have been drawn attention as a non-enzymatic biosensor electrode due to its electrochemical catalytic properties on certain metabolite as well as its advantage in process to easily control the size and porositiy for better sensitivity [[30], [31], [32]]. Especially for lactate sensing, nickel oxide has been reported to confer electrocatalytic characteristics to lactate due to inherent Ni3+ known as electrochemical active sites to induce oxidation of organic compound [[33], [34], [35]]. Furthermore, nickel oxide is earth-abundant and easily fabricated with conventional solution-based methods. Nonetheless, in comparison with enzymatic lactate sensors, the relatively low sensitivity of non-enzymatic nickel oxide-based sensors is still problematic.
Herein, we demonstrate a highly sensitive non-enzymatic sensor for lactate detection using porous nickel oxide to introduce an additional pathway for electron transfer. Using a sol-gel based inverse micelle method, the porosity of nickel oxides was well controlled with different calcination temperatures of 250, 350, and 450 °C. Electron microscopy showed that, with increasing calcination temperature, the surface area and pore size of nickel oxide decreased because of partially aggregated surface morphology. Furthermore, elemental analysis revealed that low-temperature calcination induces more active sites for Ni3+ electrocatalytic reaction to lactate, which influenced lactate detection because of decreased connected active sites to lactate. In addition to characterization of the porous nickel oxide sensor, we assessed its sensing capability to lactate with cyclovoltammetry and chronoamperometric response. As a result, we found that nickel oxide calcined at 250 °C displays maximal surface area and concentration of active sites, thus exhibiting the highest sensitivity (62.35 μA/mM [cm2]) and lowest detection of limit (27 μM) among as-synthesized nickel oxides tested. Moreover, as-synthesized nickel oxide shows good selectivity against other metabolites such as l-ascorbic acid, uric acid, and dopamine, and is close to a potential alternative to enzymatic sensing electrodes.
Section snippets
Materials
Nickel (II) nitrate hexahydrate (Ni(NO3)2·6H2O), nitric acid, 1-butanol, PEO20-PPO70-PEO20 (P123), L-(+)-lactic acid (LA), dopamine (DA), ascorbic acid (AA), and uric acid (UA) were purchased from Sigma-Aldrich. Commercially available NiO (c-NiO) powders were received from Alfa Aesar. All chemicals were used as received without further purification.
Synthesis of mesoporous structure NiOs
The mesoporous structure NiOs were fabricated by sol-gel based inverse micelle methods [36]. In the synthesis process of porous samples, nickel (II)
Results and discussion
We synthesized highly porous nickel oxides by a sol-gel based inverse micelle method which is known to fabricate porous metal oxide with control over porosity. During synthesis, the P123 surfactant species that induce hydrogen bonding can act physical barrier with metal source thus, hinder aggregation of metal oxide particles and disappear during annealing process. Moreover, the different calcination temperature of 250 (NiO250), 350 (NiO350) or 450 °C (NiO450) was designated to control over
Conclusion
Here we report successful fabrication of porous nickel oxide structures by a sol-gel based inverse micelle method under mild reaction conditions. As-synthesized-NiOs exhibit higher surface area and greater abundance of electrocatalytic Ni3+ sites with decreasing calcination temperature between 450 and 250 °C. In order to estimate the sensing capability of as-synthesized NiOs, we measured cyclovoltametric and chronoamperometric responses to different lactate concentrations. Despite the
Declaration of interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Author contributions
Y.M.P, S.J.S, H.K., and T.J.P designed research; S.K., W.Y., and H.L. synthesized materials and performed characterization; Y.M.P, S.J.S, S.K., and H.K. analyzed data; and S.K., Y.M.P, S.J.S, and T.J.P wrote the paper.
Acknowledgement
This research was financially supported by an internal research program of the Korea Institute of Industrial Technology, South Korea (Project No. PEO19052), and by the Human Resources Development program (No. 20174030201830) of the Korea Institute of Energy Technology Evaluation and Planning (KETEP) grant funded by the Korea government Ministry of Trade, Industry and Energy.
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