Online Electrochemical System Research Articles

Online ascorbate sensing reveals oxidative injury occurrence in inferior colliculus in salicylate-induced tinnitus animal model

Tinnitus is a widespread and serious clinical and social problem. Although oxidative injury has been suggested to be one of pathological mechanisms in auditory cortex, whether this mechanism could be applied to inferior colliculus remains unclear. In this study, we used an online electrochemical system (OECS) integrating in vivo microdialysis with selective electrochemical detector to continuously monitor the dynamics of ascorbate efflux, an index of oxidative injury, in inferior colliculus of living rats during sodium salicylate-induced tinnitus. We found that OECS with a carbon nanotubes (CNTs)-modified electrode as the detector selectively responses to ascorbate, which is free from the interference from sodium salicylate and MK-801 that were used to induce tinnitus animal model and investigate the N-methyl-d-aspartate (NMDA) receptor mediated excitotoxicity, respectively. With the OECS, we found that the extracellular ascorbate level in inferior colliculus significantly increases after salicylate administration and such increase was suppressed by immediate injection of NMDA receptor antagonist MK-801. In addition, we found that salicylate administration significantly increases the spontaneous and sound stimuli evoked neural activity in inferior colliculus and that the increases were inhibited by the injection of MK-801. These results suggest that oxidative injury may occur in inferior colliculus following salicylate-induced tinnitus, which is closely relevant to the NMDA-mediated neuronal excitotoxicity. This information is useful for understanding the neurochemical processes in inferior colliculus involved in tinnitus and its related brain diseases.

Ascorbate oxidase-like nanozyme with high specificity for inhibition of cancer cell proliferation and online electrochemical DOPAC monitoring

Despite the extensive investigation of the nanozymes exhibit their favorable performance compared to natural enzymes, nevertheless, the highly specific nanozyme still needs to be developed so that it can meet the requirements of exploring the mechanism as well as administration of related diseases and selective monitoring in biological system. In this study, self-assembled glutathione-Cu/Cu2O nanoparticles (GSH-Cu/Cu2O NPs) that exhibits specific ascorbic acid (AA) oxidase-like catalytic activity were constructed for AA-activated and H2O2-reinforced cancer cell proliferation inhibition and selective neurochemical monitoring. Cu/Cu2O NPs demonstrates effective AA oxidase-like activity and no common characteristics of other redox mimic enzymes often present in nanozyme. In particular, we found that the AA oxidase-like activity of GSH-Cu/Cu2O nanozyme was significantly improved by about 40% by improving the activation ability toward oxygen. The synthesized nanozyme can induce the generation of active oxygen by accelerating the oxidation of AA, which effectively suppresses the proliferation of cancer cells. We constructed an online electrochemical system (OECS) though loading nanozyme with enhanced ascorbate oxidase activity into a microreactor and setting it in the upstream of the detector. This GSH-Cu/Cu2O NPs-integrated microreactor can completely eliminate AA interference of the physical level toward 3,4-dihydroxy phenylacetic acid (DOPAC) electrochemical measurement, and the nanozyme-based OECS is able to continuously capture DOPAC alteration in rat brain acidosis model. Our findings may inspire rational design of nanozymes with high specificity as well as nanozyme-based selectivity solution for in vivo detection and show promising opportunities for their involvement in neurochemistry investigation.

Structure Defect Tuning of Metal-Organic Frameworks as a Nanozyme Regulatory Strategy for Selective Online Electrochemical Analysis of Uric Acid.

Nanozymes have been designed to address the limitations of high cost and poor stability involving natural enzymes in analytical applications. However, the catalytic efficiency of the nanozyme still needs to be improved so that it can meet the selectivity and stability requirements of accurate biomolecule analysis. Here, we presented structure defects of metal-organic frameworks (MOFs) as a tuning strategy to regulate the catalytic efficiency of artificial nanozymes and investigated the roles of defects on the catalytic activity of oxidase-like MOFs. Structural defects were introduced into a novel Co-containing zeolitic imidazolate framework with gradually loosened morphology (ZIF-L-Co) by doping cysteine (Cys). It was found that with the increase in defect degree, the properties of materials such as ascorbate oxidase-like, glutathione oxidase-like, and laccase-like were obviously enhanced by over 5, 2, and 3 times, respectively. In-depth structural investigations indicate that the doping of sulfur inducing structural defects which may destroy the equilibrium state between cobalt and nitrogen in 2-methylimidazole and distort the crystal lattice, thereby enhancing the adsorption of oxygen and thus promoting the oxidase-like activity. The ZIF-L-Co-10 mg with enhanced ascorbate oxidase- and laccase-like activity was loaded into a microreactor and integrated into an online electrochemical system (OECS) in the upstream of the detector. This nanozyme-based microreactor can completely remove ascorbic acid, dopamine, and 3,4-dihydroxyphenylacetic acid which are the main interference toward uric acid (UA) electrochemical measurement, and the ZIF-L-Co-10 mg Cys-based OECS system is capable of continuously capturing UA change in rat brain following ischemia-reperfusion injury. Structure defect tuning of ZIF-L-Co not only provides a new regulatory strategy for artificial nanozyme activity but also provides a critical chemical platform for the investigation of UA-related brain function and brain diseases.

Electrochemical Sensing of Ascorbate as an Index of Neuroprotection from Seizure Activity by Physical Exercise in Freely Moving Rats.

Physical exercise (PE) has been drawing increasing attention to prevent and alleviate neural damage of brain diseases; however, in vivo sensing of the neuroprotection ability of PE remains a challenge. Here, we find that ascorbate can be used as a small molecular index for neuroprotective function of PE and the neuroprotection ability of PE can thus be in vivo monitored with an online electrochemical system (OECS) in freely moving animals. With the OECS as the sensing system, we find that the concentration of ascorbate in the microdialysate from the striatum increases greatly in kainic acid (KA)-induced seizure rats and reaches twice the basal level (i.e., 214.4 ± 32.7%, p < 0.001, n = 4) at a time point 90 min after KA microinjection. Such an increase of ascorbate is obviously attenuated (i.e., 153.6 ± 23.9% of the basal level, p < 0.05, n = 3) after PE, showing the neuroprotective activity of PE. This finding is believed to be significant in providing chemical insight into the neuroprotection ability of PE.

ZIF-67 as a Template Generating and Tuning "Raisin Pudding"-Type Nanozymes with Multiple Enzyme-like Activities: Toward Online Electrochemical Detection of 3,4-Dihydroxyphenylacetic Acid in Living Brains.

Due to its unique structure and high porosity, metal-organic frameworks (MOFs) can act not only as nanozyme materials but also as carriers to encapsulate natural enzymes and thus have received extensive attention in recent years. However, a few research studies have been conducted to investigate MOF as a template to generate and tune nanozymes in the structure and performance. In this work, the "raisin pudding"-type ZIF-67/Cu0.76Co2.24O4 nanospheres (ZIF-67/Cu0.76Co2.24O4 NSs) were obtained by rationally regulating the weight ratio of ZIF-67 and Cu(NO3)2 in the synthesis process. Here, ZIF-67 not only acts as a template but also provides a cobalt source for the synthesis of cobalt copper oxide on the surface of ZIF-67/Cu0.76Co2.24O4 NSs with multiple enzyme-like activities. The ZIF-67/Cu0.76Co2.24O4 NSs can mimic four kinds of enzymes with peroxidase-like, glutathione peroxidase-like, superoxide dismutase-like, and laccase-like activities. Based on its laccase-like activity, an online electrochemical system for continuous monitoring of 3,4-dihydroxyphenylacetic acid with good linearity in the range of 0.5-20 μM and a detection limit of 0.15 μM was established. Furthermore, the alteration of DOPAC in the brain microdialysate before and after ischemia of the rats' brain was also successfully recorded. This work not only raises a new idea for the synthesis of nanozyme materials with multiple enzyme activities but also provides a new solution for the detection of neurotransmitters in living brains.

Role of inferior colliculus in vestibular vertigo induced by water caloric irrigation

Background: Vestibular vertigo is a common clinical symptom; however, the central neural mechanism of it is still poorly understood.Objective: To demonstrate the changes of neural excitability and ascorbate in inferior colliculus (IC) in a rat vertigo model induced by water caloric irrigation.Methods: In vertigo model induced by water caloric irrigation, we recorded the changes of spontaneous firing rate (SFR) of IC. Then a technique that combining in vivo microanalysis with an online electrochemical system (OECS) was employed to monitor the changes of extracellular ascorbate in IC.Results: Electrophysiological studies showed that after vestibular ice water stimulation, the level of SFR in IC significantly increased, reaching (989 ± 9) % and (941 ± 62) % respectively at 2.0 h after contralateral ice water vestibular stimulation and ipsilateral ice water vestibular stimulation. However, the level of ascorbate in IC dramatically decreased after ice stimulation, decreased to (30 ± 12) % and (57 ± 24) % of the basal level respectively in the contralateral group and ipsilateral group.Conclusions and significance: These findings suggest that inferior colliculus plays a role in peripheral vertigo, which would appear useful for uncovering neural mechanisms of vertigo and help finding novel therapeutic targets for vertigo.

Persistent oppression and simple decompression both exacerbate spinal cord ascorbate levels.

Background: Surgical decompression after acute spinal cord injury has become the consensus of orthopaedic surgeons. However, the choice of surgical decompression time window after acute spinal cord injury has been one of the most controversial topics in orthopaedics.Objective: We apply an online electrochemical system (OECS) for continuously monitoring the ascorbate of the rats' spinal cord to determine the extent to which ascorbate levels were influenced by contusion or sustained compression.Methods: Adult Sprague-Dawley rats (n=10) were instrumented for ascorbate concentration recording and received T11 drop spinal cord injury (SCI). The Group A (n=5) were treated with immediately decompression after SCI. The Group B (n=5) were contused and oppressed until 1 h after the injury to decompress.Results: The ascorbate level of spinal cord increased immediately by contusion injury and reached to 1.62 μmol/L ± 0.61 μmol/L (217.30% ± 95.09% of the basal level) at the time point of 60 min after the injury. Compared with the Group A, the ascorbate level in Group B increased more significantly at 1 h after the injury, reaching to 3.76 μmol/L ± 1.75 μmol/L (430.25% ± 101.30% of the basal level). Meanwhile, we also found that the decompression after 1 hour of continuous compression will cause delayed peaks of ascorbate reaching to 5.71 μmol/L ± 2.69 μmol/L (627.73% ± 188.11% of the basal level).Conclusion: Our study provides first-hand direct experimental evidence indicating ascorbate is directly involved in secondary spinal cord injury and exhibits the dynamic time course of microenvironment changes after continuous compression injury of the spinal cord.

Single-atom electrocatalysis: a new approach to in vivo electrochemical biosensing

Modulation of interfacial electron transfer has been proven to pave a new approach to in vivo electrochemical monitoring of brain chemistry; however, designing and establishing highly efficient electrocatalytic scheme towards neurochemicals remain a long-standing challenge. Here, we find that recently established single-atom catalyst (SAC) can be used for catalyzing the electrochemical process of physiologically relevant chemicals and thus offers a new avenue to in vivo electrochemical biosensing. To prove this new concept, we used Co single-atom catalyst (Co-SAC), in which the atomic active sites are dispersed in ordered porous N-doping carbon matrix at atomic level, as an example of SACs for analyzing glucose as the physiologically relevant model chemicals. We found that Co-SAC catalyzes the electrochemical oxidation of hydrogen peroxide (H2O2) at a low potential of ca. +0.05 V (vs. Ag/AgCl). This property was further used for developing an oxidase-based glucose biosensor that was used subsequently as a selective detector of an online electrochemical system (OECS) for continuous monitoring of microdialysate glucose in rat brain. The OECS with Co-SAC-based glucose biosensor as the online detector was well responsive to glucose without interference from other electroactive species in brain microdialysate. This study essentially offers a new approach to in vivo electrochemical analysis with SACs as electrocatalysts to modulate interfacial electron transfer.

Ischemic Postconditioning Recovers Cortex Ascorbic Acid during Ischemia/Reperfusion Monitored with an Online Electrochemical System.

As a promising therapeutic treatment, ischemic postconditioning has recently received considerable attention. Although the neuroprotection effect of postconditioning has been observed, a reliable approach that can evaluate the neuroprotective efficiency of postconditioning treatment during the acute period after ischemia remains to be developed. This study investigates the dynamics of cortex ascorbic acid during the acute period of cerebral ischemia before and after ischemic postconditioning with an online electrochemical system (OECS). The cerebral ischemia/reperfusion injury and the neuronal functional outcome are evaluated with triphenyltetrazolium chloride staining, immunohistochemistry, and electrophysiological recording techniques. Electrochemical recording results show that cortex ascorbic acid sharply increases 10 min after middle cerebral artery occlusion and then reaches a plateau. After direct reperfusion following ischemia (i.e., without ischemic postconditioning), the cortex ascorbic acid further increases and then starts to decrease slowly at a time point of about 40 min after reperfusion. In striking contrast, the cortex ascorbic acid drops and recovers to its basal level after ischemic postconditioning followed by reperfusion. With the recovery of cortex ascorbic acid, ischemic postconditioning concomitantly promotes the recovery of neural function and reduces the oxidative damage. These results demonstrate that our OECS for monitoring cortex ascorbic acid can be used as a platform for evaluating the neuroprotective efficiency of ischemic postconditioning in the acute phase of cerebral ischemia, which is of great importance for screening proper postconditioning parameters for preventing ischemic damages.

Neuroprotective effects of MK-801 on auditory cortex in salicylate-induced tinnitus: Involvement of neural activity, glutamate and ascorbate

Tinnitus may cause anxiety, depression, insomnia, which impair the quality of life of millions worldwide. However, the mechanism of tinnitus remains to be understood, it has been previously hypothesized that the activation of N-methyl-D-aspartate (NMDA) receptor is involved in the tinnitus processes and blockade of the NMDA receptor is regarded as a therapeutic strategy for tinnitus treatment even if the rescue treatment is still proved invalid in some cases. To demonstrate the therapeutic effect of the NMDA receptor blocker on tinnitus, we examined here the spontaneous firing rate (SFR) and the neurochemical dynamics in the auditory cortex (AC) of rats after sodium salicylate (SS) injection, which is a widely used model for tinnitus research. We also recorded their responses to MK-801 treatment. Electrophysiological studies showed that MK-801 significantly suppresses SFR in AC of rats with SS-induced tinnitus. In addition, by using a technique that combining in vivo microdialysis with an online electrochemical system (OECS) and a high-performance liquid chromatography (HPLC), we found that the levels of both glutamate and ascorbate in AC dramatically increased after SS injection and that MK-801 administration attenuated those response. Further studies found that MK-801 given at a time point of 30 min pre- or post-injection of SS were more effective than that given at a time point of 60 min post-SS injection, indicating that the time point of MK-801 intervention has a critical impact on the therapeutic effect. These findings suggest that MK-801 plays a neuroprotective role against hyperactivity during tinnitus induced by SS and that the therapeutic effect depends on the time point of MK-801 intervention, which would advance the studies on understanding of the therapeutic potential of NMDA receptor antagonist in tinnitus therapy.

Therapeutic hypothermia effectively reduces elevated extracellular ascorbate concentrations caused by acute spinal cord injury

In recent years, systemic hypothermia has taken the spotlight for its use in spinal cord injury (SCI) research fields, but detailed molecular mechanisms are still not fully understood. In this study, we use an online-electrochemical system (OECS) to in vivo continuously monitor the ascorbate of the rats’ spinal cord. We find that the basal level of ascorbate in rat spinal cord is 1.85 ± 0.88 μmol L−1 (n = 20). It increased immediately after SCI and reached 2.36 ± 0.65 μmol L−1 (164.90% ± 7.99% of the basal level) (n = 5) at 60 min after the injury. The SCI-induced extracellular ascorbate increase is obviously attenuated by therapeutic hypothermia (28 °C) after injury and ascorbate returns to 3.01 ± 0.59 μmol L−1 (100.24% ± 5.02% of the basal level) (n = 5), at 60 min after SCI. These results substantially manifest that the OECS for ascorbate detection could be employed as a platform for understanding the pathological changes during spinal cord injury. This study provides experimental evidence for the essential roles of ascorbate in SCI which could serve as a biomarker for SCI. Our findings also raise the possibility that therapeutic hypothermia can effectively exert neuroprotection in the acute phase of SCI.

In Vivo Recording of Ascorbate and Neural Excitability in Medial Vestibular Nucleus and Hippocampus Following Ice Water Vestibular Stimulation in Rats

AbstractVertigo is a common clinical symptom; however, the chemical and electrophysiological processes involved in the pathological mechanism of vertigo remain to be fully understood. In this study, we used an online electrochemical system (OECS) and electrophysiological system to record the dynamics of ascorbate and the change in the neural excitability in medial vestibular nucleus (MVN) and hippocampus of rats following vertigo induced by vestibular ice water stimulation. The OECS employed for recording the dynamics of ascorbate consists of in vivo microdialysis and online selective electrochemical detection. We observed that the level of ascorbate in MVN increased to 155±14 % of the basal level after ice water vestibular stimulation. Whereas, the level of ascorbate in hippocampus decreased to 55±12 % of the basal level after ice water vestibular stimulation. To explore the neural excitability of MVN and hippocampus, we used an electrophysiological recording system and analyzed spontaneous firing rate (SFR) of the neuronal discharges in both nucleus. We found that, in MVN, the neural excitability began to increase during vestibular ice stimulation, remained to be elevated until 1 hour after stimulation. Whereas, in hippocampus, the neural excitability decreased soon after vestibular ice stimulation and recovered 1 hour after stimulation. Dynamics of ascorbate recorded with the OECS correlate with the change in the neural excitability, which may suggest the function of ascorbate as an antioxidant and possibly as a neuromodulator for glutamate‐mediated neurotransmission. The involvement of MVN and hippocampus for the dynamics of ascorbate observed here may be related to the vestibular compensation of vertigo.

Observation of Lidocaine‐suppressed Decrease of Magnesium in Salicylate‐induced Tinnitus with an Online Electrochemical System

AbstractTinnitus, characterized by an auditory perception in the absence of actual sound, is one of the most common encountered difficulties to manage otological problems, in which, lidocaine has been shown to provide temporary relief from subjective tinnitus. However, the pathophysiological mechanism of lidocaine treatment remains to be understood. The study demonstrates the first observation on that lidocaine well suppresses the decrease in the level magnesium (Mg2+) in inferior colliculus (IC) of guinea pigs following salicylate‐induced tinnitus with an online electrochemical system (OECS) for continuously monitoring of microdialysate of Mg2+. The OECS was established by efficiently coupling a selective electrochemical detector with in vivo microdialysis to monitor Mg2+ in the microdialysate continuously sampled from animal brain. The selective detection of Mg2+ over other coexisting neurochemicals was successfully achieved through a mechanism with divalent metal cations (e. g., Mg2+, Ca2+) enhanced catalytic current of organic dyes (i. e., polymerized film of toluidine blue O, p‐TBO) toward the oxidation of nicotinamide adenine dinucleotide (NADH) and with ethyleneglcol‐bis (2‐aminoethylether) tetraacetic acid (EGTA) as selective masking agents for Ca2+. Tinnitus was induced by intraperitoneal injection of sodium salicylate (350 mg/kg body weight). For the lidocaine group, using the microdialysis probe, 1 % lidocaine was perfused into IC of guinea pigs at 2 μL/min for 20 min. With the OECS, we found that the perfusion of lidocaine could recover the decrease of the Mg2+ level caused by salicylate‐induced tinnitus. The basal level of microdialysate Mg2+ in IC of guinea pigs was previously determined to be 1.29±0.44 mM, which was decreased significantly following salicylate‐induced tinnitus. The lidocaine perfusion significantly recovers the Mg2+ level by 37.7±14.8 %. This observation suggests that Mg2+ plays an important role in the pathological mechanism of tinnitus.

Online Electrochemical Monitoring of Glucose in Rat Brain with Acanthosphere‐like CuOOH Nanospheres‐based Electrochemical Sensor as Non‐enzymatic and O2‐independent Detector

AbstractThis study for the first time establishes a reliable non‐enzymatic online electrochemical system (OECS) for selective and continuous measuring glucose in animal cerebral system. We proposed a one‐step electrochemical method to synthetize a Cu nanostructure directly onto electrode, and over‐oxidize it in NaOH to form acanthosphere‐like CuOOH nanospheres (denoted as AN‐CuOOH), which can serve as an excellent electrochemical catalyst for glucose oxidation and thus for simple and reproducible non‐enzymatic glucose detection in strong alkaline solution. To validate the AN‐CuOOH‐based sensors for in vivo applications, we connect the sensors to a merged solution of in vivo microdialysis and 0.4 M NaOH to set up an OECS for in vivo glucose monitoring in rat brain. The response of this system has strong tolerance to the oxygen and pH fluctuation which may happen in the cerebral brain system during some physiological and pathological processes. The fabricated non‐enzymatic OECS has a good linear current response to glucose within a concentration range from 10 to 1000 μΜ with sensitivity of 130.22 nA ⋅ μΜ−1 cm−2. Additionally, the OECS is stable and does not suffer from the interference coexisting in the cerebral system. With the fabricated non‐enzymatic OECS for glucose measurement, the basal level of glucose in the microdialysate from rats striatum was determined to be 248.63 μΜ (n=3). Furthermore, the glucose was clearly decreased to 26.37 % of the basal level when the rat was undergone by 20 min ischemia, and the glucose concentration was gradually restored to 99.15 % during 40 min of reperfusion. Our work propose a technically simple strategy for the fabrication of a highly reproducible non‐enzymatic OECS system free from oxygen and pH interference, which is promising in exploring the molecular mechanism associated with brain functions.

Reduction of Ammineruthenium(III) by Sulfide Enables In Vivo Electrochemical Monitoring of Free Endogenous Hydrogen Sulfide.

The discovery of endogenous sulfide in mammalian brain opens up a door to understanding of the physiological function of hydrogen sulfide (H2S). The transformation of different forms of sulfide (i.e., S2-, HS-, H2S, bound sulfane sulfur, et al.) in various physiological conditions hurdles the direct detection of hydrogen sulfide in vivo. Here, we find that ammineruthenium(III) (Ru(NH3)63+) can catalyze the electrochemical oxidation of free sulfide including HS- and H2S in a neutral solution (pH 7.4). This property is used to constitute an electrochemical mechanism for selective detection of hydrogen sulfide. By coupling in vivo microdialysis with selective electrochemical detection, we successfully developed an integrated microchip-based online electrochemical system (OECS) for continuous monitoring of free endogenous hydrogen sulfide in the central nervous system (CNS). The microchip-based OECS is well responsive toward hydrogen sulfide with high stability, sensitivity and selectivity. Compared with the existing methods, the OECS does not require offline treatment of brain tissue or adjustment of the detection solutions into acidic or strong basic atmosphere. These priorities essentially enable the system to accurately and reliably track dynamics of hydrogen sulfide in the CNS.

Rational Design of Bioelectrochemically Multifunctional Film with Oxidase, Ferrocene, and Graphene Oxide for Development of in Vivo Electrochemical Biosensors.

This study demonstrates a new strategy to develop in vivo electrochemical biosensors through rational design and simple formation of bioelectrochemically multifunctional film (BMF). The BMF is rationally designed by first efficiently incorporating oxidase, ferrocene mediator, and graphene oxide into polymaleimidostyrene/polystyrene (PMS/PS) matrix to form a homogeneous mixture and then simply formed by drop-coating the mixture onto solid conducting substrate. By using the as-formed BMF, electrochemical biosensors could be constructed with a technical simplicity and high reproducibility. To illustrate the BMF-based biosensors for in vivo applications, we directly couple the biosensors to in vivo microdialysis to establish an online electrochemical system (OECS) for in vivo monitoring of glucose in rat auditory cortex during salicylate-induced tinnitus model. The OECS with the BMF-based biosensor as the detector shows a linear response toward glucose within a concentration range from 50 to 500 μM with a detection limit of 10 μM (S/N = 3). Additionally, the OECS is stable and does not suffer from the interference from the electroactive species endogenously coexisting in the brain microdialysate. With the BMF-based OECS, the basal level of glucose in the microdialysate continuously sampled from rat auditory cortex is determined to be 120 ± 10 μM (n = 5). After the rats were administrated with salicylate to induce transient tinnitus, the microdialysate glucose concentration in the rat auditory cortex remarkably increased to 433 ± 190 μM (n = 5) at the time point of 1.5 h. This study essentially offers a new, technically simple and reproducible approach to development of in vivo electrochemical biosensors, which is envisaged to be relatively useful for understanding of the molecular basis of brain functions.

An Online Electrochemical System for Continuously Monitoring Uric Acid Change following Rabbit Kidney following Ischemia-reperfusion Injury

In vivo monitoring of uric acid (UA) with high reliability and robustness is of great physiological and pathological importance because UA, as one kind of the most important chemical species in the biological systems, plays a great role in some physiological processes and is used as the biomarkers for early diagnosis of several diseases. In this study, a microfluidic chip-based online electrochemical system (OECS) is developed for the first time for continuously monitoring UA in rabbit kidney. To establish the OECS for UA monitoring, a single-channel microfluidic chip is developed into an electrochemical cell by incorporating a single-walled carbon nanotubes (SWNTs)-modified indium-tin oxide (ITO) electrode as working electrode, an Ag/AgCl wire as reference electrode, and a Pt tube as counter electrode. To completely remove ascorbic acid and thus eliminate its interference toward UA detection in such a single-channel microfluidic chip, ascorbate oxidase (AOx) is crosslinked onto both the etched ITO glass in the upstream channel and the SWNT-modified ITO electrode in the downstream. With the as-prepared microfluidic chip-based electrochemical cell as the detector for selective detection of UA, an online electrochemical system is successfully established by directly coupling the microfluidic chip-based electrochemical detector with in vivo microdialysis. The system exhibits a good linear response, high stability and good in vivo selectivity and is thus able to continuously sense UA change in rabbit kidney following ischemia-reperfusion injury. The study essentially offers a new OECS for in vivo continuous monitoring of UA.

An Online Electrochemical System for Continuous Measurement of Glutamate with Signal Amplification by Enzymatic Substrate Cycling

AbstractThis study demonstrates a new online electrochemical system (OECS) for continuous monitoring of glutamate by efficiently integrating in vivo microdialysis and selective electrochemical detection with an enhanced sensitivity through enzymatic substrate cycling. We find that the involvement of enzymatic substrate cycling in the biosensing scheme remarkably increases the sensitivity of the OECS toward the measurements of glutamate. Moreover, the OECS demonstrated here is stable, reproducible, and selective and could thus be potentially useful for continuous monitoring of glutamate release in central nervous system. This study essentially offers a new approach to the continuous measurements of cerebral glutamate release, which is believed to find interesting applications in understanding the molecular basis underlying brain functions.

A multi-enzyme microreactor-based online electrochemical system for selective and continuous monitoring of acetylcholine.

This study demonstrates an online electrochemical system (OECS) for selective and continuous measurements of acetylcholine (ACh) through efficiently integrating in vivo microdialysis, a multi-enzyme microreactor and an electrochemical detector. A multi-enzyme microreactor was prepared first by co-immobilizing two kinds of enzymes, i.e. choline oxidase (ChOx) and catalase (Cat), onto magnetite nanoparticles and then confining the as-formed nanoparticles into a fused-silica capillary with the assistance of an external magnet. The multi-enzyme microreactor was settled between an in vivo microdialysis sampling system and an electrochemical detector to suppress the interference from choline toward ACh detection. Selective detection of ACh was accomplished using the electrochemical detector with ACh esterase (AChE) and ChOx as the recognition units for ACh and Prussian blue (PB) as the electrocatalyst for the reduction of hydrogen peroxide (H2O2). The current recorded with the OECS was linear with the concentration of ACh (I/nA = -3.90CACh/μM + 1.21, γ = 0.998) within a concentration range of 5 μM to 100 μM. The detection limit, based on a signal-to-noise ratio of 3, was calculated to be 1 μM. Interference investigation demonstrates that the OECS did not produce an observable current response toward physiological levels of common electroactive species, such as ascorbic acid (AA), dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), and uric acid (UA). The high selectivity and the good linearity in combination with the high stability may enable the OECS developed here as a potential system for continuous monitoring of cerebral ACh release in some physiological and pathological processes.

Continuous and Simultaneous Electrochemical Measurements of Glucose, Lactate, and Ascorbate in Rat Brain Following Brain Ischemia

Developing new tools and technologies to enable recording the dynamic changes of multiple neurochemicals is the essence of better understanding of the molecular basis of brain functions. This study demonstrates a microfluidic chip-based online electrochemical system (OECS) for in vivo continuous and simultaneous monitoring of glucose, lactate, and ascorbate in rat brain. To fabricate the microfluidic chip-based detecting system, a microfluidic chip with patterned channel is developed into an electrochemical flow cell by incorporating the chip with three surface-modified indium-tin oxide (ITO) electrodes as working electrodes, a Ag/AgCl wire as reference electrode, and a stainless steel tube as counter electrode. Selective detection of ascorbate is achieved by the use of single-walled carbon nanotubes (SWNTs) to largely facilitate the electrochemical oxidation of ascorbate, while a dehydrogenase-based biosensing mechanism with methylene green (MG) adsorbed onto SWNTs as an electrocatalyst for the oxidation of dihydronicotiamide adenine dinucleotide (NADH) is employed for biosensing of glucose and lactate. To avoid the crosstalk among three sensors, the sensor alignment is carefully designed with the SWNT-modified electrode in the upstream channel and paralleled glucose and lactate biosensors in the downstream channels. With the microfluidic chip-based electrochemical flow cell as the detector, an OECS is successfully established by directly integrating the microfluidic chip-based electrochemical flow cell with in vivo microdialysis. The OECS exhibits a good linear response toward glucose, lactate, and ascorbate with less crosstalk. This property, along with the high stability and selectivity, enables the OECS for continuously monitoring three species in rat brain following brain ischemia.