D. C. Vanegas

A nanoceria–platinum–graphene nanocomposite for electrochemical biosensing

Most graphene-metal nanocomposites for biosensing are formed using noble metals. Recently, development of nanocomposites using rare earth metals has gained much attention. This paper reports on the development of a nanoceria-nanoplatinum-graphene hybrid nanocomposite as a base transducing layer for mediator-free enzymatic biosensors. The hybrid nanocomposite was shown to improve detection of superoxide or hydrogen peroxide when compared to other carbon-metal hybrid nanocomposites. Based on this finding, the nanocomposite was applied for biosensing by adding either a peroxide-producing oxidase (glucose oxidase), or a superoxide-producing oxidase (xanthine oxidase). Material analysis indicated that nanoceria and nanoplatinum were equally distributed along the surface of the hybrid material, ensuring detection of either superoxide or hydrogen peroxide produced by oxidase activity. Glucose biosensors demonstrated a sensitivity (66.2±2.6μAmM(-1)cm(-2)), response time (6.3±3.4s), and limit of detection (1.3±0.6μM) that were comparable to other graphene-mediated electrodes in the current literature. Remarkably, XOD biosensor sensitivity (1164±332μAmM(-1)), response time (5.0±1.5s), and limit of detection (0.2±0.1μM) were higher than any reported biosensors using similar metal-decorated carbon nanomaterials. This material is the first demonstration of a highly efficient, diverse nanoceria/nanoplatinum/graphene hybrid nanocomposite for biosensing.

A self-referencing biosensor for real-time monitoring of physiological ATP transport in plant systems

The objective of this study was to develop a self-referencing electrochemical biosensor for the direct measurement of ATP flux into the extracellular matrix by living cells/organisms. The working mechanism of the developed biosensor is based on the activity of glycerol kinase and glycerol-3-phosphate oxidase. A stratified bi-enzyme nanocomposite was created using a protein-templated silica sol gel encapsulation technique on top of graphene-modified platinum electrodes. The biosensor exhibited excellent electrochemical performance with a sensitivity of 2.4±1.8 nA/µM, a response time of 20±13 s and a lower detection limit of 1.3±0.7 nM. The self-referencing biosensor was used to measure exogenous ATP efflux by (i) germinating Ceratopteris spores and (ii) growing Zea mays L. roots. This manuscript demonstrates the first development of a non-invasive ATP micro-biosensor for the direct measurement of eATP transport in living tissues. Before this work, assays of eATP have not been able to record the temporally transient movement of ATP at physiological levels (nM and sub-nM). The method demonstrated here accurately measured [eATP] flux in the immediate vicinity of plant cells. Although these proof of concept experiments focus on plant tissues, the technique developed herein is applicable to any living tissue, where nanomolar concentrations of ATP play a critical role in signaling and development. This tool will be invaluable for conducting hypothesis-driven life science research aimed at understanding the role of ATP in the extracellular environment.

Leaf Extract from Lithocarpus polystachyus Rehd. Promote Glycogen Synthesis in T2DM Mice

The purpose of this study was to investigate the effects of leaf extract from Lithocarpus polystachyus Rehd. on type II diabetes mellitus (T2DM) and the active ingredients of this effect. In addition, this study determined, for the first time, the underlying molecular and pharmacological mechanisms of the extracts on hyperglycemia using long-term double high diet-fed and streptozotocin (STZ) induced type II diabetic mice. In the present study, leaf extract, phloridzin and trilobatin were assessed in vivo (gavage) and in vitro (non-invasive micro-test technique, NMT) in experimental T2DM mice. The biochemical parameters were measured including blood glucose and blood lipid level, liver biochemical indexes, and hepatic glycogen. The relative expression of glycometabolism-related genes was detected. The effect of leaf extracts on physiological glucose flux in liver tissue from control and T2DM mice was also investigated. Body weight of experimental T2DM mice increased significantly after the first week, but stabilized over the subsequent three weeks; body weight of all other groups did not change during the four weeks’ study. After four weeks, all treatment groups decreased blood glucose, and treatment with leaf extract had numerous positive effects: a) promoted in glucose uptake in liver, b) increased synthesis of liver glycogen, c) reduced oxidative stress, d) up-regulation of glucokinase (GK), glucose transporter 2 (GLUT2), insulin receptor (IR) and insulin receptor substrate (IRS) expression in liver, e) down-regulation of glucose-6-phosphatase (G-6-P) expression, and f) ameliorated blood lipid levels. Both treatment with trilobatin or phloridzin accelerated liver glycogen synthesis, decreased oxidative stress and increased expression of GK. IRS and phosphoenolpyruvate carboxykinase (PEPCK) were both up-regulated after treatment with trilobatin. Expression of GLUT2, PEPCK and G-6-P were also increased in liver tissue after treatment with phloridzin. Our data indicate that leaf extract from L. polystachyus Rehd. has a preferable hypoglycemic effects than trilobatin or phloridzin alone. Leaf extract significantly increased glucose uptake and hepatic glycogen synthesis while also inducing a decline of hepatic gluconeogenesis and oxidative stress in T2DM mice. From this study, we draw conclusions that L. polystachyus promoted glycogen synthesis in T2DM mice, and that the active compounds were not only the trilobatin or phloridzin.

Emerging Biorecognition and Transduction Schemes for Rapid Detection of Pathogenic Bacteria in Food

The presence of unsafe levels of microorganisms in food constitutes a growing economic and public health problem that necessitates new technology for their rapid detection along the food continuum from production to consumption. While traditional techniques are reliable, there is a need for more sensitive, selective, rapid, and cost-effective approaches for food safety evaluation. Methods such as microbiological counts are sufficiently accurate and inexpensive, and are capable of determining presence and viability for most pathogens. However, these techniques are time consuming, involve destructive sampling, and require trained personnel and biosafety-certified facilities for analysis. Molecular techniques such as the polymerase chain reaction have greatly improved analytical capability over the last decade, achieving shorter analysis time with quantitative data and strain specificity, and in some cases the ability to discriminate cell viability. The emerging field of nanosensors/biosensors has demonstrated a variety of devices that hold promise to bridge the gap between traditional plate counting and molecular techniques. Many nanosensors/biosensors are rapid, portable, accurate devices that can be used as an additional screening tool for identifying unsafe levels of microorganisms in food products with no need for pre-enrichment. In this review, we provide a brief overview of available biorecognition-transduction techniques for detecting bacteria in food. We then discuss the advantages and disadvantages of each technique, and describe some recent biosensor or nanosensor technologies that are under development. We conclude by summarizing the opportunities and challenges in the field of pathogen monitoring in food systems and we focus the discussion on the strengths/weaknesses of the most popular biorecognition agents and transducer nanomaterials for biosensing.

Biosensors for Indirect Monitoring of Foodborne Bacteria

Microbiological safety assessment in food samples is often a cumbersome and time consuming task that has to be done in a regular basis in most food manufacturing facilities. Standard microbiological techniques for evaluating critical levels of bacteria take several days to yield results, thus the food products have to be withheld in the processing factories until the safety inspection is concluded. The food industry requires faster and reliable tools for more efficient microbiological testing that can reduce the safety inspection lag significantly. Biosensors that target specific molecules associated with bacterial activity are amongst the most promising technologies for addressing this need. State-ofthe- art biosensors allow for instantaneous quantification of molecular biomarkers. This review focuses on describing the operation mechanism and technological challenges and opportunities for developing high performance biosensors capable of real-time monitoring of biomarkers from foodborne bacteria.

Lignin and silicate based hydrogels for biosensor applications

Advances in biocompatible materials and electrocatalytic nanomaterials have extended and enhanced the field of biosensors. Immobilization of biorecognition elements on nanomaterial platforms is an efficient technique for developing high fidelity biosensors. Single layer (i.e., Langmuir–Blodgett) protein films are efficient, but disadvantages of this approach include high cost, mass transfer limitations, and Vromer competition for surface binding sites. There is a need for simple, user friendly protein-nanomaterial sensing membranes that can be developed in laboratories or classrooms (i.e., outside of the clean room). In this research, we develop high fidelity nanomaterial platforms for developing electrochemical biosensors using sustainable biomaterials and user-friendly deposition techniques. Catalytic nanomaterial platforms are developed using a combination of self assembled monolayer chemistry and electrodeposition. High performance biomaterials (e.g., nanolignin) are recovered from paper pulp waste and combined with proteins and nanomaterials to form active sensor membranes. These methods are being used to develop electrochemical biosensors for studying physiological transport in biomedical, agricultural, and environmental applications.

Laser Scribed Graphene Biosensor for Detection of Biogenic Amines in Food Samples Using Locally Sourced Materials

In foods, high levels of biogenic amines (BA) are the result of microbial metabolism that could be affected by temperatures and storage conditions. Thus, the level of BA is commonly used as an indicator of food safety and quality. This manuscript outlines the development of laser scribed graphene electrodes, with locally sourced materials, for reagent-free food safety biosensing. To fabricate the biosensors, the graphene surface was functionalized with copper microparticles and diamine oxidase, purchased from a local supermarket; and then compared to biosensors fabricated with analytical grade materials. The amperometric biosensor exhibits good electrochemical performance, with an average histamine sensitivity of 23.3 µA/mM, a lower detection limit of 11.6 µM, and a response time of 7.3 s, showing similar performance to biosensors constructed from analytical grade materials. We demonstrated the application of the biosensor by testing total BA concentration in fish paste samples subjected to fermentation with lactic acid bacteria. Biogenic amines concentrations prior to lactic acid fermentation were below the detection limit of the biosensor, while concentration after fermentation was 19.24 ± 8.21 mg histamine/kg, confirming that the sensor was selective in a complex food matrix. The low-cost, rapid, and accurate device is a promising tool for biogenic amine estimation in food samples, particularly in situations where standard laboratory techniques are unavailable, or are cost prohibitive. This biosensor can be used for screening food samples, potentially limiting food waste, while reducing chances of foodborne outbreaks.

Bio-inspired patterned networks (BIPS) for development of wearable/disposable biosensors

Here we demonstrate a novel approach for fabricating point of care (POC) wearable electrochemical biosensors based on 3D patterning of bionanocomposite networks. To create Bio-Inspired Patterned network (BIPS) electrodes, we first generate fractal network in silico models that optimize transport of network fluxes according to an energy function. Network patterns are then inkjet printed onto flexible substrate using conductive graphene ink. We then deposit fractal nanometal structures onto the graphene to create a 3D nanocomposite network. Finally, we biofunctionalize the surface with biorecognition agents using covalent bonding. In this paper, BIPS are used to develop high efficiency, low cost biosensors for measuring glucose as a proof of concept. Our results on the fundamental performance of BIPS sensors show that the biomimetic nanostructures significantly enhance biosensor sensitivity, accuracy, response time, limit of detection, and hysteresis compared to conventional POC non fractal electrodes (serpentine, interdigitated, and screen printed electrodes). BIPs, in particular Apollonian patterned BIPS, represent a new generation of POC biosensors based on nanoscale and microscale fractal networks that significantly improve electrical connectivity, leading to enhanced sensor performance.

Xanthine oxidase biosensor for monitoring meat spoilage

In this study, we have designed an electrochemical biosensor for real-time detection of specific biomarkers of bacterial metabolism related to meat spoilage (hypoxanthine and xanthine). The selective biosensor was developed by assembling a ‘sandwich’ of nanomaterials and enzymes on a platinum-iridium electrode (1.6 mm tip diameter). The materials deposited on the sensor tip include amorphous platinum nanoclusters (i.e. Pt black), reduced graphene oxide, nanoceria, and xanthine oxidase. Xanthine oxidase was encapsulated in laponite hydrogel and used for the biorecognition of hypoxanthine and xanthine (two molecules involved in the rotting of meat by spoilage microorganisms). The developed biosensor demonstrated good electrochemical performance toward xanthine with sensitivity of 2.14 ± 1.48 μA/mM, response time of 5.2 ± 1.5 sec, lower detection limit of 150 ± 39 nM, and retained at least 88% of its activity after 7 days of continuous use.

Non-invasive microsensors for studying cell/tissue physiology

Non-invasive tools that allow real-time quantification of molecules relevant to metabolism, homeostasis, and cell signaling in cells and tissue are of great importance for studying physiology. Several microsensor technologies have been developed to monitor concentration of molecules such as ions, oxygen, electroactive molecules (e.g., nitric oxide, hydrogen peroxide), and biomolecules (e.g., sugars, hormones). The major challenges for microsensors are overcoming relatively low sensitivity and low signal-to-noise ratio. Modern approaches for enhancing microsensor performance focus on the incorporation of catalytic nanomaterials to increase sensitivity, reduce response time, and increase operating range. To improve signal-to-noise ratio, a non-invasive microsensor modality called self-referencing (SR) is being applied. The SR technique allows measurement of temporal and spatial transport dynamics at the cell, tissue, organ, and organismal level.