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SUBMITTED BY A.Nakkiran CECRI
Sensor technology is one of the most important key technologies of the future with a constantly increasing number of applications, both in the industrial and in the private sectors. More and more bio sensors are used for the control of processes in environment monitoring, healthcare, and Military (against bioweapons). Consequently, the development of fast and sensitive bio sensors is the subject of intense research, propelled by strategies based on nanoscience and nanotechnology. This paper highlights the recent developments and reflects the impact of nanoscience on electrochemical sensor technology by elucidates size dependent sensitivity of gold electrode to sense dopamine, an important neurotransmitter.
Introduction According to the modern definition, biosensors are analytical devices comprising a biological or biologically-derived sensing element either integrated within or intimately associated with a physicochemical transducer1 (Fig1). The transducer is an important component in a biosensor through which the measurement of the target analyte(s) is achieved by selective transformation of a biomolecule-analyte interaction into a quantifiable electrical or optical signal. A wide range of optical and electrochemical instruments have been employed in conjunction with biological sensing.
Figure 1 Schematic diagram showing the main components of a biosensor (a) A bio-component, (b) a transducer which converts the biochemical reaction into a physical signal, (c) an amplifier which converts a physical signal into an electrical signal, which is processed and displayed by a recorder or PC (d).
Electrochemical transducers detect an electrochemical signal that is generated by the interaction between the analyte and the receptor. It could be a change of a redox potential (Potentiometric), the conductivity of the solution (conductometric) or the production of redox active molecules that generate a current (voltammetric, amperometric, coulometric).
Dopamine and Electrochemistry Dopamine is an important neurotransmitter because it is involved in physical and cognitive functions. To understand the challenges associated with measuring dopamine it is important to understand the environment in which dopaminergic neurons function. Neuron Function • Nerve cells, or neurons, are the basic building blocks of the nervous system. The neuron is responsible for sending and receiving nerve impulses or messages. A neuron that is excited will transmit its energy to neurons that are next to it (Fig.2).
Figure 2: Transformations message through neurons
•Neurons have a central cell body attached to slender, branching arms. There are two types of “arms”: dendrites are like antennae and carry messages, or impulses, to the cell body, while axons carry messages away from the cell body. • Impulses travel from neuron to neuron from the axon of one cell to the dendrites of another by crossing over a tiny gap between the two nerve cells called a synapse.
• Incoming messages from the dendrites are passed to the end of the axon where sacs containing neurotransmitters (dopamine) open into the synapse. • The dopamine molecules cross the synapse and fit into special receptors on the receiving cell. • That cell is stimulated to pass the message on • After the message is passed on, the receptors release the dopamine particles back into the synapse where the excess dopamine is “taken up” or recycled within the releasing neuron So inadequately stimulation of dopamine (DA) can cause fatal disease such as Parkinson’s and schizophrenia. Recent clinical studies have demonstrated that the content of dopamine in biological fluids can be used to assess the amount of oxidation stress in human metabolism and excessive oxidative stress has been linked to cancer, diabetes mellitus, and hepatic disease. Electrochemical Detection Dopamine (DA), the most important among the class of neurotransmitters, plays an important role in the function of the central nervous system. The development of methods for dopamine quantification in nerves and biological fluids is the subject of intense current investigation in neurochemical studies.Electrochemical method is an ideal choice for the quantitative determination of dopamine, because •Dopamine is easily oxidizable •R. MARK WIGHTMAN Research group in University of North Carolina has employed in-vivo voltammetry to measure the dopamine release and uptake in freely moving animals and found that a behavioral stimulus can evoke a transient increase in dopamine, providing how a neurotransmitter controls behavior on second and sub second timescales and revealing how critical rapid, selective, and sensitive measurements for real-time detection of chemical changes in the brain(Fig 3)
Figure 3 X-Ray image of microelectrode in brain
But Interference due to the co-existence of ascorbic acid (AA) in the biological fluids, which also undergoes oxidation more or less at the same potential, is the major flaw in dopamine sensor. Also, the concentration of AA is relatively higher than that of DA in these samples (103 times higher than DA), which results in poor selectivity and sensitivity for DA detection.So the detection of DA in the presence of excess of AA is a challenging task in electro analytical research.
Chemicals Hydrogen tetrachloroaurate, dopamine (DA) and ascorbate (AA) were obtained from Aldrich and were used as received. All other chemicals used in this investigation were of analytical grade and were used without further purification. The phosphate buffer solution (PBS) was prepared from NaH2PO4 and Na2HPO4 (0.1 M). Preparation of Au colloids Au colloids were prepared according to the literature. Typically, 1 ml of 1% HAuCl4 was added to 90ml of water at room temperature. After 1 min of stirring, 2 ml of 38.8 mM sodium citrate was added. Subsequently,1 ml of freshly prepared 0.075% NaHB4 in 38.8mM sodium citrate was added and the colloidal solution was stirred for another 5-10 min and stored in a dark bottle at 4 8C. The concentration of the Au nanoparticles was estimated to be 0.32 mM. Immobilization of Au colloidal particles The Au electrodes of 1.6 mm diameter were polished with alumina powder (1.0 and 0.06 mm) and sonicated in water for 5-10 min. The polished electrodes were then electrochemically cleaned by potential cycling between /0.2 and 1.5 V at a scan rate of 10 V s-1 in 0.05 M H2SO4 for 10 min or until the cyclic voltammogram characteristic for a clean Au electrode was obtained. The electrochemically cleaned Au electrode was immersed into an aqueous solution of 10 mM of amine-terminated monolayer of cystamine (CYST) for 1 h. The CYST monolayer-modified electrode was rinsed well with water and kept in water for at least 30 min to remove the physically adsorbed CYST. The CYST electrode was subsequently soaked in the Au colloidal solution for 12 h. The resulting electrode was washed with copious amount of water and subjected to electrochemical experiments. Here after the Au nanoparticle-immobilized electrodes will be referred as the nano-Au electrode (fig.4).
Figure 4 Schematic representation of the fabrication of the nano-Au self-assembly (note that this is a pictorial representation and is not on the correct scale). 1) Bare gold electrode 2) CYSTMINE modified –Au electrode 3) Au nanoparticle immobilized electrode 4) TEM of Au nanoparticle immobilized electrode
RESULT AND DISSCUSSION Fig. 5 shows the SW voltammograms obtained for DA at the bare and nano-Au electrodes. The voltammetric response at the bare electrode is rather broad, whereas it is sharp and well defined at the nano-Au electrode, suggesting that the electrochemical behavior of the nano-Au electrode is quite different from the bulk Au electrode. Furthermore, as can be readily seen from this figure, the peak current at the nano-Au electrode is significantly larger compared to that of the bare electrode. For instance a 1.7fold enhancement in the peak current at the nano-Au electrode was observed, which indicates that the nano-Au electrode possesses excellent sensitivity towards DA.
Fig. 5 Square-wave voltammograms obtained for the oxidation of DA (50 mM) at the bare Au (a) and nano-Au (b) electrodes in 0.1 M PBS (pH 7.2). Since ascorbic acid (AA) is the major interferent in the voltammetric measurement of DA, its voltammetric behavior at the Au nano-assembly was studied. Fig. 6 shows the cyclic voltammograms obtained for the oxidation of AA at the nano-Au electrode at different scan rates. At the nano-Au electrode the oxidation occurs at around 0.03 V and an enormous increase in the peak current compared to the bare electrode was observed. These results indicate that the nano-Au electrode effectively catalyzes the oxidation of AA. It has been reported quite recently that nanometer-sized Au particles exhibit excellent electro catalytic activity.
Fig. 6 Cyclic voltammograms obtained for the oxidation of AA (100mM) at the nano-Au electrode in 0.1MPBS (pH 7.2). Scan rate: 25, 50, 75, 100, 125, 150 and 175 mV s_1.
The important attribute of the nano-sized catalysts is the • High surface area and •Interface-dominated properties that differs from the atomic, molecular and bulk counterpart. In the present investigation the facilitated oxidation of ascorbic acid (AA) at the nano-Au electrode is believed to be due to the excellent catalytic activity of nano-sized Au particles. Because the main objective of the present investigation is the determination of DA in the presence of AA, our attention is focused on the voltammetric detection of DA in the presence of AA.
Figure 7 Cyclic voltammograms of a binary mixture solution of AA and DA at the bare Au (a) and nano-Au (b) electrodes (pH 7.2). Scan rate: 100 mV s-1. (B) Corresponding square-wave voltammograms obtained at bare Au (a) and Nano-Au (b) electrodes. Fig. 7 shows the cyclic and SW voltammograms obtained for DA and AA coexisting at bare and nano- Au electrodes. We can see that the bare electrode cannot separate the voltammetric signals of AA and DA. Only one broad voltammetric signal was observed for both analytes and the voltammetric peak decreased in the subsequent sweeps.
Therefore it is impossible to use the bare electrode for the voltammetric determination of DA in the presence of AA. But, the nano-Au electrode resolved the mixed voltammetric signals into two well-defined voltammetric peaks at 0.015 and 0.185 V corresponding to the oxidations of AA and DA, respectively. The nano-Au electrode shows good selectivity and excellent sensitivity in the detection of DA in the presence of AA.AA is readily oxidized well before the oxidation potential of DA is reached. Thus the catalytic oxidation of AA by the oxidized DA is completely eliminated and the precise determination of DA in the presence of AA is possible at the nano-Au electrode. The voltammetric signals of AA and DA remained unchanged in the subsequent sweeps, indicating that the nano-Au electrode does not undergo surface fouling. Furthermore, the separation between the voltammetric peaks of AA and DA is large (165 mV) and thus the simultaneous determination of AA and DA or the selective determination of DA in the presence of AA is feasible at the nano-Au electrode Insights on Influence of Particle Size on Electrochemical bio-sensing The result shows that nano-sized Au is largely different from the bulk counterpart and it shows a surprisingly high electro catalytic activity. The nano-Au electrode successfully distinguishes the voltammetric signals of AA and DA, which are indistinguishable at the bare Au electrode. Because each incorporated nanoparticle operate as an individual electrode (electrode of nanosize), and act as an active site for interfacial electron transfer. Electron transfer at nanoscale electrodes is much different from that of bulk, because reducing the electrode size increases the diffusion-controlled transport rate in steady-state voltammetric measurements. Consider the simple electrontransfer reaction Ox + e– Red
Occurring at a spherical electrode of radius a, Diffusion of Ox to the electrode surface occurs before the electron-transfer step, and either step may be rate-limiting. At steady state, the rate constant for diffusion (cm/s) of redox molecules to the spherical electrode is simply D/a, in which D is the diffusion constant (cm2/s) of Ox. Since a is in
nanometer, D/a is comparable to or significantly larger than the standard electron-transfer rate constant ket (cm/s). D/a ≥ket So the overall rate of the electrode reaction is solely controlled by the interfacial electron-transfer step (i.e. transport rate of analyte from the bulk of the solution to the interface is not accounted).Hence nanoelectrode open up possibilities for work in very low concentration (nanomolar) of analyte.whatever can be done at a planar electrode can be done at concentration of about 106 times lower by using nanoelectrode without reaching the limiting current. But in bulk electrode the reaction is under diffusion control, so low concentration analyte reactions are not easily distinguishable from the high concentrated interference molecule. Hence reason for the high sensitivity of nanoelectrode is well explained. CONCLUSION
The present work reveals the fact that; size miniaturization is the principal cause for high selectivity and high sensitivity of DA sensor. Also proposed reason soundly corroborates the obtained result. Thus the potential application of nanosized Au for the fabrication of a voltammetric DA sensor is demonstrated.
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