Nanocrystalline diamond modified gold electrode for glucose biosensing

Abstract

Boron-doped diamond has drawn much attention in electrochemical sensors. However there are few reports on non-doped diamond because of its weak conductivity. Here, we reported a glucose biosensor based on electrochemical pretreatment of non-doped nanocrystalline diamond (N-NCD) modified gold electrode for the selective detection of glucose. N-NCD was coated on gold electrode and glucose oxidase (GOx) was immobilized onto the surfaces of N-NCD by forming amide linkages between enzyme amine residues and carboxylic acid groups on N-NCD. The anodic pretreatment of N-NCD modified electrode not only promoted the electron transfer rate in the N-NCD thin film, but also resulted in a dramatic improvement in the reduction of the dissolved oxygen. This performance could be used to detect glucose at negative potential through monitoring the current change of oxygen reduction. The biosensor effectively performs a selective electrochemical analysis of glucose in the presence of common interferents, such as ascorbic acid (AA), acetaminophen (AP) and uric acid (UA). A wide linear calibration range from 10 μM to 15 mM and a low detection limit of 5 μM were achieved for the detection of glucose.

Introduction

Due to the various electron orbit characteristics and the anisotropy of carbon element, carbon materials contain characteristics of nearly all the matters on the earth, thus attracting intense attention of scientists and engineers. A large degree of carbon materials have been employed for the development of biosensors. Glassy carbon (GC), carbon paste, graphite and carbon fibres have been well established for many years as electrode materials (Cosnier et al., 1999, Kulys, 1999, Horozova et al., 2000, Netchiporouk et al., 1995). These materials allow for the easy enzyme immobilization and possess reproducible electrochemical behavior. Recently, novel carbon materials, such as carbon fullerenes, carbon nanotubes (CNT) and diamond lead new directions in the design of biosensors (Nednoor et al., 2004, Poh et al., 2004). They offer lots of possibilities for the biosensor construction, for example, electron mediation, nano-biosensor fabrication and enzyme entrapment.

Diamond-based materials, because of their unique properties such as biocompatibility, chemical inertness, chemical stability, wide potential window and optical transparency properties, which are superior to the traditional carbon materials, e.g. GC and highly oriented pyrolytic graphite (HOPG), have attracted increasing attention in biosensor and biochip applications (Haymond et al., 2002, Lora Huang and Chang, 2004, Weng et al., 2005). The functionalized surfaces of diamond are excellent for the self-assembly of proteins. Hamers and coworkers employed nanocrystalline diamond thin films for DNA hybridization via photochemical modification of diamond surfaces, and showed that both as semiconductors, nanocrystalline diamond has superior properties over the crystalline silicon for the DNA hybridization (Yang et al., 2002, Strother et al., 2002, Strother et al., 2003). Proteins such as cytochrome c, antibodies and enzymes also have been immobilized on diamond films for biosensor applications (Lora Huang and Chang, 2004, Huang et al., 2004, Su et al., 2004). Since for electrochemical applications, the diamond thin film must possess a good conductivity, most work in this field has been carried out using diamond layers doped with boron (Levy-Clement et al., 1999), which display almost metal-like properties. Compton and co-workers investigated the properties of non-doped nanocrystalline diamond (N-NCD) film and found that compared to boron-doped diamond, it is a highly active electrode material, with low over potentials and high adherence of metallic electrodeposites, for all redox systems studied (Hian et al., 2003a, Hian et al., 2003b). However, as an electrode material, the electrical conductivity of N-NCD film is not high enough.

Carbon materials have a great improvement in the electron-transfer properties of redox systems and an enhanced electrochemical activity after the electrochemical pretreatment. In the previous works, HOPG electrode and GC electrode have been used for the investigation of the specific phenomenon. McCreery and co-workers have reported that the anodization resulted in the destruction of the basal plane lattice structure within the first few nanometers of the surface and the emergence of an amorphous layer rich in edge plane defects, which induced the electron-transfer activation (Bowling et al., 1989a, Bowling et al., 1989b). Goss et al. (1993) also investigated the electrochemical pretreatment of HOPG and provided atomic force microscopy (AFM) images to show modest overall changes in the HOPG surface lattice in the former case and extensive lattice damage with exposure of edge plane sites in the latter case. For the GC electrode, the reason of increasing activity after pretreatment is unclear. Engstrom (1982) suggested that the pretreatment introduced or altered the nature of functional groups on the electrode surface, which might serve as mediators of electrons between the electrode and the eletroactive species. Quinone functionalities appeared to be candidates as mediators, since there is substantial evidence suggesting the presence of such groups on anodized carbon surfaces (Evans et al., 1977, Evans and Kuwana, 1977). The same authors reported the probability that electrochemical pretreatment cleaned the surface of GC electrode with contaminants introduced in the polishing stage of electrode preparation and changed the chemical nature of the GC surface (Engstrom and Strasser, 1984). It has been proposed that on carbon paste electrodes, anodization causes the removal of organic pasting liquid from the carbon surface, making the electrode surface more hydrophilic and therefore more accessible to the species in solution (Rice et al., 1983). Recently, some rising carbon materials have drawn lots of attention and were investigated for this phenomenon. Joseph Wang and co-workers have reported the electrochemical activation of CNT prepared by different process. The anodic pretreatment results in a dramatic improvement in the electrochemical reactivity of the ARC-produced CNT, while the chemical vapour deposition (CVD) produced CNT appeared to be resistant to the anodic activation. Differences in the effect of the electrochemical pretreatment are attributed to the preanodization effectively breaking the basal-plane end caps of ARC–CNT thereby exposing edge plane defects, similar to those already present in the open-end capes of CVD–CNT (Musameh et al., 2005). Fujishima and co-workers have demonstrated that anodically pretreated boron-doped diamond electrode could be used for the direct electrochemical oxidation of disulfides such as glutathione disulfide (GSSG) and glutathione (GSH), which are difficult to oxidize at other kinds of electrodes. The oxygen functional groups formed on the anodically oxidized diamond surface are believed to facilitate the attractive electrostatic interaction between the oxygenated surface and the positively charged GSSG (Terashima et al., 2003). However, the electrochemical pretreatment of N-NCD thin-film has not been introduced to the construction of biosensor, to our knowledge.

In the present work, we reported a glucose biosensor based on the immobilization of N-NCD on the surface of gold electrode. N-NCD was physically adsorbed onto the polyelectrolyte-coated electrode and GOx was covalently immobilized on the surface of N-NCD with activated surface COOH sites through amide linkages. We then investigated the properties of N-NCD thin-film after the electrochemical pretreatment. After preanodized at 0.7 V versus SCE, the electron-transfer rate in the film of N-NCD is enhanced and the prepared biosensor displays a great improvement in the reduction of dissolved oxygen. An amperometric biosensor based on monitoring the reduction current of oxygen was made and a high selectivity in the glucose detection was achieved.