Membraneless enzymatic biofuel cells based on graphene nanosheets

Abstract

The possibility of employing graphene sheets as a potential candidate for the construction of biofuel cells is reported in this paper. Initially, graphene sheets were chemically synthesized and characterized by surface characterization techniques. Following this, graphene was employed to fabricate the anode and cathode in the biofuel cell. The anode of the biofuel cell consists of a gold electrode on which we co-immobilized graphene – glucose oxidase using silica sol–gel matrix. Voltammetric measurements were conducted to quantitatively evaluate the suitability of employing graphene sheets as an electrode dopant and its performance was compared with single walled carbon nanotubes (SWCNTs). The cathode of the biofuel cell was constructed in a similar method except that graphene was co-immobilized with bilirubin oxidase. Finally, two membraneless enzymatic biofuel cells, one using graphene sheets and the other using SWCNTs, were constructed and their performances were compared. Upon comparison, graphene based biofuel cell exhibited a maximum power density of about 24.3 ± 4 μW (N = 3), which is nearly two times greater than that of the SWCNTs biofuel cell, and the performance of the graphene biofuel cell lasted for 7 days.

Introduction

In recent years, there has been considerable interest towards the development of enzymatic biofuel cells (EBFC) as they can be employed as an in vivo power source for implantable medical devices such as pacemakers, micro drug pumps, deep brain stimulators, etc. (Gao et al., 2007, Barton et al., 2004, Bullen et al., 2006, Kim et al., 2006, Ikeda and Kano, 2003). The most attractive feature of this EBFC is that they can utilize glucose or other carbohydrates abundantly present in the human body as a fuel. To date, there have been considerable efforts among many researchers to fabricate practical EBFC devices out of different theoretical concepts (Mano et al., 2002, Willner, 2002, Service, 2002, Katz et al., 2005, Katz and Willner, 2003). Glucose, a major component of the human serum, is most widely used as the fuel for theses EBFCs. However, low power density and poor stability of the EBFC are the two major challenges to be rectified in the upcoming days. A non-compartmentalized glucose|O₂ biofuel cell possessing a maximum power of 4 μW cm⁻² and a life time of 48 h was reported (Katz et al., 1999), while an abiotically catalyzed glucose fuel cell exhibited a maximum power density of 3.3 μW cm⁻² with a life time of 224 days (Kerzenmachera et al., 2008). In another study, a cell lifetime of up to 45 days was reported with enzymes entrapped in a modified Nafion membrane (Moore et al., 2004). Furthermore, an EBFC employing glucose oxidase (GOx) and laccase as biocatalysts generated a maximum power density of 5.49 μW cm⁻² using glucose as a fuel (Liu and Dong, 2007).

The low power density of the EBFC in comparison with conventional inorganic fuel cells is due to location of the active site of the enzyme buried deep under the protein shell hindering the electron transfer pathway between the enzyme's active site and the electrode (Liu and Dong, 2007). In order to overcome this issue, most researchers employ carbon nanotubes (CNTs) to decrease the electron transfer resistance and increase the electrode surface area (Gao et al., 2007, Li et al., 2008, Lim et al., 2007, Liu and Dong, 2007). The covalent binding of the enzyme with CNTs has resulted in a faster electron transfer rate. However, a complex chemical treatment process of CNTs has to be performed in order to create active binding sites on the edge of the CNTs. Such a process hinders the mass production of this electrode (Li et al., 2005, Imamura et al., 1995, Degani and Heller, 1988). Furthermore, a number of redox mediators are widely used to boost the electron transfer rate. The redox potential of the mediator used should lie between the redox potential of the enzyme and that of the electrode. As a result, the electrons are gradually shuttled from the enzyme to the mediator and then to the electrode (Lim et al., 2007).

Graphene, a two-dimensional (2D) nanostructure of carbon discovered in 2004, possesses a very large surface area of about 2630 m² g⁻¹, which is about the size of a football stadium (Stoller et al., 2008). Further, the electrons on the graphene surface move ballistically over the sheet without any collisions with mobilities as high as 10,000 cm² V⁻¹ s⁻¹ at room temperature (Geim and MacDonald, 2007, Novoselov et al., 2004). In addition, graphene was found to exhibit an excellent conductivity. In our previous work, a four-point probe method was used to measure the conductance of graphene and it was calculated to be 64 mS cm⁻¹, which is approximately 60 times more than that of SWCNTs (Alwarappan et al., 2009, Dai et al., 2007). It is also worth mentioning that graphene possesses a number of surface active functional moieties such as carboxylic, ketonic, quinonic and CC. Of these, the carboxylic and ketonic groups are reactive and can easily bind covalently with GOx. The presence of extended CC conjugation in graphene is also expected to shuttle electrons. Nonetheless, to the best of our knowledge, there are no reports available in literature which employs graphene as an electrode material for EBFC.

The biocompatible sol–gel encapsulation method is widely preferred to immobilize biomolecules such as proteins, nucleic acids and cells. The microstructured porous sol–gel matrix prevents the biomolecules from being denatured by pH or temperature and thus retains their long-term bioactivity (Kandimalla et al., 2006). On the other hand, the porous structure of the sol–gel matrix allows diffusion of the fuel towards the electrode surface for the redox reaction to occur.

In the present work, we describe an EBFC system essentially based on silica sol–gel immobilized graphene sheets/enzyme composite electrodes. Further, we employ glucose oxidase (GOx) and bilirubin oxidase (BOD) as the anodic and cathodic enzyme, respectively (Komaba et al., 2008, Kuwahara et al., 2008, Kuwahara et al., 2009, Willner et al., 2009;). In addition, we employ ferrocenemethanol (FM) and 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) (Lim et al., 2007) as anodic and cathodic mediators, respectively. Due to the specific catalytic activity of these enzymes, the proton exchange membrane is eliminated to facilitate the fabrication process. After the fabrication of graphene based membraneless EBFC, the maximum power density is found to be 24.3 ± 4 μW (N = 3). After 7 days, the power output dropped to 50% of its original power output. For comparison, a similar EBFC system was constructed using single walled carbon nanotubes (SWCNTs) and the power output was measured by the same method.