IEEE TRANSACTIONS ON NANOTECHNOLOGY, VOL. 10, NO. 1,JANUARY 2011 59 Surface Engineering of Graphene-Enzyme Nanocomposites for Miniaturized Biofuel Cell Chang Liu, Zhongfang Chen, and Chen-Zhong Li Abstract—A novel approach to the surface functionalization for membraneless enzymatic glucose/oxygen biofuel cell applications is described. The biofuel cell employs the gold plate electrodes modi- fied by specific graphene-enzyme conjugations, which are immobi- lized by electrochemical deposition of the conducting polypyrrole polymer. The electrochemical activity of these electrodes is supe- rior to the electrodes immobilized with sol–gel. Such enhancements can be attributed to the excellent electrical property and enzyme loading capability of the polypyrrole material. The power output and the biostability of the integrated biofuel cell are also improved. Index Terms—Biofuel cell, enzymatic fuel cell, graphene nanosheets, polypyrrole, surface engineering. I. INTRODUCTION B IOFUEL cells are electrochemical power generators that are able to convert chemical energy into electrical energy through redox reactions. In recent years, the development of enzymatic biofuel cells (EBFC) is likely to have a significant impact on homeland security, aerospace, and healthcare industries as the EBFCs can produce higher power output than other types of biofuel cells [1]–[4]. The output power of such EBFCs is well sufficient to supply some microscale electronic systems, such as cameras for remote surveillance, transmitters, actuators, or even wireless sensor networks, which paves the path for the U.S. Army’s goal to eliminate all army military batteries or at least reduce the fre- quency of replacing batteries, thus to realize integrated soldier sensor suites as envisioned in the warfighter concept. Consider- able efforts of researchers are also given to explore the potential of EBFCs for long-term space mission applications [5]. Current technologies, including solar and nuclear energies are either ex- cessively expensive or dangerous, which motivates scientists to Manuscript received December 31, 2009; revised March 19, 2010; accepted April 20, 2010. Date of publication May 17, 2010; date of current version January 26, 2011. This work was supported in part by Department of De- fense/Air Force Office of Scientific Research under Grant FA9550-07-1-0344, in part by the National Science Foundation (NSF) Major Research Instrumen- tation Program under Grant MRI 0821582, in part by the Wallace Coulter Foundation and 2008 FIU Faculty Research Award, in part by the NSF under Grant CHE-0716718, and in part by the Institute for Functional Nanomaterials NSF under Grant 0701525. The review of this paper was arranged by Associate Editor L. Dong. C. Liu and C.-Z. Li are with the Nanobioengineering/Bioelectronics Labora- tory, Department of Biomedical Engineering, Florida International University, Miami, FL 33174 USA (e-mail: licz@fiu.edu; cliu003@fiu.edu). Z. Chen is with the Department of Chemistry, Institute for Functional Nanomaterials, University of Puerto Rico, San Juan, PR 00923 USA (e-mail: zhongfangchen@gmail.com). Color versions of one or more of the figures in this paper are available online at http://ieeexplore.ieee.org. Digital Object Identifier 10.1109/TNANO.2010.2050147 develop an alternative portable energy source. However, due to the poor stability caused by enzyme denaturation, the EBFCs have not been successful in practical applications. Currently, how to improve the stability and to fabricate practical EBFC devices out of theoretical concepts [6]–[10] is a key challenge. Among others, covalent binding and physical entrapment are the two main strategies for stability enhancement. In this paper, we demonstrate a simple, practical EBFC system based on electropolymerized pyrrole immobilized en- zyme/graphene sheets composite electrodes. Among different enzyme immobilization methods studied so far, encapsulating enzymes in polypyrrole film offers a facile strategy for the fabri- cation of EBFCs, since the process only includes the application of a fixed potential between the working electrode and the ref- erence electrode in a solution containing the pyrrole monomer and enzymes. This technique is especially attractive for the en- zymatic functionalization of EBFC electrodes, because we can estimate the amount of the immobilized enzyme and the thick- ness of the growing polymer film easily by measuring the charge passed through the electrode. Moreover, earlier studies on the microstructure and the conductivity of polypyrrole [11] showed that polypyrrole is a porous polymer with high conductivity and surface-to-volume ratio. These properties inspired us to exam- ine the performance of polypyrrole as a diffusion and electron transfer medium and also as a protection of enzymes. Besides the physical encapsulation of polypyrrole, graphene nanosheet material has also been employed to construct co- valent linkage between the enzymes and the electrodes. As a novel carbon allotrope, graphene possesses a very large sur- face area, which is about 2630 m 2 ·g −1 [12]. The electrons on the graphene surface move ballistically over the sheet without any collisions with mobilities as high as 10 000 cm 2 ·V −1 ·s −1 at room temperature [13]–[16]. Recently, we demonstrated that graphene exhibits a larger I D /I G ratio relative to single-walled carbon nanotube, which is an indication of greater sp 2 char- acter [17]. Note that though with high purity, the chemically coverted graphene (from graphene oxide) used in our experi- ments possesses a number of surface active functional moieties, such as carboxylic and ketonic groups, which are reactive and can easily bind covalently with enzymes [18]. The presence of C=C conjugation in graphene is also expected to boost the elec- tron transfer rate, which significantly enhances the power output of the EBFCs [19]. More recently, we employed graphene and sol–gel, another common immobilization method for biofuel cell assembly. However, the power output and the stability were unsatisfied for future security and space applications [20]. In this paper, we report a novel strategy of surface func- tionalization and immobilization for EBFC applications using 1536-125X/$26.00 © 2010 IEEE