**1. Introduction**

[24] Bi, H., Huang, F. Q., Liang, J., Tang, Y. F., Lu, X. J., Xie, X. M., & Jiang, M. H. (2011). Large-scale preparation of highly conductive three dimensional graphene and its ap‐

[25] Leubner, S., Katsukis, G., & Guldi, D. M. (2012). Decorating polyelectrolyte wrapped SWNTs with CdTe quantum dots for solar energy conversion. *Faraday Discuss.*, 155,

[26] Bourdo, S. E., Saini, V., Piron, J., Al-Brahim, I., Boyer, C., Rioux, J., Bairi, V., Biris, A. S., & Viswanathan, T. (2012). Photovoltaic Device Performance of Single-Walled Car‐ bon Nanotube and Polyaniline Films on n-Si: Device Structure Analysis. *ACS applied*

[27] Mattoso, L. H. C., Manohar, S. K., Macdiarmid, A. G., & Epstein, A. J. (1995). Studies on the Chemical Syntheses and on the Characteristics of Polyaniline Derivatives.

[28] Kozawa, D., Hiraoka, K., Miyauchi, Y., Mouri, S., & Matsuda, K. (2012). Analysis of the Photovoltaic Properties of Single-Walled Carbon Nanotube/Silicon Heterojunc‐

[29] Castrucci, P., Del Gobbo, S., Camilli, L., Scarselli, M., Casciardi, S., Tombolini, F., Convertino, A., Fortunato, G., & De Crescenzi, M. (2011). Photovoltaic Response of Carbon Nanotube-Silicon Heterojunctions: Effect of Nanotube Film Thickness and

[30] Hewitt, C. A., Kaiser, A. B., Roth, S., Craps, M., Czerw, R., & Carroll, D. L. (2012). Multilayered Carbon Nanotube/Polymer Composite Based Thermoelectric Fabrics.

*Journal of Polymer Science Part a-Polymer Chemistry.*, 33(8), 1227-1234.

Number of Walls. *J Nanosci Nanotechnol.*, 11(10), 9202-9207.

plications in CdTe solar cells. *J Mater Chem.*, 21(43), 17366-17370.

253-265.

*materials & interfaces.*, 4(1), 363-368.

432 Syntheses and Applications of Carbon Nanotubes and Their Composites

tion Solar Cells. *Appl Phys Express*, 5(4).

*Nano Lett.*, 12(3), 1307-1310.

Alternative energy pathways to replace depleting oil reserves and to limit the effects of glob‐ al warming by reducing the atmospheric emissions of carbon dioxide are nowadays re‐ quired. Dihydrogen appears as an attractive candidate because it represents the highest energy output relative to the molecular weight (120 MJ kg-1 against 50 MJ kg-1 for natural gas), and because its combustion delivers only water and heat. Whereas the main renewable sources of energy available in nature (solar, wind, geothermal…) need to be transformed, di‐ hydrogen is able to transport and store energy. Dihydrogen can be produced from renewa‐ ble energies, indirectly from photosynthesis *via* biomass transformation, or directly by bacteria. It can be converted into electricity using fuel cell technology. From all these proper‐ ties and because it does not compete with food and water resources, dihydrogen has been defined as third generation biofuel. It thus emerges as a new fully friendly environmental energy vector. The use of dihydrogen as an energy carrier is not a new idea. Let us simply remember that Jules Verne, a famous French visionary novelist, wrote early in 1874: "I be‐ lieve that O2 and H2 will be in the future our energy and heat sources" [1]. His prediction simply relied on the discovery a few years before of the fuel cell concept by C. Schönbein, then W. Groove, who demonstrated that when stopping water electrolysis, a current flow occurred in the reverse way [2]. However in order to implement the dihydrogen economy and replace fossil fuels, there are significant technical challenges that need to be overcome in each of the following domains:

© 2013 De Poulpiquet et al.; licensee InTech. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2013 De Poulpiquet et al.; licensee InTech. This is a paper distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


As opposed to widespread opinions, natural dihydrogen sources exist alone on the earth's surface. Local and continuous emanations of dihydrogen can be observed in cratonic zones, ophiolitic rocks or oceanic ridges [3]. Dihydrogen is effectively produced in the upper man‐ tle of the earth through natural oxidation of iron (II)-rich minerals, like ferromagnesians, by water of the hydrosphere. The ferrous iron is oxidized in ferric iron and water is concurrent‐ ly reduced in dihydrogen, as given by following equation: 2Fe2+ (mineral) + 2 H+ (water) 2 Fe3+ (mineral) + H2. The same reaction can occur with other ions like Mn2+. Exploitation of these sources remains however difficult so far as dihydrogen does not accumulate on the earth subsurface, especially for two reasons. First because as a powerful energy source dihydro‐ gen is quickly consumed (biologically or abiotically), and second because as the lightest and most mobile gas it is not much retained by Earth's attraction and escapes in the atmosphere.

isms or enzymes could be used instead of chemical catalysts for the development of efficient electricity producing devices. These innovative batteries called biofuel cells rely on enzymes highly specific for various fuels and oxidants [11]. A mandatory condition is that these en‐ zymes have to be immobilized onto electrodes. One of the most common biofuel cell uses glucose oxidase and laccase, two enzymes specific for glucose oxidation and oxygen reduc‐ tion, respectively. A few years ago, a new concept of biofuel cells appeared based on en‐ zymes specific of dihydrogen oxidation. This biohydrogen economy relies on the opportunity to use low-cost materials for efficient conversion of solar energy to dihydrogen and of dihydrogen to electricity. Many microorganisms biosynthesize hydrogenase, the met‐ alloenzyme that catalyzes the dihydrogen conversion. At least two modes of application of dihydrogen-metabolizing protein catalysts are nowadays considered within dihydrogen as a future energy carrier. Hydrogenases may be used as catalysts in dihydrogen production by coupling oxygenic photosynthesis to biological dihydrogen production [12]. Hydrogenases can also be used directly as anode catalysts in biofuel cells instead of chemical catalysts [13]. The improved knowledge of hydrogenase structure and of catalytic mechanisms allows nowadays to design the development of biofuel cells functioning as Proton Exchange Mem‐

Carbon Nanotube-Enzyme Biohybrids in a Green Hydrogen Economy

http://dx.doi.org/10.5772/51782

435

**Figure 1.** A) Energetic metabolism of the bacterium *Aquifex aeolicus*: H2 oxidation in the periplasm is coupled to O2 reduction in the cytoplasm *via* a membrane quinone pool to generate a trans-membrane proton gradient for ATP

For all these innovative concepts, one of the key points is the increase in power density, thus in the current density furnished by a redox couple displaying a large as possible potential difference. Apart from the improvement in enzyme stability, the increase in the current densities supposes an optimization of both the interfacial electron transfer rate and the amount of connected enzymes at the electrode. Carbon nanotubes which develop large surface areas and can be functionalized constitute an attractive platform for such enzyme immobilization. CNTs are described as graphene sheets rolled into tubes. They exist under various structural config‐ urations (single-walled (SWCNTs), multi-walled (MWCNTs)) differing in electrical proper‐

brane (PEM) fuel cells.

synthesis; (B) General view of a chemical PEM fuel cell.

Combined with water and hydrocarbons dihydrogen is nevertheless the most abundant ele‐ ment on earth. Green means to ecologically convert H containers into dihydrogen still re‐ main however challenging. The energetic volume density of dihydrogen is low (10.8 MJ m-3 against 40 MJ m-3 for natural gas) so that storage and transportation appear as bottlenecks for large scale development in transportation for example. Conversion of dihydrogen to electricity in fuel cells presents high electrical efficiency (more than 50% against less than 30% for gas engines), but requires the use of catalysts both for H2 oxidation and O2 reduc‐ tion. These are mainly based on platinum catalysts, which are highly expensive, weakly available on earth, and non biodegradable. Extensive researches thus aim to decrease the amount of platinum catalysts in fuel cells. Following the discovery of carbon nanotubes (CNTs) [4, 5], their large scale availability opened a new avenue in these three domains. Due to their intrinsic properties, such as high stability, high electrical and thermal conductivities [6] and high developed surface areas, carbon nanotubes constitute attractive materials, able to enhance the credibility of an hydrogen economy.

Besides, platinum catalysts are inhibited by very low amount of CO and S (0.1% of CO is sufficient to decrease one hundred fold the catalytic activity of Pt in ten minutes!), thus re‐ quiring strong steps of H2 purification [7]. They are not specific to either O2 or H2 catalysis, thus requiring the use of a membrane to separate the anodic and cathodic compartments. Nafion® perfluoronated membrane is currently the only really performing polymer [8], in‐ creasing its cost. Replacement of platinum-based catalysts is thus highly needed. In that way, a new concept appeared less than five years ago, when looking at the pathways micro‐ organisms use for the production of ATP, their own energetic source [9, 10]. As an example, the hyperthermophilic, microaerophilic bacterium *Aquifex aeolicus*, couples H2 oxidation to O2 reduction *via* a membrane quinone pool (Figure 1). The redox coupling generates a pro‐ ton gradient through the cell membrane for ATP synthesis. Clearly, this pathway can be considered as an "*in vivo* biofuel cell". The question rises if we could take benefit of bacterial energetic pathways for our own energetic needs. The idea thus emerged that microorgan‐ isms or enzymes could be used instead of chemical catalysts for the development of efficient electricity producing devices. These innovative batteries called biofuel cells rely on enzymes highly specific for various fuels and oxidants [11]. A mandatory condition is that these en‐ zymes have to be immobilized onto electrodes. One of the most common biofuel cell uses glucose oxidase and laccase, two enzymes specific for glucose oxidation and oxygen reduc‐ tion, respectively. A few years ago, a new concept of biofuel cells appeared based on en‐ zymes specific of dihydrogen oxidation. This biohydrogen economy relies on the opportunity to use low-cost materials for efficient conversion of solar energy to dihydrogen and of dihydrogen to electricity. Many microorganisms biosynthesize hydrogenase, the met‐ alloenzyme that catalyzes the dihydrogen conversion. At least two modes of application of dihydrogen-metabolizing protein catalysts are nowadays considered within dihydrogen as a future energy carrier. Hydrogenases may be used as catalysts in dihydrogen production by coupling oxygenic photosynthesis to biological dihydrogen production [12]. Hydrogenases can also be used directly as anode catalysts in biofuel cells instead of chemical catalysts [13]. The improved knowledge of hydrogenase structure and of catalytic mechanisms allows nowadays to design the development of biofuel cells functioning as Proton Exchange Mem‐ brane (PEM) fuel cells.

**1.** dihydrogen production and generation,

434 Syntheses and Applications of Carbon Nanotubes and Their Composites

**2.** dihydrogen storage and transportation,

**3.** dihydrogen conversion to electrical energy.

to enhance the credibility of an hydrogen economy.

As opposed to widespread opinions, natural dihydrogen sources exist alone on the earth's surface. Local and continuous emanations of dihydrogen can be observed in cratonic zones, ophiolitic rocks or oceanic ridges [3]. Dihydrogen is effectively produced in the upper man‐ tle of the earth through natural oxidation of iron (II)-rich minerals, like ferromagnesians, by water of the hydrosphere. The ferrous iron is oxidized in ferric iron and water is concurrent‐

Fe3+ (mineral) + H2. The same reaction can occur with other ions like Mn2+. Exploitation of these sources remains however difficult so far as dihydrogen does not accumulate on the earth subsurface, especially for two reasons. First because as a powerful energy source dihydro‐ gen is quickly consumed (biologically or abiotically), and second because as the lightest and most mobile gas it is not much retained by Earth's attraction and escapes in the atmosphere.

Combined with water and hydrocarbons dihydrogen is nevertheless the most abundant ele‐ ment on earth. Green means to ecologically convert H containers into dihydrogen still re‐ main however challenging. The energetic volume density of dihydrogen is low (10.8 MJ m-3 against 40 MJ m-3 for natural gas) so that storage and transportation appear as bottlenecks for large scale development in transportation for example. Conversion of dihydrogen to electricity in fuel cells presents high electrical efficiency (more than 50% against less than 30% for gas engines), but requires the use of catalysts both for H2 oxidation and O2 reduc‐ tion. These are mainly based on platinum catalysts, which are highly expensive, weakly available on earth, and non biodegradable. Extensive researches thus aim to decrease the amount of platinum catalysts in fuel cells. Following the discovery of carbon nanotubes (CNTs) [4, 5], their large scale availability opened a new avenue in these three domains. Due to their intrinsic properties, such as high stability, high electrical and thermal conductivities [6] and high developed surface areas, carbon nanotubes constitute attractive materials, able

Besides, platinum catalysts are inhibited by very low amount of CO and S (0.1% of CO is sufficient to decrease one hundred fold the catalytic activity of Pt in ten minutes!), thus re‐ quiring strong steps of H2 purification [7]. They are not specific to either O2 or H2 catalysis, thus requiring the use of a membrane to separate the anodic and cathodic compartments. Nafion® perfluoronated membrane is currently the only really performing polymer [8], in‐ creasing its cost. Replacement of platinum-based catalysts is thus highly needed. In that way, a new concept appeared less than five years ago, when looking at the pathways micro‐ organisms use for the production of ATP, their own energetic source [9, 10]. As an example, the hyperthermophilic, microaerophilic bacterium *Aquifex aeolicus*, couples H2 oxidation to O2 reduction *via* a membrane quinone pool (Figure 1). The redox coupling generates a pro‐ ton gradient through the cell membrane for ATP synthesis. Clearly, this pathway can be considered as an "*in vivo* biofuel cell". The question rises if we could take benefit of bacterial energetic pathways for our own energetic needs. The idea thus emerged that microorgan‐

(water) 2

ly reduced in dihydrogen, as given by following equation: 2Fe2+ (mineral) + 2 H+

**Figure 1.** A) Energetic metabolism of the bacterium *Aquifex aeolicus*: H2 oxidation in the periplasm is coupled to O2 reduction in the cytoplasm *via* a membrane quinone pool to generate a trans-membrane proton gradient for ATP synthesis; (B) General view of a chemical PEM fuel cell.

For all these innovative concepts, one of the key points is the increase in power density, thus in the current density furnished by a redox couple displaying a large as possible potential difference. Apart from the improvement in enzyme stability, the increase in the current densities supposes an optimization of both the interfacial electron transfer rate and the amount of connected enzymes at the electrode. Carbon nanotubes which develop large surface areas and can be functionalized constitute an attractive platform for such enzyme immobilization. CNTs are described as graphene sheets rolled into tubes. They exist under various structural config‐ urations (single-walled (SWCNTs), multi-walled (MWCNTs)) differing in electrical proper‐ ties, thus tuning the platform properties for enzyme immobilization. The end of the tubes is capped by a fullerene-type hemisphere that yields selective functionalization of the CNTs [14].

**3. Carbon nanotubes for safe and efficient H2 storage**

from MH2 upon increase in temperature and/or decrease in pressure.

**H2 liquid, -253 C**

**MgH2 Mg2NiH4 FeTiH2 LaNi5H6**

Carbon Nanotube-Enzyme Biohybrids in a Green Hydrogen Economy

http://dx.doi.org/10.5772/51782

437

0.99 4.2 6.5 5.9 6.0 5.5

Much progress has been made during the last years in that domain, including the high‐ light of the advantages offered by using CNTs. An efficient approach appears to be the formulation of new carbon/transition metal catalyst composites of specific composition and molecular structure, which can greatly stimulate and improve the chemical reactions involv‐ ing dihydrogen relocation in alkali-metal aluminium materials. Absorption kinetics and dihydrogen storage capacity were shown to be enhanced by mixing MH2 with SWCNTs as a result of an increase in interfacial area, decrease in MH2 particle agglomeration and nanoplatform for efficient H2 diffusion [20, 21]. The hydriding and dehydriding kinetics of SWCNT/catalyzed sodium aluminium composite were found to be much better than those of the material ground without carbon additives. Temperature of H2 desorption was low‐ ered [22]. The presence of carbon creates new dihydrogen transition sites and the high dihydrogen diffusivity of the nanotubes facilitates hydrogen atom transition. Faster ther‐

**Material H2 gas,**

H-atom per cm3 (x1022)

**200 bar**

**Table 1.** H density as a function of storage method.

The use of H2 in fuel cells to generate electricity has been proved early in the middle of the nineteenth century. Surprisingly this discovery by C. Schönbein in 1839 of current genera‐ tion by use of H2 and O2 in sulphuric acid was applied by NASA only late in 1960. Despite intensive studies over the last two decades, fuel cells still suffer from high cost and low du‐ rability. The first difficulty responsible for this slow large scale development lies on dihy‐ drogen storage and transportation, both regarded as bottlenecks considering dihydrogen specific volumic density as a gas. For convenience the gas must be intensely pressurized to several hundred atmospheres and stored in a pressure vessel. The ways to store dihydrogen with minimum hazard are under liquid state under cryogenic temperatures (at a tempera‐ ture of -253 °C), or more efficiently in a solid state. Storage of dihydrogen in hydride form uses an alloy that can absorb and hold large amounts of dihydrogen by bonding with hy‐ drogen and forming hydrides. A dihydrogen storage alloy is capable of absorbing and re‐ leasing dihydrogen without compromising its own structure, according to the reaction: M + H2 ↔ MH2, where M represents the metal and H, hydrogen. Qualities that make these al‐ loys useful include their ability to absorb and release large amounts of dihydrogen gas many times without deteriorating, and their selectivity toward dihydrogen only. In addi‐ tion, their absorption and release rates can be controlled by adjusting temperature or pres‐ sure. The dihydrogen storage alloys in common use occurs in four different forms: AB5 (e.g., LaNi5), AB (e.g., FeTi), A2B (e.g., Mg2Ni) and AB2 (e.g., ZrV2). Metal hydrides, such as MgH2, Mg2NiH4 or LiBH4, constitute secure reserves of dihydrogen [17-19]. Dihydrogen is released

With the objective of dihydrogen as a future green energy vector, this review focuses on the last developments in the fuel -and more especially biofuel- cell field thanks to the advanta‐ geous use of carbon nanotubes. In a first part, carbon nanotubes for H2 storage enhancement are discussed. Then fuel cells in which carbon nanotubes help to decrease the amount of high cost noble metal catalysts are described. Green H2 economy is then emphasized consid‐ ering the key role of hydrogenase, the enzyme responsible for dihydrogen conversion. This requires the functional immobilization of the biocatalysts onto electrodes. The use of carbon nanotubes in this immobilization step is underlined, including the modes of carbon nano‐ tube functionalization and enzyme or microbes grafting. Then the advantages of developing biofuel cells in which chemical catalysts are replaced by enzymes or microbes are described. A short review of the sugar/O2 biofuel cells, the most widely investigated biofuel cell, is giv‐ en with a particular attention on the devices based on carbon nanotube-modified bioelectro‐ des. The last developments based on carbon nanotube networks for hydrogenase immobilization, or mimicking synthetic complex immobilization, in view of efficient dihy‐ drogen catalytic oxidation are finally described in order to allow the design of a future H2/O2 biofuel cell.

#### **2. Carbon nanotubes: an attractive carbon material**

The discovery of carbon nanotubes (CNTs) has induced breakthroughs in many scientific domains, including H2 economy, biosensors, bioelectrochemistry…This is due to their re‐ markable properties, such as good electronic, mechanical and thermal properties. Their nanometric size compares with that of proteins and enzymes, offering the possibility of elec‐ trical connection. Their large developed surface area allows the development of devices in smaller volumes. SWCNTs are sp2 hybridized carbon in a hexagonal honeycomb structure that is rolled into hollow tube morphology [15]. MWCNTs are multiple concentric tubes en‐ circling each other [5]. Depending on the chirality, CNTs can be metallic or semiconducting. The distinction between metallic and semiconducting is very important for application, but the physical separation of allotropes is one of the most difficult challenges to overcome. In MWCNTs, a single metallic layer results in the entire nanotubes metallic behavior. Most of‐ ten mixtures of these two forms are present in CNTs preparation. More information on the physical and electronic structures can be found in many published reviews [16]. CNTs are produced by various methods such as arc discharge, laser ablation, and chemical vapor dep‐ osition (CVD). Commercially CNTs are generally produced by CVD during the pyrolysis of hydrocarbon gases at high temperature. The control of synthesis parameters (reagent gas, T °, metal catalysts) allows for the control of CNT properties. Metal impurities may remain in the CNTs sample, thus requiring purification steps. CNTs may be treated to functionalize the surface.
