Photoactive Films of Photosystem I on Transparent Reduced

Jul 10, 2014 - ABSTRACT: Photosystem I (PSI) is a photoactive electron-transport protein found in plants that participates in the process of photosynt...
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Photoactive Films of Photosystem I on Transparent Reduced Graphene Oxide Electrodes Emily Darby,† Gabriel LeBlanc,† Evan A. Gizzie,† Kevin M. Winter,† G. Kane Jennings,‡ and David E. Cliffel*,† †

Department of Chemistry and ‡Department of Chemical and Biomolecular Engineering, Vanderbilt University, Nashville, Tennessee 37235, United States S Supporting Information *

ABSTRACT: Photosystem I (PSI) is a photoactive electron-transport protein found in plants that participates in the process of photosynthesis. Because of PSI’s abundance in nature and its efficiency with charge transfer and separation, there is a great interest in applying the protein in photoactive electrodes. Here, we developed a completely organic, transparent, conductive electrode using reduced graphene oxide (RGO) on which a multilayer of PSI could be deposited. The resulting photoactive electrode demonstrated current densities comparable to that of a gold electrode modified with a multilayer film of PSI and significantly higher than that of a graphene electrode modified with a monolayer film of PSI. The relatively large photocurrents produced by integrating PSI with RGO and using an opaque, organic mediator can be applied to the facile production of more economic solar energy conversion devices.

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INTRODUCTION Photosynthesis is a highly efficient photoconversion process occurring in plants, algae, and cyanobacteria. This process is carried out by protein complexes, one of which is Photosystem I (PSI), an ∼500 kDa photoactive electron-transport protein complex found in the thylakoid membrane. PSI operates as a photodiode and naturally performs the charge separation and transfer that researchers have been trying to recreate artificially for years with photoconversion devices. Charge transfer is carried out when energy from light excites an electron at the P700 site, which then proceeds down in energy through an electron-transport chain to the terminal iron−sulfur complex, known as FB.1 Due to PSI’s abundance in nature and its nearly perfect quantum efficiency, there has been interest in applying the protein in photoactive devices.2−4 The use of modified gold electrodes,5−7 polymers,8,9 and semiconductor10,11 electrode materials has shown significant improvements in current densities of PSI-based devices. Several recent developments have also demonstrated improved photocurrents by integrating PSI with various carbon-based electrodes. Carmeli et al. described how PSI could be covalently bound to carbon nanotubes.12,13 Previously, our research team integrated PSI with monolayer graphene.14 The unique properties of graphene have generated great interest in its use for electrochemical applications. However, the time-consuming process of producing monolayer graphene currently makes its use on a large scale unrealistic. Reduced graphene oxide (RGO) is an attractive alternative to graphene because it is transparent and conductive but can be produced via rapid chemical processes.15 Graphite is easily oxidized to produce graphene oxide (GO), a monolayer of carbon rings with oxygen functional groups. Because of GO’s dipole introduced by the oxygen functional groups, it is easily dissolved in polar solvents © 2014 American Chemical Society

such as water and ethanol. While GO lacks the electrical conductivity of graphite, the thermal or chemical reduction of GO produces RGO, which regains much of the lost conductivity.16 In addition, the relative transparency of RGO gives us the ability to illuminate through the photoactive electrode directly, rather than through the electrochemical mediator. Ciesielski et al. demonstrated that increasing the thickness of PSI deposited on the substrate enhances photocurrent densities.17 By depositing a multilayer of PSI instead of a monolayer, the photocurrent density can be increased by an order of magnitude. Interestingly, the photoexcited electrons generated during the illumination of these thick multilayer films are able to migrate to the underlying electrode or to an electrochemical mediator. This ability to transfer electrons is likely due to the fact that the PSI film can serve as a conductive medium under illumination conditions.18,19 Unlike graphene, large-area RGO is not found in true monolayer thickness due to the incomplete removal of oxygen functional groups during reduction. For comparison, the RGO films used in this study are approximately 50 nm thick while a film of graphene is less than 1 nm thick. This increased thickness does absorb significantly more light but also makes the electrode more robust. Along with the increased thickness, the RGO films possess a significant amount of roughness not possible in a monolayer graphene electrode (Supporting Information), which can improve the contact surface area between the PSI multilayer film and the conductive RGO electrode. In the PSIRGO system, the photoexcited electrons can be shuttled away Received: March 19, 2014 Revised: July 9, 2014 Published: July 10, 2014 8990

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Electrochemical Measurements. A CH Instruments CHI 660A electrochemical workstation equipped with a Faraday cage was used to perform electrochemical measurements. A custom-built threeelectrode cell was used to analyze the biohybrid electrodes. The RGO substrate was used as the working electrode, Ag/AgCl was used as the reference electrode, and a platinum mesh was used as the counter electrode. The electrochemical mediator solution consisted of 100 mM KCl (Sigma) and either 2 mM dichloroindophenol (DCPIP), 1 mM ferricyanide/1 mM ferrocyanide, 2 mM methylene blue, 2 mM methyl viologen, 2 mM ruthenium(II) hexamine, 1 mM ruthenium(II) hexamine/1 mM ruthenium(III) hexamine, or 2 mM sodium ascorbate. Photochronoamperometric experiments were performed at the experimentally determined dark open circuit potential for each system, which ensures that each electrode is analyzed using an overpotential value of 0. This set potential prevents the generation of dark current during the photochronoamperometric experiments and allows the photocurrent values using different electrochemical mediators to be compared accurately. The photocurrent values reported in this manuscript were determined by taking the difference between the current after 10 s of illumination and the dark current baseline. The average open circuit potential for each system is reported in the Supporting Information. Illumination was generated using a 250 W cold light source (Leica KL 2500 LCD). Using a Coherent Radiation 210 power meter, we determined that the lamp generated 342 mW/cm2. However, after passing through the sample holder and the RGO electrode, much of the illumination was reflected, dispersed, or absorbed such that the light intensity reaching the PSI multilayer film or electrochemical mediator was only 34 mW/cm2. Following PSI deposition, the light intensity passing through the electrode was 10 mW/cm2.

by the conductive RGO electrode, while an electrochemical mediator is used to resupply the system with electrons (Figure 1). Here we describe the effects of depositing a multilayer film of PSI on a semitransparent, conductive RGO electrode.

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Figure 1. Cartoon depiction of the electron-transfer process for the biohybrid electrode system. Light is able to reach the PSI multilayer film (green) by passing through the RGO electrode. An electrochemical mediator (M) is used to provide electrons to the PSI film where they are excited and passed on to the RGO electrode.

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RESULTS AND DISCUSSION Previously, our research group observed photocurrent improvements by modifying graphene with PSI.14 RGO has similar properties to graphene but generally has imperfections because of unreduced oxygen functional groups remaining on the substrate following the vapor reduction process. We hypothesized that RGO would demonstrate even greater photocurrent improvement because of the ability to use thicker multilayer films of PSI. We prepared the RGO electrode by spin coating GO onto functionalized glass and then reducing the GO with hydrazine vapor. The resulting RGO film was characterized by its thickness, resistance, absorbance, Raman peak shift, and CV peak splitting (Supporting Information). To measure the photoelectrochemical performance of the PSI-RGO and RGO electrodes we preformed photochronoamperometric measurements (Figure 2). We determined the current density in seven different mediator solutions with varying formal potentials (Figure 3). The mediators tested were methyl viologen, methylene blue, ruthenium(II) hexamine, ruthenium(II)/(III) hexamine, DCPIP, ferri/ferrocyanide, and sodium ascorbate. The negative photocurrent observed for all of the mediators tested indicates that the PSI-modified RGO electrode oxidizes the mediator solution. In other words, electrons are flowing into the RGO electrode as depicted in Figure 1 for all of the mediator solutions. While all of the mediators indicate that PSI may improve the photocurrent performance of the system, only DCPIP demonstrated a statistically significant improvement (p < 0.05). We attribute the high current density observed in the control experiments to the photoactivity of the electrochemical mediators (Supporting Information). DCPIP is an attractive mediator because it is organic, inexpensive, and nontoxic. While previous studies using gold electrodes found DCPIP to be an inferior mediator due to its

EXPERIMENTAL SECTION

Extraction and Isolation of Photosystem I. Photosystem I was extracted from spinach leaves, as described previously.11 Briefly, commercially available baby spinach leaves are deveined and blended in sorbitol buffer at 8000g to isolate the thylakoid membranes.20 The resulting precipitate was then resuspended using a buffer containing 1% w/w Triton X-100 in order to break open the thylakoid membranes. The suspension was then centrifuged at 20 000g followed by column chromatography on a chilled hydroxylapatite column. Excess surfactants and salts were removed using dialysis at a 1:2000 extract to water ratio.17 The resulting PSI solution had a chlorophyll concentration of 0.32 mg/mL and a P700 concentration of 4.1 × 10−6 M as characterized by the methods of Porra21 and Baba et al.,22 respectively. Preparation of a Reduced Graphene Oxide Electrode. Reduced graphene oxide-coated slides were prepared by following previously described23 methods with a few modifications. (See the Supporting Information for more details.) Briefly, GO was deposited onto functionalized glass slides through a spin-coating process. The GO film was subsequently reduced using hydrazine vapor. The resulting film was characterized using a number of analytical techniques described in the Supporting Information. Deposition of Photosystem I on the Electrode. Photosystem I was deposited on the RGO using a drop-casting method.17 PSI (50 μL) was deposited on a 0.283 cm2 circle of RGO. The solvent in the PSI solution was then removed by applying a vacuum. This process was repeated twice more to achieve PSI films that were roughly 350 nm thick as measured by profilometry. Because the high surfactant concentrations were removed during the dialysis procedure of our PSI solution, the resulting multilayer film of PSI is resistant to removal in aqueous solutions and enables the assembly of the biohybrid electrode into an aqueous electrochemical device. 8991

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Figure 4. Photochronoamperometric analysis of PSI-RGO (green) and RGO (gray) in DCPIP when the direction of the light is varied. The mediator solution was 2 mM DCPIP with 100 mM KCl. The inset shows the UV−vis spectra of DCPIP (blue) and PSI (green).

Figure 2. Photochronoamperometry analysis of PSI-modified (green) and unmodified (gray) RGO electrodes in a 2 mM DCPIP mediator solution, with 100 mM KCl as the supporting electrolyte, Ag/AgCl as the reference electrode, and platinum mesh as the counter electrode. The light was turned on for 20 s intervals as indicated by the yellow shaded regions in the graph.

The PSI-modified RGO electrode demonstrates photocurrents comparable to those of a PSI-modified gold electrode (1.2 to 7.9 μA/cm2)17 and significantly higher than that of a PSI-modified graphene electrode (0.5 μA/cm2).14 The RGO electrode achieves comparable photocurrents and is a more attractive electrode because RGO is significantly less expensive than gold and more processable than graphene. The significant increase in photocurrent observed from RGO versus the graphene substrate is likely due to the ability to use a multilayer film of PSI instead of a monolayer film, which results in an increase in useful light absorption. These photocurrent values are still significantly lower than our previous reports using a semiconducting substrate.11,19 This difference occurs because the RGO acts in a similar fashion to a metal electrode and is therefore capable of both accepting and donating electrons from/to the PSI film. Thus, the mixed orientation of the PSI complexes within the multilayer film causes much of the photocurrent to be canceled out on metallic substrates but not semiconducting substrates. We anticipate dramatic improvements in the photocurrent production of these films on RGO if the PSI complexes can be oriented within the film. Another method for improving the photocurrent density of the system is to increase the concentration of the mediator solution.11,14 A higher mediator concentration increases the photocurrent density by minimizing the diffusional limitations of the reduced mediator of the PSI film, and thus the mediator is able to donate electrons to the PSI-RGO more rapidly. In a systematic study presented in Figure 5, a dramatic increase in photocurrent was observed from 0.2 to 2 mM. Further increasing the concentration showed a marginal improvement in the photocurrent, indicating that the concentration of the mediator solution is not a significant limiting factor in photoactivity beyond a concentration of 2 mM. Additionally, concentrations greater than 20 mM give a mediator that is pastelike rather than a pourable liquid. Thus, the optimal current density is achieved with a mediator concentration of 20 mM.

Figure 3. Average photocurrent (n = 4) of PSI-modified RGO compared to unmodified RGO electrodes in various mediator solutions. The * symbol indicates p < 0.05.

absorbance in the red region of the spectrum, 24 the transparency of the RGO electrodes provides the opportunity to use this opaque mediator as it is no longer necessary to illuminate the electrode through the mediator (Figure 4). DCPIP absorbs light in the same 600−700 nm region as PSI (inset of Figure 4). Because PSI and DCPIP absorb light in the same region, when light shines through the mediator solution, the PSI-modified electrode demonstrates a significant loss in photoactivity, although there is still some photoactivity as some of the light does pass through the mediator solution. Therefore, the transparency of the RGO electrode is necessary for the use of DCPIP as a mediator solution.

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OUTLOOK We successfully developed an organic, biohybrid photoelectrode by depositing a multilayer of PSI on a carbon-based electrode. As the carbon-based electrode, we used RGO 8992

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ACKNOWLEDGMENTS

We gratefully acknowledge the financial support of the National Science Foundation (DMR 0907619), the NSF EPSCoR (EPS 1004083), the United States Department of Agriculture (201367021-21029 USDA), the U.S. Environmental Protection Agency (SU8360221), and the Scialog Program from the Research Corporation for Science Advancement. E.D. was supported by a summer research experience for undergraduates (REU) funded by the National Science Foundation (DMR 1005023). G.L. was supported in part by an ACS DAC summer fellowship.

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ABBREVIATIONS PSI, Photosystem I; GO, graphene oxide; RGO, reduced graphene oxide; DCPIP, dichloroindophenol; CV, cyclic voltammetry

Figure 5. Photocurrent density as a function of DCPIP concentration for PSI-RGO and RGO electrodes (n = 4).

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because of its transparency, conductivity, and facile production via chemical processes. The resulting photoelectrode demonstrates significant photocurrent improvement compared to an unmodified RGO electrode when DCPIP is used as the electrochemical mediator. The transparency of the RGO electrode allows us the ability to use DCPIP, an opaque mediator that is organic, inexpensive, and nonhazardous. Using DCPIP, our PSI-modified RGO electrode demonstrates comparable photocurrent densities to a PSI-modified gold electrode and nearly 10 times the photocurrent densities of a PSI-graphene electrode. Less expensive than gold and easier to produce than graphene, RGO proves to be an attractive choice for an electrode. The integration of PSI with RGO and the use of an opaque organic mediator allow the facile production of more economical organic solar cells. Furthermore, the development of a carbon-based or polymer-based counter electrode will allow for this metal-free biohybrid electrode to be integrated into a completely metal-free photovoltaic device.

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ASSOCIATED CONTENT

* Supporting Information S

Additional data describing the preparation and characterization of the RGO electrodes, the photoactivity of various electrochemical mediators, the open circuit potential values for various systems, and photovoltage measurements. This material is available free of charge via the Internet at http://pubs.acs.org.

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REFERENCES

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AUTHOR INFORMATION

Corresponding Author

*Fax: +1 615 343 1234. Tel: +1 615 343 3937. E-mail: d. cliff[email protected]. Present Address

(E.D.) Pomona College, 333 N. College Way, Claremont, California 91711, United States. Author Contributions

The manuscript was written through the contributions of all authors. All authors have given approval to the final version of the manuscript. Notes

The authors declare no competing financial interest. 8993

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