pre-printThylakoid membranes have been proposed for electrochemical solar energy conversion, but they have been plagued with short term instability. In this paper, thylakoid membranes extracted from Spinacia oleracea were physically adsorbed onto Toray paper electrodes with and without catalase, followed by entrapment in a vapor deposited silica matrix. The bioelectrodes were tested using voltammetry and amperometry and tested in a complete photobioelectrochemical cell. Upon subsequent polarization experiments, a significant decrease in the maximum current density from 1.53 ± 0.13 μAcm−2 to 0.75 ± 0.14 μAcm−2 was observed without catalase present. When catalase was included in the anode, this current decrease was not observed, showing the importance of catalase to scavenge reactive oxygen species produced by the thylakoids during photoelectrocatalysis

Minteer, Shelley D.

Sjoholm, Kyle H.

English

The University of Utah: J. Willard Marriott Digital Library

ECS Electrochemistry Letters, 1 (5) G7-G9 (2012) G72162-8726/2012/1(5)/G7/3/$28.00 © The Electrochemical SocietyBio-Solar Cells Incorporating Catalase for Stabilization ofThylakoid Bioelectrodes during Direct PhotoelectrocatalysisKyle H. Sjo¨holm,∗ Michelle Rasmussen,∗ and Shelley D. Minteer∗∗,zDepartment of Chemistry and Materials Science and Engineering, University of Utah, Salt Lake City, Utah 84112, USAThylakoid membranes have been proposed for electrochemical solar energy conversion, but they have been plagued with shortterm instability. In this paper, thylakoid membranes extracted from Spinacia oleracea were physically adsorbed onto Toray paperelectrodes with and without catalase, followed by entrapment in a vapor deposited silica matrix. The bioelectrodes were tested usingvoltammetry and amperometry and tested in a complete photobioelectrochemical cell. Upon subsequent polarization experiments,a significant decrease in the maximum current density from 1.53 ± 0.13 μAcm−2 to 0.75 ± 0.14 μAcm−2 was observed withoutcatalase present. When catalase was included in the anode, this current decrease was not observed, showing the importance ofcatalase to scavenge reactive oxygen species produced by the thylakoids during photoelectrocatalysis.© 2012 The Electrochemical Society. [DOI: 10.1149/2.006205eel] All rights reserved.Manuscript submitted June 7, 2012; revised manuscript received August 6, 2012. Published August 29, 2012.Thylakoid membranes contain the redox complexes responsible forthe light-dependent reactions of photosynthesis found in cyanobacte-ria and the chloroplasts of plants. They are composed of two pho-tosystems along with several other enzymes and cofactors. Light isabsorbed by photosystem II to oxidize water to dioxygen. The elec-trons produced in the reaction are shuttled to plastoquinone, thencytochrome b6f, plastocyanin, and finally photosystem I where theyare excited by absorbed photons.1 They are then used in the reductionof ferredoxin by ferredoxin reductase with the simultaneous conver-sion of NADP+ to NADPH.2 The protons generated form a protongradient which allows for the production of ATP, the unit of energycurrency in the living cell, by ATP synthase.1 As the quantum yieldfor photosynthesis in certain components of thylakoids, specificallyphotosystem I, is almost 100%,3 developing a method for using thesemembranes for photoelectrocatalytic energy conversion in a bio-solarcell is highly desirable.Photovoltaic devices or solar cells are defined as a device capableof converting light (photons) into electrical energy. They are gener-ally constructed of semiconductors and come in a number of varieties,ranging from organic thin film (19.6 ± 0.6% efficiency) and dye sensi-tized (11.0 ± 0.3%) solar cells (DSSC) to more efficient silicon based(25.0 ± 0.5%) and III-V semiconductor (28.3 ± 0.8%) solar cells.4Biological and bio-inspired photoelectrocatalysts have been proposedfor solar energy conversion.5 The bio-inspired photoelectrocatalystsare mostly focused on organometallic complexes with structure in-spired by photosystem I or photosystem II,5a,6 whereas the biologicalphotoelectrocatalysts have focused on either the use of photosystemI5b,7 or II8 or the intact thylakoid membranes5c,9 or chloroplasts.10 Themajority of the literature on photosystem I or II photobioelectrocatal-ysis and thylakoid photobioelectrocatalysis has employed the use ofmediators.5c,5d,9b However, these mediators typically result in a volt-age loss as well as issues associated with their own light, temperature,and long term stability. Therefore, there has been research on directphotobioelectrocatalysis,5b,7a,8 but primarily focused on photosystemsand not thylakoid immobilization.In this work, a bio-solar cell was developed which incorporateda thylakoid anode to generate photobioelectrocatalytic current.The thylakoid membranes were entrapped in a silica matrix usingvapor deposited tetramethyl orthosilicate (TMOS) developed byAtanassov et al.11 The anodes were connected with an air-breathingcathode (although other cathodes could be easily employed) andtested electrochemically using a combination of amperometry, cyclicvoltammetry, and current-voltage curves. Catalase, an enzyme that iscommonly employed to reduce the damaging effects due to reactiveoxygen species production by biocatalysts,12 was incorporated in thebioanodes. It is hypothesized that addition of this enzyme will lead∗Electrochemical Society Student Member.∗∗Electrochemical Society Active Member.zE-mail: minteer@chem.utah.eduto greater stability for the bio-solar cell by consuming any reactiveoxygen species that may be formed by the thylakoid membranes.Results and DiscussionExperimental procedures for all of the reported experiments aredetailed in the Supplementary Material.13 Thylakoids purified fromspinach were immobilized onto Toray paper electrodes with vapordeposited TMOS, as shown in Figure 1. SEM micrographs of immo-bilized thylakoids with and without the TMOS layer were obtainedand are shown in Figure S1 in the Supplementary Material.13 Withno TMOS layer, the thylakoids look similar to previously reportedthylakoid layers in the literature.14 However, after the TMOS layeris added, a more 3-dimensionally structured surface is observed, asshown in Figure S1b. Elemental analysis confirmed an increase inSi after TMOS deposition to 3.47 atomic%, which corresponds to6.78 wt%, and confirms the silicate layer formation.Activity assays of immobilized thylakoids with no catalase showedan increase in oxygen production of 219 ± 22 μmol/min when ex-posed to light compared to dark. This activity was not stable anddecreased to the background value after less than 10 minutes. Whencatalase was included with the immobilized thylakoids, the activity inlight showed the same increase in oxygen production and remainedconstant for approximately 30 minutes and then gradually decreasedto 50% activity after 2 hours. These results indicate that the additionof catalase does increase the stability of the thylakoids. Additionally,the amount of peroxide generated by the immobilized thylakoids wasdetermined using a peroxidase assay. Thylakoids with no catalaseproduced 1.2 ± 0.4 mmol/min while thylakoids immobilized withcatalase showed no detectable peroxide generation, indicating that theperoxide is being consumed by the catalase.Representative cyclic voltammograms of the modified electrodeswere obtained in the presence and absence of light, as shown inFigure S2 in the Supplementary Material.13 No significant differenceswere observed in these experiments. Amperometric measurementswere conducted by measuring the current at 0.3 V vs. SCE, as shownin Figure 2. Because the process being observed is an oxidation, it isFigure 1. Schematic of bio-solar cell.  ecsdl.org/site/terms_use address. Redistribution subject to ECS license or copyright; see 155.97.11.183Downloaded on 2013-05-21 to IP G8 ECS Electrochemistry Letters, 1 (5) G7-G9 (2012)0 50 100 150 200 2500.00-0.05-0.10-0.15-0.20-0.25-0.30-0.35-0.40Current density, µAcm-2Time, sFigure 2. Current measured with a thylakoid-modified electrode (thick line)at 0.3 V vs. SCE in 0.1 M phosphate buffer with 0.1 M nitrate. Gray arrowsindicate when the light was turned on and black arrows indicate when the lightwas turned off. Data from a control experiment with an electrode containingTMOS and catalase only is also included (thin line).expected that the current will increase in the negative direction whenthe light is turned on. This result is observed and the current returnedto background levels when the light was turned off. The increase wasapproximately 150 nA/cm2. Control experiments performed with elec-trodes modified with catalase and TMOS, but no thylakoids showedno current change in the presence of light, as shown as the controlplot in Figure 2.The thylakoid-modified electrodes were connected to an air-breathing platinum cathode separated by a Nafion membrane and theresulting bio-solar cells were tested. The average open circuit voltage(OCV) was 0.46 ± 0.03 V. Current-voltage plots (I-V plots) were ob-tained with and without light, as shown in Figure 3. When no catalasewas included in the thylakoid layer, the currents decreased with eachsubsequent test, showing photoelectrocatalysis but instability. The av-erage maximum current for the first test in the presence of the lampwas 1.53 ± 0.13 μA/cm2, but decreased to 0.90 ± 0.14 μA/cm2 forthe second test in the presence of the lamp. The performance of thebiosolar cell continued to decrease with each subsequent lamp test.0.0 0.1 0.2 0.3 0.40.00.51.01.52.02.5 First test with catalase Second test with catalase First test w/o catalase Second test w/o catalaseCurrent density, µAcm-2Voltage, VFigure 3. Current-voltage plots for a thylakoid bio-solar cell. Each cell wastested twice while exposed to light. Anodes containing no catalase and anodeswith catalase are compared.0.0 0.1 0.2 0.3 0.4 0.50.00.51.01.52.02.5 Bio-solar cell in light Bio-solar cell in dark Catalase with TMOS Toray paper TMOSCurrent density, µAcm-2Voltage, VFigure 4. Control experiments: bare Toray paper, TMOS only, catalase cov-ered with TMOS, and a complete thylakoid solar cell in the dark and light.Literature has previously reported that reactive oxygen species areproduced by photosystem I during cellular function, but these reac-tive oxygen species are consumed by ascorbate peroxidase in intactchloroplasts.15 However, the purified thylakoids do not contain thisperoxidase, which likely results in a buildup of reactive oxygen speciesafter thylakoid isolation, which is detrimental to the functioning of en-zymes in the thylakoid system. Additionally, it has been shown thatheat can cause photosystem II to produce reactive oxygen speciesas well.16 In the system being used for testing the bio-solar cell, thelamp generates significant heat with extended use and results in anequilibrium temperature of 38◦C in the light box when operated ina room temperature environment (25◦C). We hypothesized that theaddition of catalase to consume the reactive oxygen species shouldminimize the issues associated with reactive oxygen species degrada-tion in thylakoid photoelectrocatalysis. When catalase was added tothe thylakoids before deposition on the Toray carbon electrode, an in-crease in current is observed for the bio-solar cell instead of a decreasein current. The average maximum current was 2.14 ± 0.11 μA/cm2for the first test in the presence of light and 2.46 ± 0.10 μA/cm2 forthe second test in the presence of light. Both tests were done at thesame environmental temperature of 38◦C. These results show that thecatalase helps to minimize the effective of reactive oxygen speciesbefore and during photon exposure.In order to verify direct photobioelectrocatalysis, a number ofcontrols were performed to confirm that the current observed is due tothe reactions from the thylakoids. As seen in Figure 4 and Table I, theTable I. Average maximum current values and open circuitvoltages for thylakoid bioanodes utilized in bio-solar cells withand without catalase along with results from control experiments.The results of three different electrochemical cells were averagedfor each.Maximum Current Open CircuitDensity (μAcm−2) Potential (V)Thylakoids w/ catalase bioanodesFirst light treatment 2.14 ± 0.11 0.46 ± 0.03Second light treatment 2.46 ± 0.10 0.45 ± 0.02Thylakoids w/o catalasebioanodesFirst light treatment 1.53 ± 0.13 0.47 ± 0.05Second light treatment 0.90 ± 0.14 0.42 ± 0.01Bare Toray paper 0.29 ± 0.07 0.44 ± 0.02TMOS coated Toray paper 0.41 ± 0.01 0.31 ± 0.02Catalase and TMOS coated Toraypaper0.58 ± 0.05 0.39 ± 0.04  ecsdl.org/site/terms_use address. Redistribution subject to ECS license or copyright; see 155.97.11.183Downloaded on 2013-05-21 to IP ECS Electrochemistry Letters, 1 (5) G7-G9 (2012) G9individual components of the modified electrodes (i.e. Toray carbonpaper alone, Toray carbon paper with a TMOS coating layer, Toraycarbon paper with catalase in a TMOS coating layer) show little tono photocurrent. Additionally, a solar cell containing a thylakoid-modified anode showed significantly smaller currents in the dark thanwhen exposed to light, showing solar photobioelectrocatalysis. Thesebackground experiments confirmed direct photobioelectrocatalysisand the ability of the thylakoids to transfer electrons to the electrodewithout the presence of an external mediator.ConclusionsThylakoids were successfully entrapped in a vapor deposited silicamatrix on a Toray carbon paper electrode to make an anode for a bio-solar cell capable of direct photobioelectrocatalysis. When combinedwith an air-breathing cathode, the cell was able to produce a maximumcurrent density of 2.46 ± 0.10 μA/cm2 with an open circuit voltageof 0.46 ± 0.3 V. The current was not stable due to the formation ofreactive oxygen species. The addition of the enzyme catalase to theanode corrected this problem allowing for a stable current density andextending the feasible lifetime of this bio-solar cell.Supporting InformationExperimental procedures and results of additional experiments, in-cluding scanning electron micrographs of bioanodes with and withoutthe addition of the TMOS immobilization layer and representativecyclic voltammograms of the immobilized thylakoids bioanodes.AcknowledgmentsThis work was supported by a National Science Foundation MR-SEC grant for funding. The authors would also like to thank RandyPolson for obtaining SEM and elemental analysis data.References1. C. F. Meunier, J. C. Rooke, A. Leonard, H. Xie, and B.-L. 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Bio-solar cells incorporating catalase for stabilization of thylakoid bioelectrodes during direct photoelectrocatalysis

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Bio-solar cells incorporating catalase for stabilization of thylakoid bioelectrodes during direct photoelectrocatalysis

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