Bioresource Technology 101 (2010) 4013–4020

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Bioresource Technology

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Electricity generation by two types of microbial fuel cells using nitrobenzeneas the anodic or cathodic reactants

Jie Li, Guangli Liu *, Renduo Zhang, Yong Luo, Cuiping Zhang, Mingchen LiSchool of Environmental Science and Engineering, Sun Yat-Sen University, Guangzhou, Guangdong 510275, China

a r t i c l e i n f o a b s t r a c t

Article history:Received 15 September 2009Received in revised form 20 December 2009Accepted 29 December 2009

Keywords:Microbial fuel cellNitrobenzeneDegradationElectricity generation

0960-8524/$ - see front matter � 2010 Elsevier Ltd. Adoi:10.1016/j.biortech.2009.12.135

* Corresponding author. Tel.: +86 20 84110052; faxE-mail address: liugl@mail.sysu.edu.cn (G. Liu).

The effect of nitrobenzene (NB) on electricity generation and simultaneous biodegradation of NB werestudied with two types of microbial fuel cells (MFCs): a ferricyanide-cathode MFC with NB as the anodicreactant and a NB-cathode MFC. Compared to controls without NB, the presence of NB in the anode of thefirst MFC decreased maximum voltage outputs, maximum power densities and Coulombic efficiencies.No electricity was generated from the first MFC using NB as the sole fuel; however, the second MFC usingNB as the electron acceptor generated electricity successfully with a maximum voltage of 400 mV. NBwas degraded completely within 24 h in both anode and cathode chambers. Denaturing gradient gel elec-trophoresis (DGGE) profiles demonstrated that the presence of NB caused changes in relative abundanceof the dominant bacterial species and emergence of new bacteria on the anodes.

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1. Introduction

The use of microbial fuel cells (MFCs) is a promising method forwastewater treatment and power generation. Various organicmaterials have been used as fuels in MFCs, including easily degrad-able glucose (Rabaey et al., 2003), acetate (Liu et al., 2005), mono-saccharides (Catal et al., 2008), complex carbohydrates (Min et al.,2005; Shimoyama et al., 2008), and some biorefractory organics,such as cellulose (Ren et al., 2007) and petroleum contaminants(Morris and Jin, 2008). Even a few toxic and biorefractory organicscan be used as the fuel and degraded in the MFC. For example, Luoet al. (2009) operated an MFC using phenol as the fuel and demon-strated that phenol was degraded effectively.

With different substrates as the fuel, the MFC performs differ-ently. Catal et al. (2008) showed that some of the hydrolysates oflignocellulosic biomass cannot be utilized directly for electricityproduction in the MFC in the absence of other electron donors,and even inhibit electricity generation. Some non-electrode elec-tron acceptors in the anode solution, such as nitrate, can resultin electron consumption and affect performance (Liu and Logan,2004). Sukkasem et al. (2008) found that nitrate in single cham-ber air cathode MFCs did affect the maximum voltage outputsat low external resistances, but not at higher external resistances.Coulombic efficiency was greatly reduced by the addition ofnitrate.

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Nitroaromatic compounds are often found in wastewaters andcan be degraded by microorganisms from active sludge (Majumderand Gupta, 2003) and contaminated water or soils (Spain, 1995; Yeet al., 1995). The biodegradation pathways include one anaerobicreductive pathway and two aerobic oxidative pathways (He andSpain, 1999; Liu et al., 2002). Attributable to the strong electronaffinity of the nitro group, the oxidation of NB is difficult toachieve, and normally the reductive reaction occurs first underthe natural conditions (Gurevich et al., 1993). Therefore, reductivemethods including chemical reduction (Nefso et al., 2005), cata-lytic hydrogenation reduction, electrochemical reduction (Liet al., 2007), and anaerobic reduction (Kuscu and Sponza, 2009),have attracted more and more attention. However, the reductionpotential of NB (<�0.700 V) (Oliveira, 2003) is lower than that ofmolecular hydrogen at most electrodes and thus results in de-creased current efficiency. An external power supply is necessaryto carry out electrochemical reduction (Li et al., 2007). Based onits characteristics, it may be possible to use NB as the cathode sub-strate of the MFC without an external power supply. Such a MFCshould achieve simultaneous electricity generation and NBreduction.

In this study, the effect of NB on electricity generation wasinvestigated in two types of MFCs: a ferricyanide-cathode MFCwith NB as the anodic reactant and a NB-cathode MFC. We alsoexamined NB degradation rates in the anode chamber of the ferri-cyanide-cathode MFC using various glucose + NB mixtures as thefuel, and NB reduction rates in the cathode chamber of the NB-cathode MFC. Microbial communities on the anode of the ferricya-nide-cathode MFC with different substrates were also analyzed.

4014 J. Li et al. / Bioresource Technology 101 (2010) 4013–4020

2. Methods

2.1. MFC configuration

A dual-chambered MFC was constructed with an anode cham-ber and a cathode chamber, which were separated by a proton ex-change membrane (PEM, Nafion 212, Dupont Co., USA). To collectsolution samples easily from the anode chamber during the exper-iments, an anode receptacle was set up, which was a brown bottle(250 mL capacity) connected with the anode chamber. Similarly, acathode receptacle was set up to balance the flow rates in the twochambers. The anode receptacle was filled with solutions of200 mL throughout the experiments. Fifty mmol/L ferricyanidewas used as terminal electron acceptor in the cathode. A peristalticpump was used to cycle the flow at a constant flow rate of 20 mL/min in the receptacles. The electrodes were made of carbon fiberbrush of the same size (7.0 � 3.5 cm). The effective liquid volumesof the anode and cathode chambers were 27.0 mL. A copper wirewas used to connect the circuit with an external resistance of1000 X, and all exposed metal surfaces were sealed with noncon-ductive epoxy resin. The MFC was operated at a constant temper-ature (30 ± 1 �C).

To examine electricity generation of the MFC with NB as the fi-nal electron acceptor in the cathode chamber, another dual-cham-bered MFC was constructed. The cathode electrode was made ofcarbon cloth (TGP-H-060, Toray, Japan) with an area about24.5 cm2. The flexible carbon-cloth electrode contained around0.5 mg/cm2 Pt catalyst. Other operation conditions of the NB-cath-ode MFC were the same as those of the ferricyanide-cathode MFC.

2.2. MFC inoculation and operation

For quick start-up of the MFCs, the MFCs were inoculated with200 mL of mixed aerobic activated sludge and anaerobic sludge(1:1, v/v) with diverse electrochemically active bacteria. Thesludge inocula were collected from the Liede Municipal Wastewa-ter Treatment Plant of Guangzhou City, China.

Substrates used in the ferricyanide-cathode MFC included glu-cose, glucose + NB mixtures, and NB. At first, the MFC was operatedwith 1000 mg/L glucose as the fuel. After obtaining steady electri-cal outputs, the MFC was operated sequentially using 1000 mg/Lglucose + 50 mg/L NB, 1000 mg/L glucose + 150 mg/L NB,1000 mg/L glucose + 250 mg/L NB, 250 mg/L NB, and 1000 mg/Lglucose as the fuel. For each type of the substrates, the MFC ranfor 3–4 cycles. In the NB-cathode MFC, 1000 mg/L glucose wasused as the sole anode fuel and 250 mg/L NB as the electron accep-tor in the cathode.

Besides the substrates, the anodic solution also contained (perliter of deionized water): 4.0896 g Na2HPO4, 2.544 g NaH2PO4,0.310 g NH4Cl, 0.130 g KCl, trace metal solution 12.5 mL, vitaminsolution 12.5 mL, and the initial pH of all solutions was adjustedto 7.0 (Lovley and Phillips, 1988). To the NB-cathode MFC, the ini-tial pH of the cathode NB solution was adjusted to 3.0 with H2SO4.In all operations, once the voltage outputs obtained were below100 mV, the substrate was replaced with a new solution for thenext cycle. Before starting the experiment, the anode compart-ments of the two MFCs were flushed with N2 for 10 min to ensurean anaerobic environment, and the cathode compartment of theNB-cathode MFC was flushed with N2 for 10 min to eliminate theelectrons competition between O2 and NB.

2.3. Electrical and chemical analyses

Cell voltages across the external resistance were measuredusing a multimeter and data were automatically recorded by a data

acquisition system (DT85). The volumetric power density (PV, W/m3) is calculated as follows:

PV ¼U2=R

Vð1Þ

where U is the voltage (V), R is the resistance (X), and V is the effec-tive volume (i.e., the volume of the liquid media) of the anodechamber (m3). The volumetric power density indicates how muchpower is generated from unit volume of wastewater.

The Coulombic efficiency (CE) is defined as the percent of elec-trons recovered as current vs. those in the starting organic sub-strate. For complex substrates, it is more convenient to use theremoval of COD as a measure of the substrate concentration, whichis used in the CE calculation (Logan, 2008). The CE reflects the effi-ciency of chemical energy conversion into electricity and is calcu-lated by:

CE ¼ 100%MPn

i¼1Uiti

RFbDSVð2Þ

Here Ui is the output voltage (V) of MFC at time ti (s), R is theexternal resistance (X), F is Faraday’s constant (96,485 C/mol), bis the number of moles of electrons produced per mol of COD(4 e�mol/mol), DS is the removal of COD concentration (g/L), V isthe liquid volume (L), and M is the molecular weight of oxygen(32 g/mol) (Liu et al., 2005).

The polarization curve was measured by changing the externalresistances from 10,000 to 20 X. For each resistance, the MFC ranat least twice to ensure to achieve repeatable power outputs, andthen voltages were recorded (typically 5–10 min per resistance)(Logan et al., 2006). The power density was calculated for eachresistance as a function of the current using the maximum voltageoutput. The internal resistance of the cell (Rint) was calculated fromthe slope of plots of U vs. I as follows:

U ¼ Ecell � IRint ð3Þ

where Ecell is the electromotive force of the cell (Logan et al., 2006).Samples (2.0 mL each) were taken at 0, 0.5, 1.0, 2.0, 3.0, 6.0,

12.0, 24.0, 48.0 h, filtered through a syringe with 0.22 lm filterunit to remove cells, and diluted 5 or 10 times using deionizedwater. The samples were used for the following chemical analyses.The COD was measured with the dichromate method (Clesceriet al., 1998). Glucose content was measured using the anthronemethod (Raunkjer et al., 1994). Concentration of NB was measuredusing HPLC with an acetonitrile–water (7:3) mixture as the mobilephase, maintaining a flow rate of 1 mL/min. A UV spectrophoto-metric detector was employed with a wavelength of 242 nm.

2.4. Microbial community analysis

Biofilm samples were scratched from the anode of the ferricya-nide-cathode MFC fed with 1000 mg/L glucose, 1000 mg/L glu-cose + 250 mg/L NB, and 250 mg/L NB, respectively. For eachsubstrate, the bacterial samples were collected after operation ofthe MFC for four cycles. Bacterial genomic DNA was extracted fromthe biofilm samples using the FastDNA� SPIN Kit for Soil (MP Bio-medicals, Santa Ana, CA) according to the manufacturer’sinstructions.

The universal primer sets V3-2 (50-ATTACCGCGGCTGCTGG-30)and V3-3 (50-CGCCCGCCGCGCGGCGGGCGGGGCGGGGGCACGGGGGGCCTACGGGAGGCAGCAG-30) (Invitrogen Biotechnology Co.,Ltd.) were used to amplify the V3 region of bacteria 16S ribosomalDNA (rDNA) from the extracted genomic DNA. PCR amplificationwas performed in a 25 lL reaction volume, including 17.37 lLddH2O, 2.5 lL 10 � PCR Buffer, 2.0 lL DNTP mixture, 0.5 lL V3-2,0.5 lL V3-3, 0.13 lL Taq polymerase, and 2 lL template. Theamplification involved an initial denaturation at 94 �C for 5 min;

J. Li et al. / Bioresource Technology 101 (2010) 4013–4020 4015

followed by 10 cycles, each of which included 30 s of denaturationat 94 �C, 30 s of annealing at 61 �C (the temperature of anneal de-creased 0.5 �C after each cycle), and 1 min of extension at 72 �C;then 25 cycles, each of which included 30 s of denaturation at94 �C, 30 s of annealing at 55 �C, and 1 min of extension at 72 �C,with a final 7 min extension at 72 �C.

The DGGE analysis of the PCR products was carried out in adenaturing gradient gel electrophoresis system (C.B.S. Scientific,Del Mar, CA, USA). The 8% (w/v) polyacrylamide gels (16 cm �16 cm gel, thickness of 0.75 mm) contain 40–60% denaturing gra-dients (urea and formamide). Electrophoresis was conductedusing a 1 � TAE buffer at 200 V and 60 �C for 5 h. After electro-phoresis, the gel was stained with 5 lL/(100 mL) ethidium bro-mide (EB) in 1 � TAE buffer for 15 min and destained in 1 � TAEbuffer for 10 min. The fragments were visualized under a UVtransilluminator.

16S rDNA gene fragments cut out from the DGGE gel were trit-urated, added into 20 lL TE, then centrifugalized at 15,496g for2 min after 30 min water bath at 50 �C. The supernatant fluidwas used for PCR amplification, and the PCR program was the same

0 10 20 300

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V)

Time (

A

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0

5

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30

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/m3 )

Current de

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Fig. 1. Influence of nitrobenzene (NB) on (A) voltage outputs (at the external resistance oon the voltages marked with the circles in (A).

as that mentioned above, but using the universal primer sets of V3-1 (50-CCTACGGGAGGCAGCAG-30) and V3-2 (50-ATTACGCGGC-TGCTGG-30). The PCR products were used for sequencing, and thenthe sequences were compared directly to all known sequencesdeposited in the GenBank databases using the basic local align-ment search tool (BLAST).

3. Results and discussion

3.1. Effect of NB on electricity generation in the ferricyanide-cathodeMFC

After the ferricyanide-cathode MFC was operated using1000 mg/L glucose as the fuel to obtain stable outputs of electricvoltage, mixtures of 1000 mg/L glucose + 50 mg/L NB, 1000 mg/Lglucose + 150 mg/L NB, 1000 mg/L glucose + 250 mg/L NB, and250 mg/L NB, and 1000 mg/L glucose sequentially were used inthe anode as the substrates. Some representative curves of voltageoutputs are shown in Fig. 1A. The MFC using the glucose-NB mix-

40 50 60h)

0 mg/L NB 50 mg/L NB 150 mg/L NB 250 mg/L NB

80 100 120 140nsity (A/m3

)

0 mg/L NB 50 mg/L NB 150 mg/L NB 250 mg/L NB

f 1000 X), and (B) power density in the ferricyanide-cathode MFC, calculated based

4016 J. Li et al. / Bioresource Technology 101 (2010) 4013–4020

tures as the fuel was operated successfully; however, the additionof NB affected electricity generation of the MFC. With the NB con-centrations increasing from 0 to 250 mg/L, the maximum voltageoutputs decreased from 670 to 489 mV, the maximum volumetricpower densities decreased from 28.57 to 8.47 W/m3, and the inter-nal resistances increased from 400 to 1000 X. The CE and electriccharge of the MFC decreased with the NB concentrations linearlywith the coefficients of determination (R2) 0.71 and 0.79, respec-tively (Table 1).

As shown in Fig. 1A, the maximum voltage outputs decreasedand the operation periods became shorter with increasing NB con-centrations. In addition, the voltage outputs were not able to re-turn to their original level immediately after the replacement of250 mg/L NB by 1000 mg/L glucose as the fuel at the final stage.The effect of NB on electricity generation of the MFC was more pro-found than that of some other electron-withdrawing compounds.Sun et al. (2009) reported that a concentration of active brilliantred X-3B (ABRX3) of 1500 mg/L resulted in a decrease of the max-imum voltage about 25%, whereas 250 mg/L NB in the anodechamber resulted in a decrease of the maximum voltage output27%. Sukkasem et al. (2008) showed that a nitrate concentrationof 496 mg/L did not affect the maximum voltage outputs at all.The decreasing voltage outputs and shortened operation periodsare possibly attributable to electron consumption by NB reduction,rapid consumption of the available carbon source by microbes at ahigher NB concentration, and/or inhibition of electricity-generat-ing microbes in the MFC because of the NB toxicity.

Table 1Comparison of the MFC performance with the different substrates: the mixtures of 1000 m

NB concentrations in the mixture (mg/L) Maximum voltage (mV) Maximum pow

0 670.0 ± 7.07a 28.57 ± 0.70450 597.7 ± 24.75 20.24 ± 0.750150 507.7 ± 7.78 9.29 ± 0.493250 489.0 ± 7.07 8.47 ± 0.551

a The values are mean ± standard deviation (n = 3).

020406080

100120140

020406080

100

0 50 1000

20406080

100

Rem

oval

(%)

150 mg

Tim

50 mg/L NB

Fig. 2. Removal rates of glucose, nitrobenzene (NB

As shown in Fig. 1B, NB affected power density significantly.Both the maximum power density and the ranges of current den-sity decreased quickly with the NB concentrations. The power den-sity was strongly reduced by NB and the internal resistanceincreased correspondingly. The presence of NB as alternative elec-tron acceptor and its toxic effects on the microbial physiology werethe dominant factors for the reduction in power output and the in-crease of the internal resistance. The competition for electrons byNB could lead to a reduction in electron supply. In addition, theproton–electron transfer process in the MFC might be impeded be-cause of NB toxicity. The addition of NB might change the commu-nity structure and decrease the population of certain microbessuch as Hansenula anomala, which could transfer electrons directlyto electrode surfaces in presence of glucose as the carbon source(Prasad et al., 2007). Compared to previous studies (Bond and Lov-ley, 2003; Chaudhuri and Lovley, 2003), CEs of the MFC were lowerin this study. Many factors might attribute to low CEs, includingsubstrate utilization for methanogenesis and the electron transferfrom substrate to other non-electrode electron acceptors in thesolution, such as nitrate and oxygen (Liu and Logan, 2004). Theconsumption of electrons for anaerobic degradation of NB maybe the most important reason for the low CEs since under anaero-bic conditions, the reductive reaction of NB occurs first due to thestrong electron affinity of the nitro group (Gurevich et al., 1993).

To examine if NB could be used as the fuel of the MFC, the con-centration of NB was maintained at 250 mg/L and the concentra-tions of glucose were set up at 1000, 750, 500, 250, and 0 mg/L,

g/L glucose with different concentrations of nitrobenzene (NB).

er density (W/m3) Internal resistance (X) CE (%) Electric charge (C)

400.0 ± 70.71 2.70 ± 0.096 65.10 ± 2.322600.0 ± 0.00 1.48 ± 0.042 43.50 ± 11.456950.0 ± 70.71 1.24 ± 0.078 35.48 ± 0.7061000.0 ± 0.00 1.00 ± 0.156 30.32 ± 0.588

150 200 250 300

glucose

NB

250 mg/L NB/L NB

e (h)

COD

), and COD in the ferricyanide-cathode MFC.

J. Li et al. / Bioresource Technology 101 (2010) 4013–4020 4017

respectively. When 250 mg/L NB was used as the sole fuel, a volt-age about 70 mV was produced, but it gradually decreased to30 mV within 140 h. The result showed that NB was not suitableas a sole fuel of the MFC because NB was a strong electron acceptorand NB degradation proceeded by a partial reductive pathway inthe anaerobic anode chamber.

3.2. Chemical removal in the ferricyanide-cathode MFC

The NB degradation and organic matter removal in the ferricy-anide-cathode MFC were investigated with different initial NB con-centrations (50–250 mg/L) using glucose as a co-substrate. Fig. 2shows the removal rates of glucose, NB, and COD in the ferricya-nide-cathode MFC. Using the traditional anaerobic degradationmethod in a hybrid reactor, Majumder and Gupta (2003) showedthat the removal rates of NB and COD were 80–90% and 60–96%,respectively. In our experiment, the removal rates of NB werenearly 100% for all the NB initial concentrations and the COD re-moval rates were 87–98% in the MFC. The result showed that NBcould be biodegraded successfully in the MFC with glucosetogether.

NB anaerobic degradation relies on the addition of biodegrada-tion organic compounds as electron donors (Li et al., 2007). Thehigher removal rate of NB in the MFC in this study might be attrib-uted to the presence of glucose, which was easily biodegraded tosupply electrons to increase the metabolic rate of anaerobic bacte-ria with sufficient anaerobic terminal electron acceptors (Morriset al., 2009).

3.3. Microbial community

Fig. 3 shows the DGGE profiles of the 16S rDNA gene fragmentsamplified from the extracted DNA of the biofilms on the anodes.

Fig. 3. PCR-DGGE analysis of 16S rDNA extracted from the electrodes of the MFC fed witnitrobenzene mixture, NB: 250 mg/L nitrobenzene).

Table 2Characterization of isolated 16S rRNA gene fragments derived from the anodes of the MFC

Sequence designation Accession number BLAST search

Band 1a GQ870291 GQ247734.1EGQ465230.1 E

Band 2b GQ870292 GQ129889.1 UAF368755.1 PFJ786064.1 PsEU723136.1 PEU112283.1 U

Band 3b GQ870293 GQ503869.1 DFJ393114.1 DEF592815.1 U

Band 4a GQ870294 GQ390422.1 UEU704648.1 UFJ262598.1 U

a Samples from the anodes of the MFC with glucose as the fuel.b Samples from the anodes of the MFC with mixture of glucose and nitrobenzene, nit

The results suggest that the presence of NB caused changes withrespect to the dominant bacterial species on the anodes, includingchanges in relative abundance and emergence of new bacteria. TheDNA gene sequences represented by bands 1–4 were submitted tothe GenBank of the NCBI and the accession numbers obtained fromthe GenBank database were GQ870291–GQ870294. Based on acomparative analysis of the 16S rRNA gene sequences, severalgroups of bacterial species were identified. The BLAST results aresummarized in Table 2.

The BLAST results indicated that the dominant bacterial specieson the anodes of the MFC with glucose as the fuel shared 100% and88% 16S rDNA sequence homology with Enterobacter sp. (band 1)and Geobacter sp. (band 4), respectively. Geobacter sp. has been fre-quently reported to be responsible for power generation in theMFC (e.g., Bond and Lovley, 2003; Kaufmann and Lovley, 2001).Addition of NB resulted in decreased intensities of bands 1 and 4,and increased intensities of bands 2 and 3. Band 2 might belongto Pseudomonas sp. or Comamonadaceae bacteria, both of whichare able to degrade NB (He and Spain, 1999). Band 3 shared 97%16S rDNA sequence identity with Desulfovibrio sp., a sulfur-reduc-ing bacterium that can oxidize acetate in a two-electrode fuel cell(Bond et al., 2002).

Above results suggested that the addition of NB changed theelectricity-generating bacteria structure and led to rapid increaseof NB degrading bacteria. Therefore, the changes in the microbialcommunity structure should be another factor that affects electric-ity generation of the MFC.

3.4. Power generation from the NB-cathode MFC

To examine if NB was involved in the electricity generation pro-cess by accepting electrons from circuit without any externalpower input, the dual-chamber NB-cathode MFC was operated

h different substrates (G: 1000 mg/L glucose, G + NB: 1000 mg/L glucose + 250 mg/L

s with different substrates.

results Identity (%)

nterobacter sp. FW17a 100nterobacter sp. IPB54 100

ncultured Pelomonas sp. clone GI8-sp-D04 98seudomonas saccharophia 98eudomonas sp. III-116-17 97seudomonadales bacterium kmd_137 97ncultured Comamonadaceae bacterium B00055B41 97

esulfovibrio sp. enrichment culture clone Ecwsrb032 97esulfovibrio sp. clone MFC-B162-E12 96ncultured sulfate-reducing bacterium clone 2R2V06 96

ncultured Geobacter sp. clone C12 88ncultured bacterium clone MFC-GIST 439 88

ncultured Geobacter sp. clone MFC-A36 84

robenzene as the fuel.

4018 J. Li et al. / Bioresource Technology 101 (2010) 4013–4020

using 1000 mg/L glucose as the anode fuel and 250 mg/L NB as thecathode electron acceptor. Some typical curves of electrical voltageoutputs are presented in Fig. 4A. The MFC was operated for threecycles continuously for a total of about 60 h, the operation periodper cycle varied from 18 to 25 h, and the maximum voltage outputin each cycle was about 400 mV (387–410 mV). The voltage outputreached the maximum value rapidly within 1 h after replacing thenew substrate, and then decreased to about 280 mV within 6 h,and slightly increased to 285 mV for about 5 h, and finally gradu-ally decreased.

Fig. 4B shows the relationships between the cell potential andpower density vs. the current density. The cell potential decreasedlinearly with the current density. Based on Fig. 4B and Eq. (3), aninternal resistance of 626 X was obtained. The maximum powerdensity achieved from the NB-cathode MFC was 6.30 W/m3

(69.442 mW/m2) at the current density of 19.72 A/m3 (0.2173 A/m2). The performance of the NB-cathode MFC was worse thanthe ferricyanide-cathode MFC using 1000 mg/L glucose as the fuelreported above, which should be closely related to the negative re-dox potential of NB. Nevertheless, the result indicated that in thetested NB-cathode MFC system, NB could accept electrons throughthe circuit and involve in the electricity generation process.

0 10 20 3050

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Fig. 4. (A) Electricity generation (at the external resistance of 1000 X), and (B) polarizavoltage marked with the circle in (A).

3.5. Cathodic reduction of NB in the NB-cathode MFC

In the NB-cathode MFC, NB was degraded almost completely. Asshown in Fig. 5, the removal efficiency of 250 mg/L NB was up to73% when the voltage output reached the maximum value (about400 mV). Then, after about 10 h of operation the voltage output in-creased slightly to 285 mV, and the removal efficiency reached 88%correspondingly. The removal efficiency reached 98% at the end ofthe cycle. These results can be explained by the production ofintermediates during NB reduction that allowed maximum voltageoutput. These intermediates could include phenylhydroxylamine,nitrosobenzene, azoxybenzene, and azobenzene (Li et al., 2007;Oliveira, 2003). These intermediates were then reduced to anilineand voltage output was maintained. It is known that aniline isthe end product of NB reductive degradation at the cathode of bio-electrochemical systems (Mu et al., 2009), but in our reactor sys-tem, Pt was used as catalyst and the solution was strongly acidic.Under these conditions, aniline was prone to polymerization topolyaniline (Gangopadhyay and Amitabha, 2000). Further researchis needed to confirm this hypothesis.

Cathodic reduction of NB in the MFC was more efficient thanconventional electrochemical reduction (Li et al., 2007). In our

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tion and power density of the nitrobenzene-cathode MFC, calculated based on the

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Fig. 5. Removal rates of nitrobenzene (NB) in the nitrobenzene-cathode MFC.

J. Li et al. / Bioresource Technology 101 (2010) 4013–4020 4019

experiment, the average removal rate within the initial 1 hreached 3.04 mg/(L min), which was higher than the average re-moval rate (2.34 mg/(L min)) reported by Li et al. (2007). Fur-thermore, the conventional electrochemical reduction requiredan external power input, whereas the cathodic reduction of NBin the MFC did not consume but produced electricity power.The removal efficiency obtained in this study is comparable tothat of some studies on anaerobic degradation. For example,Zhang et al. (2007) reported that with Fe2+/Fe3+ concentrationsof 100–200 mg/L, the NB anaerobic degradation increased,resulting in a NB removal efficiency of 96%. Liu et al. (2007) ob-served that the NB removal efficiency reached up to 99% at con-centrations ranging from 50 to 180 mg/L in a sequencing batchreactor (SBR). Therefore, our results may provide a novel ap-proach for simultaneous electricity generation and treatment ofwastewaters containing NB, and even other nitroaromaticcompounds.

In this study, NB was degraded efficiently in the anode andcathode chambers of the ferricyanide-cathode MFC and the NB-cathode MFC, respectively. In the anode and cathode chambers,the removal rates of NB reached 99% and 98% within 24 h, andthe average removal rates within the initial 1 h were 2.35 and3.04 mg/(L min), respectively. However, the degradation mecha-nisms in the anode and cathode chambers of the two MFCs wereobviously different. In the anode chamber of the ferricyanide-cath-ode MFC, NB was degraded by anaerobic microorganism through apartial reductive pathway. In the cathode chamber of the NB-cath-ode MFC, the NB reduction was just a pure electrochemical reduc-tion process, no microorganism involving.

4. Conclusions

The effect of NB on the performance of the ferricyanide-cathodeMFC and simultaneous electricity generation and reduction in theNB-cathode MFC were assessed. The addition of NB significantlyinhibited electricity generation by the ferricyanide-cathode MFCprimarily due to electron competition between NB and the anode.The presence of NB resulted in changes of the dominant bacterialspecies on the anodes possibly because of NB toxicity. In the NB-cathode MFC, NB participated in the electricity generation process.In addition, NB was degraded efficiently in the anode and cathodechambers of the ferricyanide-cathode MFC and the NB-cathodeMFC, respectively.

Acknowledgements

This work was partially supported by grants the Natural ScienceFoundation of China (Nos. 50608070 and 50779080).

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