Optimal Production of Bioelectricity using Clostridium sporogenes

NCIM 5125

 

M. Seenuvasan¹, J. Sebastin¹, K. Ananthi¹, S.C. Gayathri¹, A. Krithika Siva¹, M. Anil Kumar¹ and K. Sathish Kumar²*

¹Department of Biotechnology, Madha Engineering College, Chennai.

²Department of Chemical Engineering, SSN College of Engineering, Chennai.

 

ABSTRACT:

A sequential anode-cathode double-chamber microbial fuel cell (MFC) was constructed for the production of electricity using a novel microorganism, Clostridium sporogenes NCIM 5125 which is profoundly known to be a part of MFC studies. The growth of the microorganism was supported by energy derived from the electron transfer process itself and results in stable, long term power production. The electricity generation was optimized by analyzing the growth of the microbe at different parameters using mediated and non-mediated MFC. The results suggested that it is feasible to generate more electricity amounting to 5.6 mA at 160^th h with the adopted C. sporogenes using mediated MFC.

 

 

 

INTRODUCTION:

As we head into the future, large portion of energy produced and used in the world will be from sustainable sources due to the world’s limited supply of fossil fuels and their impact on environmental and economic changes. Microbial fuel cell studies have been carried out since 1970’s. The microbial Fuel Cell (MFC) technology generates either electricity or hydrogen from bacterial growth in carbon-containing solutions. A microbial fuel cell is a device that uses bacteria to catalyze the conversion of organic matter into electricity^1-8. Bacteria generate electrons and protons at the anode by oxidizing the substrate. Electrons are transferred through the external circuit while the protons diffuse through the solution to the cathode, where the electrons combine with protons and oxygen to form water^9-10.

Currently most MFC research focuses on increasing the power density of the system based on the projected surface area of electrodes and/or the reactor volume, determining the effects of varying fuel cell components and the parameters of MFC construction on the voltage output¹¹. The MFC has operational and functional advantages over the technologies currently used for generating energy from organic matter; first, they are capable of working at ambient temperature. Secondly, they enable the direct conversion of the substrate into energy. Thirdly, they do not require any gas treatment. Finally, the cathode does not require forced aeration as it is passively aerated. Some recent developments allow high conversion efficiencies of simple carbohydrates like glucose and complex carbohydrates like starch.

 

As an alternative strategy, anaerobic digestion of wastewater with concomitant methane formation has been developed, but until now, this strategy is not widely applied, mainly due to the low COD concentrations in the domestic sewage, which require a reactor dimension that is economically not feasible.

 

The aim of the present study is to determine the possibility of maximum electricity generation using microbial fuel cell. To the best of our knowledge, it is for the first time that Clostridium sporogenes NCIM 5125 is reported to serve as a biocatalyst in MFC studies. The maximum potential difference was recorded at stable operating conditions.

 

MATERIALS AND METHODS:

Microorganism and Culture Conditions:

The pure culture Clostridium sporogenes NCIM (5125) procured from National Collection of Industrial Microorganism, Pune, was raised on Fluid Thioglycollate Medium containing (g/L):  pancreatic digest of casein, 15.0; yeast extract, 5.0;  NaCl, 2.5; dextrose, 5.5; L-cysteine, 0.5; sodium thioglycollate, 0.5; agar, 0.75; resazurin, 0.01. It was incubated anaerobically at 37˚C at pH 6.8 for 48 h.

 

Medium:

The medium used for electricity production was Clostridium broth medium (specific media) containing (g/L): meat extract, 10.0; peptone, 10.0; yeast extract, 3.0; D (+) glucose, 5.0; starch,1.0; NaCl, 5.0; sodium acetate, 3.0;  L-cysteine chloride, 0.5; agar-agar,­  0.5. The reinforced Clostridium media (general media) which contained (g/L): casein peptone, 10.0; beef extract, 10.0; yeast extract, 3.0; dextrose, 5.0; NaCl, 5.0; sodium acetate, 3.0; soluble starch, 1.0;  L-cysteine hydrochloride, 0.5 was used for optimization.

 

MFC configuration and operation:

The microbial fuel cell (Fig. 1) is a dual chamber system that was designed and fabricated of acrylic material to avoid biofouling. A proton exchange membrane (Nafion 117, DuPont TM) separated the anode and cathode compartments, each consisting of an equal volume of 3000 mL. The membrane was pretreated in order to increase its efficiency to transfer protons. The anode and cathode were made of plain graphite electrodes (7x200x30mm). Before the start of this study, the anode compartment was purged with nitrogen (N₂) gas to maintain anoxic conditions. The anode chamber was filled with culture media (working volume: 2500 mL) and continuously purged with N₂ gas to maintain anaerobic condition. The cathode chamber was filled with 0.1 M potassium phosphate buffer and the pH was adjusted to 6.8. The cathode chamber was provided with subsequent aeration. The current generations were noted at an interval of 20 h and fresh inoculum was injected once the current started decreasing. The fresh medium was injected continuously with respect to the CO₂ production by means of CO₂ collector by replacing the exhausted feed. The MFC was maintained at a constant resistance of 10 Ω and the current generation was measured using a digital multimeter.

 

Fig. 1. Dual chambered MFC

 

Optimization and Calculations:

The optimization procedures were carried out in order to provide suitable growth environment for the microorganisms, since better the growth of microbes, larger is the current production. The adaptation of the microorganism was analyzed by optimizing parameters such as media, pH and mediator concentrations. The media is an important parameter which plays a major role in the growth of the microorganism and hence results in power generation. As it influences the growth of the microorganisms, it thereby contributes to increased electricity production. The reinforced Clostridium media and the Clostridium broth media were taken for analyses. On a similar basis, pH and mediator concentrations were also analyzed for the optimal growth of the microbe.  The current measurements were recorded every 20 h at a constant resistance of 10 Ω. The calculations were made by using the equations,

where, I is the current in Amperes, V is the voltage in volts, P is the power in Watts, R is the resistance in ohms and A is the area in cm².                                                          

 

RESULTS AND DISCUSSION:

Effect of Media Composition on Electricity Production:

The growth of Clostridium sporogenes at its maximum level was observed in two different media. Both the media showed significant results, out of which clostridium broth media was found to be effective which might be because it facilitated the growth of only Clostridium species and restricted the growth of other microbes. Whereas the reinforced clostridium media was found to support the growth of clostridium species and other microbes as well.

 

When employing the reinforced Clostridium media for the growth of C. sporogenes, the current generation was calculated at an interval of 20 h and after each interval, there was an increase in current production. The microbes use the given media as the substrate and generate electricity. The growth of microorganism directly influenced the electricity production, and as the microbe reached its end phase, there was a decrease in the electricity production and hence the feed was injected (first cycle) into the anode (Fig. 2). After the feed injection, there was an increment in the current generation thus reaching a maximum value of 1.89 mA (at 200^th h). Furthermore, as the microbe grew, there was an alternative change in the current generation. A second cycle of feed injection (at 240^th h) did not enhance the current production greatly as that of first cycle.

 

While using Clostridium broth media the current production was found to be maximum when compared to that of reinforced Clostridium media. During the initial feeding, the current production reached a maximum peak and as the organism reached the stationary phase, there was a subsequent decrease in the current generation. As the first cycle of feed injection (at 140^th h) was introduced, the electricity generation reached a maximum value of 2.71 mA which was the maximum output recorded.

 

Fig. 2. Bioelectricity production on different media

 

 

Effect of pH on Electricity Production:

The pH plays a significant role in bacterial growth and hence in current production. The Clostridium sporogenes is partly an acidophilic bacterium and was found to grow rapidly at an optimal pH of about 6.8 at which the maximum current production was 2.83 mA (Fig. 3), which was found to be higher than that when grown under pH 5.8 and 7.8. The pH environment of 5.8 was considerably acidic in nature and hence supported the growth of C. sporogenes thus generating a maximum output of 1.95 mA (at 180^th h) and this output was observed after the first feed cycle. The pH 6.8 might have had a profound impact on the Clostridium species growth since the current production reached a maximum of 2.83 mA (at 160^th h). The organism would have given rise to this maximum power output as the pH 6.8 could have provided a favorable and optimal environment to support its growth. The maximum current generation reached during the first cycle of feed injection. However, the second cycle did enhance the growth of C. sporogenes but maximum output was not achieved.

 

Fig.  3. Bioelectricity production at varying pH levels

 

Effect of Mediator Concentration on Electricity Production:

The mediator is a component used to trap out the electrons present on the plasma membrane of the microbe. The mediator usually used is a dye compound which is toxic in nature. However, when added in fewer concentrations it enhances the electricity production. This analysis was carried out in order to find the optimum concentration of mediator  used for MFC study. The mediator used in this case was methylene blue, which acted as electrophore and enhanced the current generation largely. As the mediator was added (0.1 mg/L in Clostridium broth media at pH 6.8), it enhanced the activity of trapping out the electrons from the organism cell wall and in increasing the current production (Fig. 4). During the initial feeding, there was an increased current output. After the first cycle, a maximum current of 3.17 mA (at 180^th h) was obtained. This was found to be the maximum output during the two cycles.

 

The mediator composition of 0.5 mg/L was the optimal concentration for the microbial growth, thus producing a maximum output of 5.6 mA (160^th h). The addition of mediator enabled the current generation at the earliest possible (i.e., peak obtained during the first cycle indicates maximum output). During the second cycle, a peak was obtained, producing maximum output of 4.21 mA (at 240^th h) which was not the maximum as that produced during the first cycle.

 

The mediator concentration of 1.0 mg/L was the maximum composition added and as the concentration increased the current production decreased which implies that the mediator, at its increased concentration, would have inhibited the growth of the organism. In addition, the current output was found to be a maximum of 1.08 mA (at 140^th h). This indicates that due to the addition of an external chemical, there is subsequent increase in current output at a relatively early time. However, during the second cycle, the growth of Clostridium sporogenes was found to be inhibited, thus showing a fluctuation in electricity production.

 

Fig. 4. Bioelectricity  production  at  vary ing mediator concentrations

 

Effect of pH at Constant Mediator Composition on Electricity Production:

The effect of pH with respect to optimal mediator concentrations did not give additional current production (Fig. 5). Thus, it is confirmed that optimal mediator concentration would support the maximum growth of Clostridium sporogenes.

 

Fig. 5. Bioelectricity production at optimal mediator  concentrations with varying pH

CONCLUSION:

From this investigation, the designed microbial fuel cell was found to be capable of generating a maximum current output of 5.6 mA at 160^th h using Clostridium sporogenes NCIM 5125  in the presence of methylene blue (0.5 mg/L) as an electrophore, when cultivated in clostridium broth media at 37˚C at pH 6.8. This study has thus showed that the electricity generation would be possible with the use of C. sporogenes on its growth in a mediated MFC system.

 

ACKNOWLEDGEMENT:

The authors are indebted to all the colleagues, especially those from the Department of Biotechnology, Madha Engineering College, Chennai for their valuable suggestions on the laboratory work and manuscript preparation as well.

 

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Received on 17.11.2012                             Accepted on 20.01.2013        

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