Advances in Microbiology, 2012, 2, 332-339 Published Online September 2012 (
Conversion of Carbon Dioxide to Metabolites by
Clostridium acetobutylicum KCTC1037 Cultivated with
Electrochemical Reducing Power
Bo Young Jeon1, Il Lae Jung2, Doo Hyun Park1*
1Department of Biological Engineering, Seokyeong University, Seoul, South Korea
2Department of Radiation Biology, Environmental Radiation Research Group,
Korea Atomic Energy Research Institute, Daejeon, South Korea
Email: *
Received June 21, 2012; revised July 29, 2012; accepted August 6, 2012
In this research, metabolic fixation of CO2 by growing cells of C. acetobutylicum cultivated with electrochemical re-
ducing power was tested on the basis of the metabolites production and genes expression. In cyclic voltammetry, elec-
trochemical oxidation and reduction reaction of neutral red (NR) immobilized in intact cells of C. acetobutylicum was
stationarily repeated like the soluble one in the condition without CO2 but the electrochemical reduction reaction was
selectively increased by addition of CO2. In electrochemical bioreactor, the modified graphite felt cathode with NR
(NR-cathode) induced C. acetobutylicum to generate acetate, propionate, and butyrate from CO2 in defined medium.
When H2 and CO2 were used as an electron donor and an electron acceptor, respectively, C. acetobutylicum also pro-
duced the same metabolites in a defined medium. C. acetobutylicum was not grown in the defined medium without sub-
stituted electron donors (H2 or electrochemical reducing power). C. acetobutylicum cultivated with electrochemical re-
ducing power produced more butyrate than acetate in complex medium but produced more acetate than butyrate in de-
fined medium. The genes of encoding the enzymes catalyzing acetyl-CoA in C. acetobuylicum electrochemically culti-
vated in defined medium than conventionally cultivated in complex medium. These results are a clue that C. acetobu-
tylicum may metabolically convert CO2 to metabolites and produce free energy from the electrochemical reducing power.
Keywords: C. acetobutylicu m ; CO2-Assimilation; Electrochemical Reducing Power; Coupling Redox Reaction
1. Introduction
Chemoautotrophs that regenerate reducing power and
produce free energy in coupling with oxidation of H2 can
more effectively fix CO2 than ammonium, nitrite, and
ferrous-oxidizing bacteria because redox potential of
H2/2H+ (–0.42 V vs. NHE) is lower than NAD+/NADH
(–0.32 V vs. NHE) [1]. The reducing power generated in
coupling with oxidation of ammonium, nitrite, and fer-
rous ion can’t induce regeneration of NAD(P)H without
a reverse electron transport system coupled to consump-
tion of external energy [2-5]. Experimentally measured
redox potential of NR is –0.325 V (vs. NHE), which is
theoretically enough to mediate generation of electron-
driving force from electrode to NADH [6]. Practically,
electrochemically reduced NR catalyzes NADH regen-
eration by non-enzymatic catalysis [7]. Theoretically,
electrochemically reduced NR may be more effective in
reducing power than H2 on the basis of non-enzymatic
catalysis of NADH regeneration.
C. acetobutylicum is a typical fermentation bacterium
that produces acetate, propionate, and butyrate by the
coupling redox reaction of reducing power and ace-
tyl-CoA generated from metabolic oxidation of glucose,
and also autotrophically produces acetyl-CoA from the
coupling redox reaction of H2 and CO2 in defined me-
dium with organic nitrogen nutrient [8,9]. The acetyl-
CoA is metabolically oxidized to acetate coupled to re-
generation of NADH or reduced to butyrate coupled to
oxidation of NADH [10]. Biological production of or-
ganic polymers, fatty acids and alcohols from H2 and
CO2 has been studied in order to decrease global warm-
ing and produce renewable energy and useful biomass
[11,12]. H2 is the most effective electron donor to bacte-
riologically fix CO2 on the basis of its redox potential but
is less practical to apply to biological system owing to
very low solubility in water and possible explosiveness
in the process of use, transport and storage. The cost for
production of H2 is not economical in comparison of
glucose price. In order to solve the problem caused by
the H2 production cost, an alternative technology has
*Corresponding author.
opyright © 2012 SciRes. AiM
B. Y. JEON ET AL. 333
been developed [13,14].
Electricity generated from the solar cell can be directly
converted to biochemical reducing power in coupling
with the redox reaction of NR immobilized in bacterial
cell or graphite felt electrode [6]. Bacterial CO2 fixation
induced by biochemical reducing power electrochemi-
cally regenerated by the solar cell electricity may corre-
spond to the photosynthesis because O2 is generated from
anode compartment and CO2 is biochemically assimi-
lated into biomass and converted to metabolites in cath-
ode compartment [15]. Covalently immobilized NR in
graphite felt electrode can function as a catalyst for NADH
regeneration and a redox carrier for electron transfer
from electrode to bacterial cell [16]. Redox potential of
NR is –0.325 volt (vs. NHE), which is 0.05 volt lower
than NAD+. The electrochemical redox reaction of NR
can be coupled to biochemical redox reaction as follows:
[NRox + 2e + 1H+ NRred; NRred + NAD+ NRox +
NADH]. Commonly, NRox and NAD+ are reduced to NRred
and NADH, respectively by accepting two electrons from
electrode and NRred (ox: oxidation; red: reduction).
In this study, electrochemical reducing power was
charged to C. acetobutylicum culture using the NR-
cathode to induce autotrophic production of acetate and
butyrate from CO2 in a defined medium and increase of
butyrate production in a complex medium. The NR-
cathode, to which –2 V of DC-electricity was charged,
may be an optimized habitat for strict anaerobes because
the lower oxidation-reduction potential than –300 mV (vs.
NHE) is electrochemically generated and the electro-
chemically reduced NR may catalyze bacterial NADH
2. Materials and Methods
2.1. Medium
Reinforced clostridial (RC) medium (Tryptone 10 g/L,
Sodium chloride 5 g/L, Beef extract 10 g/L, Yeast extract
3 g/L, Glucose 20 g/L, Starch 1 g/L, L-cystein hydro-
chloride 0.5 g/L, Sodium acetate 3 g/L) was used as a
complex medium and for successive cultivation of C.
acetobutylicum. M9 mineral medium (Disodium phos-
phate 6.8 g/L, Monosodium phosphate 3 g/L, Ammo-
nium chloride 5 g/L, Sodium chloride 0.5 g/L, Magne-
sium sulfate 0.246 g/L, Calcium chloride 0.0147 g/L)
supplemented with sodium bicarbonate (25 mM) and
yeast extract (3 g/L) was used as a defined medium. Fifty
ml of RC or defined medium was prepared in anaerobic
serum vials (total volume 165 ml) whose headspace was
filled with 2 atmospheres of oxygen-free N2 or H2.
2.2. Electrochemical Bioreactor
An electrochemical bioreactor that was designed for con-
tinuous culture in previous research [16] was partially
modified for cultivation of strict anaerobic bacterium in
batch culture, as is shown in Figure 1. The electro-
chemical bioreactor (inner diameter, 80 mm; height, 200
mm; medium volume, 500 ml; electrode volume, 250 ml;
total volume, 1000 ml; Pyrex, USA) with a built-in anode
compartment was designed to equalize distance between
anode and all round of cylindrical cathode. A sintered
glass filter (diameter, 50 mm; thickness, 5 mm; pore, 1 -
1.6 μm, Duran, Germany) that was modified with cellu-
lose acetate film (35 μm thickness, Electron Microscopy
Sciences, USA) was fixed at the bottom end of the
tube-type anode compartment (inner diameter 20 mm;
height, 150 mm; working volume, 50 ml). Cellulose ace-
tate film attached to the sintered glass functions as a
semipermeable membrane capable of selectively trans-
ferring water, gas, and proton. Five hundred ml of the
media was prepared in the electrochemical bioreactor
(Figure 1) to which O2-free CO2 (50 ml·min–1) was con-
tinuously supplied during cultivation. Inoculation ratio
was adjusted to 5% (w/w) of medium volume. DC –2 V
of electricity was charged to NR-graphite felt cathode to
induce electrochemical reduction reaction of NR for C.
acetobutylicum. The defined medium was used as ano-
lyte to avoid generation of osmosis between anode and
cathode compartment.
2.3. Electrode
Graphite felt (thickness, 10 mm; height, 200 mm; length,
Figure 1. Electrochemical bioreactor composed of graphite
felt modified with NR (NR-cathode), glass filter membrane
modified with cellulose acetate film, and platinum anode.
Copyright © 2012 SciRes. AiM
500 mm; Electrosynthesis, USA) was rolled up to be a
cylinder type (internal diameter, 40 mm; external diame-
ter, 75 mm). Neutral red was immobilized to thegraphite
felt (10 × 200 × 500 mm, Electrosynthesis, USA) by the
covalent bond between neutral red and polyvinyl alcohol
(mean molecular weight, 80,000, Sigma, USA) according
to the technique used in previous research [16]. The
graphite felt modified with NR was used as a cathode
and a platinum wire (thickness 0.5 mm, length 150 mm)
was employed as an anode. Electric potential charged to
NR-cathode was precisely adjusted to –2 V.
2.4. Analysis of Electrochemical Reaction of
C. acetobutylicum
The cyclic voltammetry was employed in order to ana-
lyze electrochemical redox reactions between electrode
and intact cell of C. acetobutylicum. The cyclic voltam-
metry was conducted using a voltammetric potentiostat
(BAS model CV50W, USA) linked to a data acquisition
system. Aglassy carbon electrode (5mm diameter, Elec-
trosynthesis, USA), a platinum wire, and an Ag/AgCl
electrode (redox potential, +0.2 V vs. NHE, Electrosyn-
thesis, USA) were utilized as a working electrode,
counter-electrode, and reference electrode, respectively.
The reactant was composed of 25 mM Tris-HCl buffer
(pH 7.5) containing 5 mM NaCl and 100 μM NR. C.
acetobutylicum that was anaerobically cultivated in the
modified M9 medium for 48 hr under H2 atmosphere was
anaerobically centrifuged at 1500 × g and 4˚C for 60 min.
The precipitated bacterial cells were suspended in 0.05
volume of the oxygen-free reaction mixture, in which NR
was spontaneously immobilized in bacterial cells. Prior
to and during the cyclic voltammetry measurement, ar-
gon (99.999%) was sparged into headspace of reaction
beaker in order to protect contamination of the reaction
mixture by oxygen. The scanning rate was 25 mV·s–1 over
a range of 0 to –1200 mV. During cyclic voltammetry for
NR dissolved in reactant or immobilized in C. acetobu-
tylicum, the variations of upper voltammetric peaks (an
indicator for electron transfer from electrode to bacterial
cells through NR) and lower voltammetric peaks (an in-
dicator for electron transfer from bacterial cells to elec-
trode through NR) by addition of CO2 were recorded.
2.5. Analysis of Metabolites
Bacterial metabolites were analyzed using a Gas Chro-
matography/Mass Spectrometry (Clarus 600 series +
TurboMatrix HSS Trap, PerkinElmer, USA) equipped
with Elite-FFAP column (ID 0.25 μm, OD 0.32 μm,
length 30 m) and electron ionization system. Bacterial
culture was centrifugation at 10,000 g and 4˚C for 30
min and filtered with membrane filter (pore 0.22 μm),
and then directly injected into the GC/MS injector. Con-
centration of metabolites was determined based on peak
area of standard compounds and chemical species was
determined based on mass profile database.
2.6. Microarray of mRNA
C. acetobutylicum was cultivated in the electrochemical
bioreactor using the defined medium under strict anaero-
bic CO2 atmosphere and in the complex medium under
strict anaerobic N2 atmosphere for 5 days. Total RNA
was isolated and purified from harvested bacterial cells
using a RNA purification kit (Total RNA, spin-column
format, Oligotex mRNA mini kit, Qiagen Korea, Seoul).
Microarray analysis of mRNA was conducted at Ge-
nomictree (Daejeon, Korea) using the systems, kits,
DNA chips, and analysis software offered by Agillent
Technologies (Korea branch, Seoul) via a turnkey-based
analyzing order. The significantly expressed genes that
are concerned with CO2 fixation and energy metabolism
were selectively analyzed to compare the relationship
between metabolic pathway related with CO2 fixation
and cultivation conditions.
3. Results
3.1. Electrochemical Redox Reaction in Coupling
with CO2
Cyclic voltammetry is a useful technique to measure
electrochemical coupling redox reaction of electron me-
diator immobilized in bacterial cells. In the cyclic volt-
ammetry without bacterial cells, the electrochemical re-
dox reaction of NR was measured to be –0.52 V (vs.
Ag/AgCl), which is very similar to the experimental
value –0.525 V (vs. Ag/AgCl) measured in standard con-
dition. Both the upper and lower voltammetric peaks
were not altered by addition of CO2 as expected; in con-
trast, the upper voltammetric peak generated by NR im-
mobilized in C. acetobutylicum was shifted upward from
2.4 to 2.9 μA and rightward from –0.52 to –0.55 V by
addition of CO2 as shown in Figure 2. Increase of 0.4 μA
of current indicates that electrons are transferred from
electrode to bacterial cells coupled to redox reaction of
NR. Increase of –0.3 V of redox potential is a clue that
electrons are transferred from electrode to NR by lower
electrode (working electrode) potential than intrinsic
redox potential of NR. Relatively higher electron-driving
force (electrode potential) may be required for electrons
to move through the electric resistance generated be-
tween electrode and NR immobilized in bacterial mem-
brane. Meanwhile, other upper voltammetric peak (bold
arrow mark) located at –0.9 V (vs. Ag/AgCl) was also
shifted upward from 2.7 to 3.0 μA and rightward from
–0.9 V to –0.93 V by addition of CO2. It seems possible
that electrons are transferred from the electrode via one
of the electron carriers located in the bacterial membrane,
Copyright © 2012 SciRes. AiM
Copyright © 2012 SciRes. AiM
allowing for current and potential increase by addition of
CO2. CO2 could act as an electron acceptor to induce
biochemical oxidation of NADH that may be electro-
chemically regenerated, by which electrons may be
transferred from electrode to bacterial cells via the cou-
pling redox reaction of NR and NAD+.
3.2. Growth and Metabolite Production of
C. acetobutylicum
C.acetobutylicum did not grow and didn’t produce me-
tabolites in the defined medium under N2 atmosphere
(DM-N2) but grew and produced acetate, propionate, and
butyrate under H2 atmosphere (DM-H2), as shown in
Table 1. The metabolites detected in the chromatography
for culture fluid of C. acetobutylicum cultivated in the
DM-N2 may have originated from the metabolites con-
tained in the inoculum. The electrochemical reducing
power generated from NR-cathode (reduced NR) is con-
verted to the biochemical reducing power (NADH),
which can be presumed on the basis of the growth and
metabolite production of C. acetobutylicum cultivated in
the DM-ER. C. acetobutylicum cultivated with electro-
chemical reducing power produced more acetate than
butyrate in DM-ER but more butyrate than acetate in
CM-ER. These are more clues that biochemical reducing
power (NADH) may be regenerated by electrochemical
reducing power generated from –2 V of NR-cathode and
the high balance of NADH/NAD+ may induce metabolic
conversion of CO2 to metabolites in coupling with free
energy synthesis.
3.3. Quantitative and Qualitative Verification of
Figure 2. Cyclic voltammetry for NR dissolved in reactant
and immobilized in C. acetobutylicum during cyclic potential
scanning from 0 mV to –1200 mV. Upper and lower volt-
ammetric peaks for dissolved NR were not altered but for
immobilized NR were shifted upward and rightward, re-
spectively, by addition of CO2. Other upper peak (bold ar-
row mark) also was shifted upward and rightwar d by addi-
tion of CO2.
Metabolites generated by C. acetobutylicum in different
cultivation conditions were quantitatively and qualita-
tively analyzed by a specially trained expert, and found
to be acetate, propionate, and butyrate as shown in Fig-
ure 3, as expected. This analytical process is absolutely
Table 1. Growth and metabolite production of C. acetobutylicum cultivated in complex medium (CM) and defined medium
(DM) under 2 atm of N2 atmosphere, 2 atm of H2 atmosphere, and electrochemical reduction condition (ER) for 5 days.
Metabolites (mM)
Cultivation conditions Growth at OD660 (initial-final)
Acetic acid Propionic acid Butyric acid
DM-N2 0.08 - 0.06 0.6 ± 0.04 0.1 ± 0.01 0.5 ± 0.02
DM-ER 0.08 - 0.36 9.2 ± 0.2 0.8 ± 0.03 6.4 ± 0.3
DM-H2 0.08 - 0.38 9.8 ± 0.3 0.9 ± 0.02 6.0 ± 0.3
CM-N2 0.08 - 1.28 35.8 ± 1.4 2.8 ± 0.1 21.6 ± 1.1
CM-ER 0.08 - 1.06 19.6 ± 0.8 4.8 ± 0.2 38.4 ± 0.9
CM-H2 0.08 - 1.21 24.4 ± 0.9 4.2 ± 0.2 36.6 ± 1.3
Figure 3. Mass spectrometer profiles of three major peaks detected in gas chromatography of volatiles contained in the cul-
ture fluid of C. acetobutylicum.
required because some metabolic intermediates derived
from amino acids (yeast extract) may be produced by
bacterial cells cultivated in the DM-H2 and DM-ER con-
3.4. Analysis of Genes Induced by
Electrochemical Reducing Power
Significant genes (higher than twice the signal intensity)
commonly expressed in C. acetobutylicum that was elec-
trochemically cultivated in the defined medium under
CO2-atmosphere and in the complex medium under
N2-atmosphere numbered in 318. All of the fundamental
genes related to CO2-fixation were not detected; however,
the genes of encoding the enzymes catalyzing acetyl-
CoA synthesis from CO2, ATP synthesis, and butyrate
production were quantitatively analyzed and compared as
shown in Table 2. The genes encoding the enzymes
catalyzing acetyl-CoA generation from CO2 and ATP
synthesis in pathway from acetyl-CoA to acetate were
more expressed but those catalyzing NADH regeneration
coupled to oxidation of substrates (metabolic intermedi-
ates) and butyric acid production were less expressed in
C. acetobuylicum cultivated in DM-ER condition than
CM-N2condition (Table 1).
4. Discussion
Electron transfer from electrode to bacterial cells can be porarily function in proportion to physiological activity
generated by the simultaneous contact of an electron me-
diator with both electrode and bacterial cells. Contact of
an electrode with the electron mediator (NR) immobi-
lized in bacterial cell or contact of bacterial cells with the
electron mediator immobilized in an electrode is a unique
way to induce electron transfer between bacterial cells
and electrode [17-19]. Patterns of cyclic voltammetry of
NR immobilized in bacterial cells were identical to those
of NR dissolved in reactant in the condition uncoupled to
the external redox reaction, because the redox reaction is
proportional to the concentration of NR contacted stably
with the electrode (Figure 2). Some NRs immobilized in
C. acetobutylicum are electrochemically reduced and
biochemically oxidized coupled to NADH regeneration
[6]. This electrochemical and biochemical coupling re-
dox reaction of NR and NAD+ may be continuously re-
peated in this condition with both electron donor and
acceptor. The electrode may be an electron donor and
CO2 may be an electron acceptor in the cyclic voltam-
metry for the modified C. acetobutylicum with NR, con-
sidering that addition of CO2 induced electron transfer
from electrode to bacterial cells via NR immobilized in
bacterial cells or other bacterial electron carrier (upper
voltammetric peaks in Figure 2) can be quantitatively
analyzed based on the increase of current (electron num-
ber) and variation of redox potential (electron-driving
force). The NR immobilized in bacterial cells can tem-
Copyright © 2012 SciRes. AiM
B. Y. JEON ET AL. 337
Table 2. Comparison of genes expressed in C. acetobutylicu
saturated defined medium and cultivated with glucose in the c
m cultivated with electrochemical reducing power in the CO2-
omplex medium.
Signal intensity for specific genes expressed in C. acetobutylicum
Ratio of A/B Gene products (Functions) Electrochemically
cultivated (A)
Cultivated in
complex medium (B)
4.87 Carbon monoxide n from CO2) 7764 dehydrogenase (CO formatio1595
4.71 Biotin-acetyl-CoA-carboxylase (Malonyl-CoA production) 2846 604
31.7 Formyl-H2folate synthase (Methyl-formation from CO2) 317 10
2.89 Formyl-H2folate cyclohydrolase (Methyl-formation from CO2) 772 267
244.8 Phosphl-CoA)
3.24 Putative
methyltransferase (Methyl-formation from CO2) 13,908
otransacetylase (Formation of acetyl-Pi from acety2448 10
212.4 Acetate kinase (Acetate production coupled to ATP synthesis) 2124 10
0.74 NAD-dependent dehydrogenase (NADH regeneration
coupled to substrate oxidation) 1856 524
0.21 Acetyl-CoA acetyltransferase (Butyric acid production) 4179 19,492
of tcte
zed in the graphite felt cathode can semipermanently
de, the electron transfer from electrode to bac-
fer from elec-
coupled to Ndation in C. acetum grown
in glucose. Accordingly, higher production of butyric
O fixa-
he immobi-
he baria modified with NR; however, NR immobi-
function as long as it is contacting with intact cells of
In the electrochemical bioreactor equipped with the
rial cells can’t be quantitatively analyzed because the
number of electrons (current) transferred from power
supply to NR-cathode is not proportional to the meta-
bolic reduction reaction catalyzed by bacterial cells, and
–2 V of electrode potential charged to the bioreactor was
stronger than the redox potential of NR. This can gener-
ate electron-driving force to transfer electrons via NR
immobilized in electrode to bacterial cells but may in-
duce electrochemical reduction of medium ingredients
and other organic compounds. The –2 V of electrode
potential may be too strong to induce the electrochemical
reduction of NR (–0.325 V vs. NHE) but is required to
induce electron transfer from electrode to bacterial cells
through the electron barrier (cytoplasmic membrane)
because electric resistance may be generated by reactor
membrane between anolyte and catholyte, connecting
error between NR and bacterial cells, and structural
mismatch between NR and NAD+ [20].
It is unquestionable that the NR immobilized in graph-
ite felt electrode mediated electron trans
de to bacterial cells and catalyzed regeneration of
biochemical reducing power on the basis of acetic and
butyric acid production from CO2 and production of
more butyric acid than acetic acid from glucose. This
result is very similar to that obtained from C. acetobu-
tylicum culture using H2 and CO2 as an electron donor
and acceptor. Metabolic production of acetic acid is cou-
pled to NADH regeneration but that of butyric acid is
acid than acetic acid by C. acetobutylicum cultivated in
CM-ER and CM-H2 is another clue that H2 and electro-
chemically reduced NR may be an additional reducing
power to increase ratio of NADH/NAD+ [21].
Metabolic conversion of CO2 to metabolites is coupled
to oxidation of biochemical reducing power regenerated
by the electrochemically reduced NR or H2; however, the
metabolic pathways related to the autotrophic C
ADH oxiobuylic
n can be assumed only by the metabolites produced by
C. acetobutylicum cultivated in different media and un-
der different conditions. Microarray analysis of mRNA is
effective to analyze variations of metabolic pathway and
free energy production related to autotrophic CO2fixation
or heterotrophic growth. Practically, the specific genes
related to the CO2 fixation and energy metabolism ex-
pressed in C. acetobutylicum electrochemically or con-
ventionally cultivated are a clue that the metabolic con-
version of CO2 to metabolites in coupling with the free
energy production and redox reaction of reducing power
may be generated by the electrochemical reducing power.
Theoretically, –2 V of electricity charged to the NR-
cathode located in culture medium may induce H2 gen-
eration by electrolysis of H2O. However, H2 was not de-
tected in the electrochemical bioreactor even by precision
analysis. Accordingly, the NR-cathode may directly
transfer electrons from electrode to intact cells of C.
acetobutylicum and induce catalyzing of NADH regen-
eration in metabolism of C. acetobutylicum.
5. Conclusion
The electrochemical redox reaction of NR, the catalytic
function of NR for NADH regeneration, and t
Copyright © 2012 SciRes. AiM
lization technique of NR in the electrode permit C. ace-
& Renewabl
ergy of the Korea Institute of Energy Technology
) grant funded by the Korea
. Jungermann and K. Decker, “Energy Con-
servation in Chemotrophic Anaerobic Bacteria,”
riological Rev, pp. 100-180.
, No. 2, 1979, pp. 177-182.
tobutylicum KCTC1037 to grow and produce metabolites
using electrochemical reducing power. Mixed acid fer-
mentation bacteria produced the relatively reduced me-
tabolite (butyrate) or oxidized metabolite (acetate) de-
pending on balance of NADH/NAD+. In autotrophic mi-
crobes, CO2 can be reduced to CO by catalysis of carbon
monoxide dehydrogenase in coupling with oxidation of
biochemical reducing power (NADH or NADPH). Prac-
tically, C. acetobutylicum produced more butyrate than
acetate from glucose and more acetate than butyrate from
CO2, reasonable on the basis of metabolic pathway for
ATP regenerations. C. acetobutylicum cultivated hetero-
trophically with glucose synthesizes ATP by substrate-
level phosphorylation and regenerates NADH in both
glycolysis and pathway from pyruvate to acetate but that
cultivated autotrophically with electrochemical reducing
power and CO2 synthesizes ATP in the pathway from
acetyl-CoA to acetate and regenerates NADH coupled to
electrochemical redox reaction of NR.
6. Acknowledgements
This work was supported by the New e En-
luation and Planning (KETEP
governmental Ministry of Knowledge Economy (2012-
[1] R. Thauer, KBacte-
iew, Vol. 41, No. 1, 1977
[2] A. M. Blackmer, J. M. Bremner and E. L. Schmidt, “Pro-
duction of Nitrous Oxide by Ammonia-Oxidizing Che-
moautotrophic Microorganisms in Soil,” Applied and En-
vironmental Microbiology, Vol. 40, No. 6, 1980, pp.
[3] E. Siefert and N. Pfennig, “Chemoautotrophic Growth of
Rhodopesudomonas Species with Hydrogen and Chemo-
trophic Utilization of Methanol and Formate,” Archives of
Microbiology, Vol. 122
[4] C. Castelle, M. Guiral, G. Malarte, F. Ledgham, G. Leroy,
M. Brugna and M.-T. Giudici-Orticon, “A New Iron-Oxi-
dizing/O2-Reducing Supercomplex Spanning Both
and Outer Membranes, Is
olated from the Extreme Aci-
dophile Acidithiobacillus ferrooxidans,” Journal of Bio-
logical Chemistry, Vol. 283, No. 38, 2008, pp. 25803-
25811. doi:10.1074/jbc.M802496200
[5] A. Elbehti, G. Brasseur and D. Lemesle-Meunier, “First
Evidence for Existence of an Uphill Electron Transfer
through the bc1 and NADH-Q Oxidoreductase Complexes
of the Acidophilic Obligate Chemoautotrophic Ferrous
Ion-Oxidizing Bacterium Thibacillus ferroxidans,” Jour-
nal of Bacteriology, Vol. 182, No. 12, 2000, pp. 3602-
3606. doi:10.1128/JB.182.12.3602-3606.2000
[6] D. H. Park and J. G. Zeikus, “Utilization of Electrically
Reduced Neutral Red by Actinobacillus succinogenes:
Physiological Function of Neutral Red in Membrane-
Driven Fumarate Reduction and Energy Conservation,”
Journal of Bacteriology, Vol. 181, No. 8, pp. 2403-2410.
[7] A. A. Karyakin, O. A. Bobrova and E. E. Karyakina,
“Electroreduction of NAD+ to Enzymatically Active
NADH at Poly(Neutral Red) Modified Electrodes,”
Journal of Electroanalytical Chemistry, Vol. 399, No. 1-2,
1995, pp. 179-184. doi:10.1016/0022-0728(95)04300-4
[8] M. Hügler, C. Menendez, H. Schägger and G. Fuchs,
“Malonyl-Coenzyme A Reductase from Chloroflexus
aurantiacus, a Key Enzyme of the 3-Hydroxypropionate
Cycle for Autotrophic CO2 Fixation,” Journal of Bacteri-
ology, Vol. 184, No. 9, 2002, pp. 2404-2410.
[9] H. Buschhhorn, P. Dürre and G. Gottschalk, “Production
and Utilization of Ethanol by the Homoacetogen Aceto-
bacterium woodii,” Applied and Environment
ology, Vol. 55, No. 7, 1989, pp. 1835-18
al Microbi-
[10] H. Zhang, M. A. Bruns and B. E. Logan, “Biological
Hydrogen Production by Clostridium acetobutylicum in
an Unsaturated Flow Reactor,” Water Research, Vol. 40,
No. 4, 2006, pp. 728-734.
[11] J. Zhang, J. Sun, X. Zhang, Y. Zhao and S. Zhang, “The
Recent Development of CO2 Fixation and Conversion by
Ionic Liquid,” Greenhouse Gases
ogy, Vol. 1, No. 2, 2011, pp. 142-1
: Science and Technol-
[12] B. Wang, Y. Li, N. Wu and C. Q. Lan, “CO2 Bio-Mitiga-
tion Using Microalgae,” Applied Microbiology and Bio-
technology, Vol. 79, No. 5, 2008, pp. 707-718.
[13] J. E. Funk, “Thermochemical Hydrogen Production: Past
and Present,” International Journal of Hydrogen Energy,
Vol. 26, No. 3, 2001, pp. 185-190.
rnal of Hydrogen
[14] A. Steinfeld, “Solar Hydrogen Production via a Two-Step
Water-Splitting Thermochemical Cycle Based on Zn/ZnO
Redox Reactions,” International Jou
Energy, Vol. 27, 2002, pp. 611-619.
[15] B. Y. Jeon, I. L. Jung and D. H. Park, “Enrichment and
Isolation of CO2-Fixing Bacteria with Electrochemical
Reducing Power as a Sole Energy S
Environmental Protection, Vol. 3, 2012
ource,” Journal of
, pp. 55-60.
[16] B. Y. Jeon, I. L. Jung and D. H. Park, “Enrichment of
CO2-Fixing Bacteria in Cylinder-Type Electrochemical
Bioreactor with Built-In Anode Compartment,” Jo
of Microbiology and Biotechnurnal
ology, Vol. 21, No. 6, 2011,
8, 1991, pp. 11445-11450.
pp. 590-598.
[17] C. J. Kay, L. P. Solomonson and M. J. Barber, “Electro-
chemical and Kinetic Analysis of Electron-Transfer Re-
actions of Chlorella Nitrate Reductase,” Biochemistry,
Vol. 30, No. 4
Copyright © 2012 SciRes. AiM
Copyright © 2012 SciRes. AiM
Methanol Elec-
Solid State Electrochemistry
[18] X. Zhong, J. Chen, B. Liu, Y. Xu and Y. Kuang, “Neutral
Red as Electron Transfer Mediator Enhanced Electro-
catalytic Activity of Platinum Catalyst for
tro-Oxidation,” Journal of,
Vol. 11, No. 4, 2007, pp. 463-468.
[19] L. Huang, J. M. Regan and X. Quan, “Electron Transfer
Mechanisms, New Applications, and performance of Bio-
cathode Microbial Fuel Cells,” Bio
Vol. 102, 2011, pp. 316-323.
resource Technology,
[20] G. Reguera, K. D. McCarthy, T. Meh
Tuominen and D. R. Lovley, “E
ta, J. S. Nicoll, M. T.
xtracellular Electron
Transfer via Microbial Nanowires,” Nature, Vol. 435,
2005, pp. 1098-1101. doi:10.1038/nature03661
[21] J. Song, Y. Kim, M. Lim, H. Lee, J. I. Lee and W. Shin,
“Microbes as Electrochemical CO Conversi
lysts,” ChemSusChem, Vol. 4, No. 5, 2011, pp. 587-590.
on Cata-