Bioresource Technology xxx (2016) xxxxxx

Contents lists available at ScienceDirect

Bioresource Technology
journal homepage: www.elsevier.com/locate/biortech

Hydrogen production in microbial reverse-electrodialysis electrolysis

cells using a substrate without buffer solution
Young-Hyun Song a, Syarif Hidayat a, Han-Ki Kim a,b, Joo-Yang Park a,
a
b

Department of Civil and Environmental Engineering, Hanyang University, 222 Wangsimni-ro, Seongdong-gu, Seoul 133-791, Republic of Korea
Jeju Global Research Center, Korea Institute of Energy Research, 200, Haemajihaean-ro, Gujwa-eup, Jeju Special Self-Govenrning Province 695-971, Republic of Korea

h i g h l i g h t s

Using a substrate without buffer solution in MRECs achieved hydrogen production.

Increasing in anolyte HRT and OLR increased cell current with stable anode potential.
3

Hydrogen production rate was 0.61 m -H2/m -Van/d (yield = 0.51 mol-H2/mole-COD).

Efficiencies of Coulombic and COD removal were 41% and 81% (rate: 1.55 g-COD/L/d).

a r t i c l e

i n f o

Article history:
Received 26 November 2015
Received in revised form 1 February 2016
Accepted 5 February 2016
Available online xxxx
Keywords:
Microbial reverse-electrodialysis
electrolysis cell
Hydrogen
Phosphate buffer solution
Sustainable energy
Exoelectrogen

a b s t r a c t
The aim of this work was to use substrate without buffer solution in a microbial reverse-electrodialysis
electrolysis cell (MREC) for hydrogen production under continuous flow condition (10 cell pairs of RED
stacks, HRT = 5, 7.5, and 15 h). Decreasing in the HRT (increasing in the organic matter) made cell current
stable and increased. Hydrogen gas was produced at a rate of 0.61 m3-H2/m3-Van/d in H-MREC, with a
COD removal efficiency of 81% (1.55 g/L/d) and a Coulombic efficiency of 41%. This MREC system without
buffer solution could successfully produce hydrogen gas at a consistent rate.
2016 Elsevier Ltd. All rights reserved.

1. Introduction
The world energy consumption rate is projected to grow with
both population and economic increases in the 21st century
(Lewis and Nocera, 2006). Oil prices are also increasing due to limited fossil energy and the continuous consumption of fossil fuel. As
a result, sustainable and renewable alternative energies are of
great interest and are being studied in a wide range of areas including solar-thermal, wind power, and hydrogen. Solar-thermal and
wind power energies, however, are non-continuous energy sources
that can only produce power with sun and wind, respectively. In
contrast, hydrogen can be produced from inexhaustible water
sources, and use in transportation burns cleanly compared with
traditional fuel sources. Thus, hydrogen has been widely discussed

Corresponding author. Tel.: +82 2 2220 0411.

E-mail address: jooypark@hanyang.ac.kr (J.-Y. Park).

in the context of energy as a possible future alternative energy

carrier.
Hydrogen gas as sustainable energy from an inexhaustible
source is important for avoiding the environmental problems associated with energy depletion from the use of fossil fuels (Ho et al.,
2012). Although water electrolysis is a simple and reliable method
for generating high-purity hydrogen, it requires energy input to
sustain the process.
MREC can electrochemically generate hydrogen gas at the cathode from the current produced from exoelectrogenic bacteria at
the anode; this oxidizes organic matter and can transfer electrons
to the anode. However, the electrical power (0.30 V using acetate; pH 7) from exoelectrogens at the anode is not enough to split
water (>1.2 V) for hydrogen production. Thus, an additional voltage
is required to overcome the thermodynamic threshold (Nam et al.,
2012). In order to cover the need for electrical energy, reverse electrodialysis (RED) stacks, which provide electrical energy from
salinity gradient energy (theoretically 0.10.2 V per membrane
pair based on the open circuit potential), are placed between the

http://dx.doi.org/10.1016/j.biortech.2016.02.021
0960-8524/ 2016 Elsevier Ltd. All rights reserved.

Please cite this article in press as: Song, Y.-H., et al. Hydrogen production in microbial reverse-electrodialysis electrolysis cells using a substrate without
buffer solution. Bioresour. Technol. (2016), http://dx.doi.org/10.1016/j.biortech.2016.02.021

Y.-H. Song et al. / Bioresource Technology xxx (2016) xxxxxx

anode and cathode chambers of an MREC (Veerman et al., 2009;

Dlugolecki et al., 2009).
In many studies related to energy production, highconcentration buffer solutions are used to maintain a neutral pH
condition for exoelectrogens and to increase electrolyte conductivity. Examples include microbial fuel cells (MFCs), microbial electrolysis cells (MECs), and MRECs (Zhang et al., 2015; Ambler and
Logan, 2011; Luo et al., 2013). However, the addition of buffer
solution is expensive for wastewater treatment applications.
Phosphate buffer solution (PBS) in particular can contribute to
eutrophication as the effluents are discharged without phosphate
removal.
In this study, we demonstrated the possibility of using a
substrate without buffer solution on an MREC for hydrogen gas
production. The experiments were conducted under continuous
flow for the anode, and in fed-batch mode for the cathode. The anolyte was complied with only a ratio of 300:5:1 = COD:N:P for
anaerobic conditions. The RED stack had thin flow channels to
improve the driving force in comparison with previous MRECs
(Luo et al., 2013; Kim and Logan, 2011) using NaCl as a highconcentration (HC) solution. Tests were initially conducted to
evaluate the effect of the anolyte HRT on current generation using
substrates with PBS. Following these tests, the substrate without
buffer solution was used in the MRECs. Using a substrate without
buffer solution has not been investigated for hydrogen production
in an MREC, and the performance of this MREC was discussed using
hydrogen production, yield, Coulombic efficiency, and COD
removal rate.
2. Methods
2.1. MREC setup
The rectangular parallelepiped acrylic reactor consisted of
anode and cathode chambers with a working volume of 36 mL each
(cross section area: 12 cm2, 3 cm length) (Fig. 1). An inlet and outlet were drilled perpendicularly in the bottom and top on the wall
of the anode chamber for continuous flow. A pH probe was placed
in the anode chamber to continually monitor and control the pH of
the anolyte. A carbon fiber brush was used as the anode electrode
(2.5 cm diameter, 3 cm length; T700 SC-12000, Toray, Japan),
which was pretreated by heating in a furnace at 450 C for
30 min. A titanium mesh (width: 3.5 cm, length: 2 cm, thickness:

0.2 cm) plated with Pt (2 lm of thickness) was used as a cathode

electrode. A Tedlar bag was inserted in the top of the cathode
chamber for gas collection with a silicon stopper. Each chamber
was equipped with an Ag/AgCl reference electrode (RE-5B, BASi)
to measure the electrode potential.
A RED stack was located between the anode and cathode chambers that consisted of 10 pairs of high-concentration (HC) and
low-concentration (LC) cells made with 11 anion- and 10 cationexchange membranes (Selemion AMV and CMV, Asahi glass,
Japan). Silicon gaskets, which have a rectangular open section of
12 cm2 (3  4 cm) for flow, were placed between the anion and
cation exchange membranes with a nylon mesh spacer
(0.18 mm), and the empty bed volume of the stack was 4.3 mL.
HC and LC solutions flowed continuously through each cell, but
in opposing directions. Each solution was provided into the RED
stack at a fixed flow rate of 1.2 mL/min.
2.2. MREC operation
The anode was pre-acclimated with exoelectrogen in an
acetate-fed single air-cathode MFC (Song et al., 2015). The acclimated anode was transferred to the MREC when the MFC produced
the same stable maximum voltages repeatedly for three cycles.
The anolyte complied with a ratio of COD:N:P = 300:5:1 for
anaerobic microorganisms using CH3COONa, NH4Cl, and NaH2PO4
2H2O (Tchobanoglous et al., 2004). 20 mM NaOH was used to
maintain the anolyte at a neutral pH. The catholyte was synthetic
HC solution degassed using sonication and a vacuum pump. The
synthetic HC solution was 600 mM NaCl, and the LC solution was
12 mM NaCl created using a salinity ratio of 50. The total volume
of the MREC was 36 (anode) + 36 (cathode) + 4.3 (RED stack)
= 76.3 mL. The anolyte was continuously supplied and stirred at
80 rpm with a magnetic bar to reduce concentration loss. The catholyte was replaced every fed-batch cycle at room temperature
when the current decreased to less than 1.0 mA. The anode and
cathode were connected to a fixed external resistance of 10 O.
2.3. Experimental measurements and calculations
The cell and electrode voltages were monitored and recorded
every 10 min with a voltage recorder (VR-71, T&D Corporation)
connected to a computer. Current was calculated based on the
measured cell voltage across the 10 O resistor using Ohms law.

Fig. 1. Schematic diagram of MREC process set-up showing flow paths through the RED stack.

Please cite this article in press as: Song, Y.-H., et al. Hydrogen production in microbial reverse-electrodialysis electrolysis cells using a substrate without
buffer solution. Bioresour. Technol. (2016), http://dx.doi.org/10.1016/j.biortech.2016.02.021

Y.-H. Song et al. / Bioresource Technology xxx (2016) xxxxxx

Gas produced at the cathode was collected and analyzed using

gas chromatographs (YL6500, YoungLin, Ins.) with a thermal conductivity detector (TCD) and a flame ionization detector (FID) to
measure the concentration of H2, N2, O2, CO2, and CH4. Argon
was used as a carrier gas. The volume of hydrogen produced was
measured using the water displacement method. All samples were
filtered with a 0.2 lm syringe before analysis. The total chemical
oxygen demand (COD) of the anolyte was measured by a standard
method using a Hach spectrophotometer (DR-2800, Hach Co.). The
pH and conductivity of the solutions were monitored with a pH
meter (Orion 720A+, Thermo) and conductivity meter (Orion three
star, Thermo), respectively.
Coulombic efficiency (rCE, %), yield (YH2 , mole-H2/mole-COD),
hydrogen production rate (Q, m3-H2/m3-Van/d), and overall hydrogen recovery ((rH2 , %) were used to evaluate the performance of
this MREC as previously described (Kim and Logan, 2011). The
hydrogen production rate was normalized to the volume of the
anolyte. The overall hydrogen recovery was determined by
r H2 rCE  r cat .

and 0.21 to 0.47 V, respectively. The electrode potentials of

the cathode were relatively constant for all HRTs. The effluent
COD of the anolyte was 0.040 0.005 g/L (average, n = 3). The variability of the anode potential is likely due to the low COD concentration of the anolyte. It has been demonstrated in MFC studies
that COD degradation occurs more rapidly in a higher current density, described by first-order kinetics. Furthermore, current generation was dropped when the COD concentration decreased to less
than 0.1 g/L (Ren et al., 2014; Zhang et al., 2015). Consequently,
volumes of hydrogen gas were produced in the MREC 34 (HRT of
15 h), 36 (HRT of 7.5 h), and 50 mL (HRT of 5 h), respectively.
Therefore, to generate sufficiently high current for the production
of hydrogen gas, a sufficient concentration of COD for exoelectrogens at the anode is needed. Over 92% of the produced gas was
hydrogen, with the remainder consisting of oxygen and nitrogen
in all reactors. CO2 and CH4 were not detected. The use of a RED
stack between anode and cathode chambers could stop the
methane generation at the cathode by blocking hydrogenotrophic
methanogens consuming hydrogen (Chae et al., 2010; Nam et al.,
2011b).

3. Results and discussion

3.1. Effect of anolyte HRT on current generation using a substrate with
PBS
In order to evaluate the performance of this MREC, this experiment was conducted using 1.0 g/L sodium acetate with PBS as an
anolyte in accordance with predetermined HRTs (HRT = 15, 7.5
and 5). Currents generated in the MREC varied depending on the
HRT under the anolyte continuous flow condition (Fig. 2). At an
HRT of 15 and 7.5 h (organic loading rate, OLR = 1.07 and 2.14 g
COD/L/d), cell currents showed unsteady form at 2.65.6 (3.0
6.4 A/m2, normalized by the surface area of the cathode) and
3.96.2 mA (4.57.1 A/m2) as the anode potential generation was
unstable at 0.02 to 0.38 V and 0.14 to 0.39 V. However, at
an HRT of 7.5 h, the overall current was increased and the anode
potential was decreased. Further decrease in the anolyte HRT to
5 h (OLR = 3.22 g COD/L/d) rendered both the cell current and
anode potential relatively stable at 3.96.9 mA (4.57.9 A/m2)

Fig. 2. Cell current (left axis) and anode and cathode potentials (right axis) of the
MREC fed acetate with PBS depending on HRTs of anolyte (HRT = 15, 7.5 and 5 h).

3.2. Current generation and hydrogen production using a substrate

without buffer solution
Various types of buffer solutions including phosphate, bicarbonate, MES (2-[N-morpholino] ethane sulfonate), and HEPES
(4-(2-hydroxyethly)-1-piperazineethanesulfonic acid), have been
used in many studies using electrochemically-active bacteria
(EAB) to increase the electrolyte conductivity and maintain a neutral pH condition (Nam et al., 2010; Ambler and Logan, 2011;
Watson et al., 2015). However, the immoderate addition of buffer
solution can contribute to eutrophication, especially PBS, and is
expensive due to further treatments required before disposal. Thus,
in order to evaluate the possibility of using a substrate without
buffer solution in MRECs for hydrogen production, MRECs were
operated using 1.0 (L-MREC) and 2.2 g/L (H-MREC) sodium acetate
under the continuous flow condition for anolyte during 3 cycles at
fixed HRT of 15 h. The substrate complied with the ratio of COD:N:
P = 300:5:1. The pH of the anolyte was maintained between 5.5
and 8.5 by the continuous addition of a NaOH solution as the pH
decreased due to the hydrogen ions released from organic matter
oxidation.
After increasing the concentration of sodium acetate on the
substrate from 1.0 to 2.2 g/L (0.721.6 g COD/L), the current generated by the H-MREC was 3.84.7 mA (4.35.4 A/m2), and was
higher than that of the L-MREC (0.81.8 mA or 0.92.1 A/m2) as
the anode potential decreased from 0.110.30 V to 0.48 to
0.36 V (Figs. 3 and 4). The effluent COD of L-MREC was 0.07 g/L
(n = 3), while the effluent COD of H-MREC was much higher
(0.31 g/L). The large changes in current and anode potentials
between H- and L-MRECs were due to the low COD concentration
in the anode chamber; therefore, the increase in organic matter to
1.6 g COD/L at an HRT of 15 h (OLR = 1.152.56 g COD/L/d) made
the anode potential negative, 0.45 V. Consequently, 25 and
10 mL of hydrogen gases of H- and L-MRECs were produced,
respectively, which is proportionate to the current generation.
Current generation of the H-MREC was relatively high when compared with those of previous MREC studies under various conditions (number of RED stacks, continuous and fed-batch modes),
resulting in 2.56.0 A/m2 and 5 mA (Watson et al., 2015; Kim
and Logan, 2011; Nam et al., 2012; Luo et al., 2013). The use of a
substrate without buffer solution in this MREC achieved relatively
high current generation compared to those from previous MRECs,
which is a significant advantage and will aid further research and
applications for wastewater treatment in terms of cost reduction.

Please cite this article in press as: Song, Y.-H., et al. Hydrogen production in microbial reverse-electrodialysis electrolysis cells using a substrate without
buffer solution. Bioresour. Technol. (2016), http://dx.doi.org/10.1016/j.biortech.2016.02.021

Y.-H. Song et al. / Bioresource Technology xxx (2016) xxxxxx

Fig. 3. Cell current (left axis) and anode and cathode potentials (right axis) of the
MREC fed 1.0 g/L sodium acetate without PBS under the continuous flow of anolyte
(HRT = 15 h).

from organic matter oxidation and the absence of a buffer solution

even with pH control. The low pH inhibits significantly exoelectrogenic activity (Mehanna et al., 2010). Nonetheless, the substrate
based on COD was completely removed (81% at the H-MREC to
90% at the L-MREC). The catholyte pH greatly increased from 5.4
to 12.8 due to the consumption of hydrogen ions at the cathode.
The rate of COD removal increased with organic matter,
although the COD removal efficiency of the L-MREC was higher
than that of the H-MREC, which is likely due to the amount of
organic matter injected into the reactor. With low organic matter
(L-MREC), the COD removal rate was 0.78 g/L/d (removal efficiency: 90%), and increased to 1.55 g/L/d (removal efficiency:
81%) with the addition of organic matter (H-MREC). It follows that
the current increases with organic matter. The Coulombic efficiencies were obtained 24% at the L-MREC and 41% at the H-MREC.
When the organic matter was increased, Coulombic efficiency also
increased.
The hydrogen gas production rate and Coulombic efficiency
were relatively low compared with those of previous MRECs,
although the current generated was high, which is likely due to
the absence of buffer solution in the anolyte. The lack of buffer
solution contributed to the instability of the pH and low conductivity (2220 lS/cm) in the anolyte. The solution conductivity is one of
the most important factors in microbial electrochemical technologies (METs). Nam et al. (2010) has demonstrated that the various
power densities generated can be equalized by adjusting the conductivity of the solution, which results in a significant reduction
of internal resistance (ohmic loss). Thus, increasing the conductivity of anolyte will help improve the MREC performance for hydrogen production even in the absence of buffer solution.

4. Conclusions
The use of substrates without buffer solution in MRECs can
achieve consistent hydrogen production. The increase in organic
matter (the decrease in anolyte HRT) resulted in increased cell current and stable anode potential. The hydrogen production rate
reached 0.61 m3-H2/m3-Van/d and a Coulombic efficiency of 41%
with a COD removal of 81%, although the anolyte conductivity
was significantly lower than that of existing MRECs with buffer
solution. Consequently, this study elucidates the possibility of
using substrates without buffer solution in MRECs for hydrogen
production, which will be an important step for further research
or application to wastewater treatment.

Fig. 4. Cell current (left axis) and anode and cathode potentials (right axis) of the
MREC fed 2.2 g/L sodium acetate without PBS under the continuous flow of anolyte
(HRT = 15 h).

Acknowledgement
This research was generously supported by the National
Research Foundation of Korea (NRF) (No. 2013R1A2A2A03068675).

3.3. Performance of MREC using substrates without buffer solution

References
Hydrogen gas was successfully produced in the MREC using a
substrate without any buffer solution and no additional external
power supply. The hydrogen yield increased from 0.37 to
0.51 mol-H2/mole-COD with an increase in organic matter. The
maximum hydrogen rates were 0.61 m3-H2/m3-Van/d in H-MREC
and 0.22 m3-H2/m3-Van/d in L-MREC. The hydrogen production
achieved in this system (H-MREC) was similar to or less than existing MECs (0.531.5 m3-H2/m3-Van/d) with an externally-applied
voltage and MRECs (0.91.6 m3-H2/m3-Van/d) with buffer solution
under similar conditions (Cheng and Logan, 2007; Hu et al.,
2008; Nam and Logan, 2011a; Kim and Logan, 2011; Watson
et al., 2015; Luo et al., 2013; Nam et al., 2012). The pH of the anolyte ranged from 5.5 to 8.5 due to consistent hydrogen ion release

Ambler, J.R., Logan, B.E., 2011. Evaluation of stainless steel cathode and a
bicarbonate buffer for hydrogen production in microbial electrolysis cells
using a new method for measuring gas production. Int. J. Hydrogen Energy 36,
160166.
Chae, K.J., Choi, M.J., Kim, K.Y., Ajayi, F.F., Chang, I.S., Kim, I.S., 2010. Selective
inhibition of methanogens for the improvement of biohydrogen production in
microbial electrolysis cells. Int. J. Hydrogen Energy 35, 1337913386.
Cheng, S., Logan, B.E., 2007. Sustainable and efficient biohydrogen production via
electrohydrogenesis. PNAS 104 (47), 1887118873.
Dlugolecki, P., Gambier, A., Nijmeijer, K., Wessling, M., 2009. Practical potential of
reverse electrodialysis as process for sustainable energy generation. Environ.
Sci. Technol. 43, 68886894.
Ho, K.L., Lee, D.J., Su, A., Chang, J.S., 2012. Biohydrogen from lignocellulosic
feedstock via one-step process. Int. J. Hydrogen Energy 36, 160166.
Hu, H., Fan, Y., Liu, H., 2008. Hydrogen production using single-chamber
membrane-free microbial electrolysis cells. Water Res. 42, 41724178.

Please cite this article in press as: Song, Y.-H., et al. Hydrogen production in microbial reverse-electrodialysis electrolysis cells using a substrate without
buffer solution. Bioresour. Technol. (2016), http://dx.doi.org/10.1016/j.biortech.2016.02.021

Y.-H. Song et al. / Bioresource Technology xxx (2016) xxxxxx

Kim, Y., Logan, B.E., 2011. Hydrogen production from inexhaustible supplies of fresh
and salt water using microbial reverse-electrodialysis electrolysis cells. PANS
108 (39), 1617616181.
Lewis, N.S., Nocera, D.G., 2006. Powering the planet: chemical challenges in solar
energy utilization. PANS 103 (43), 1572915735.
Luo, X., Nam, J.Y., Zhang, F., Zhang, X., Liang, P., Huang, X., Logan, B.E., 2013.
Optimization of membrane stack configuration for efficient hydrogen
production in microbial reverse-electrodialysis electrolysis cells coupled with
thermolytic solutions. Bioresour. Technol. 140, 399405.
Mehanna, M., Kiely, P.D., Call, D.C., Logan, B.E., 2010. Microbial electrodialysis cell
for simultaneous water desalination and hydrogen gas production. Environ. Sci.
Technol. 44, 95789583.
Nam, J.Y., Logan, B.E., 2011. Enhanced hydrogen generation using a saline catholyte
in a two chamber microbial electrolysis cell. Int. J. Hydrogen Energy 36, 15105
15110.
Nam, J.H., Kim, H.W., Lim, K.H., Shin, H.S., Logan, B.E., 2010. Variation of power
generation at different buffer types and conductivities in single chamber
microbial fuel cells. Biosens. Bioelectron. 25, 11551159.
Nam, J.Y., Tokash, J.C., Logan, B.E., 2011. Comparison of microbial electrolysis cells
operated with added voltage or by setting the anode potential. Int. J. Hydrogen
Energy 36, 1055010556.

Nam, J.Y., Cusick, R.D., Kim, Y., Logan, B.E., 2012. Hydrogen generation in microbial
reverse-electrodialysis electrolysis cells using a heat-regenerated salt solution.
Environ. Sci. Technol. 46, 52405246.
Ren, L., Zhang, X., He, W., Logan, B.E., 2014. High current densities enable
exoelectrogens to outcompete aerobic heterotrophs for substrate. Environ. Sci.
Technol. 46, 52405246.
Song, Y.H., An, B.M., Shin, J.W., Park, J.Y., 2015. Ethanolamine degradation and
energy recovery using a single air-cathode microbial fuel cell with various
separators. Int. Biodeterior. Biodegrad. 102, 392397.
Tchobanoglous, G., Burton, F.L., Stensel, H.D., 2004. Wastewater Engineering
Treatment and Reuse, fourth ed. McGraw Hill, New York.
Veerman, J., Saakes, M., Metz, S.J., Harmsen, G.J., 2009. Reverse electrodialysis:
performance of a stack with 50 cells on the mixing of sea and river water. J.
Membr. Sci. 327, 136144.
Watson, V.J., Hatzell, M., Logan, B.E., 2015. Hydrogen production from continuous
flow, microbial reverse-electrodialysis electrolysis cells treating fermentation
wastewater. Bioresour. Technol. 195, 5156.
Zhang, X., He, W., Ren, L., Stager, J., Evans, P.J., Logan, B.E., 2015. COD removal
characteristics in air-cathode microbial fuel cells. Bioresour. Technol. 176,
2331.

Please cite this article in press as: Song, Y.-H., et al. Hydrogen production in microbial reverse-electrodialysis electrolysis cells using a substrate without
buffer solution. Bioresour. Technol. (2016), http://dx.doi.org/10.1016/j.biortech.2016.02.021