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Biohydrogen production in an outdoor panel photobioreactoron dark fermentation effluent of molasses

Sevler Gokce Avcioglu a, Ebru Ozgur a,*, Inci Eroglu a, Meral Yucel b, Ufuk Gunduz b

aDepartment of Chemical Engineering, Middle East Technical University, Ankara 06531, TurkeybDepartment of Biological Sciences, Middle East Technical University, Ankara 06531, Turkey

a r t i c l e i n f o

Article history:

Received 7 October 2010

Received in revised form

9 December 2010

Accepted 12 December 2010

Available online 26 January 2011

Keywords:

Photofermentation

Biohydrogen

Molasses

Photobioreactor

Global solar radiation

* Corresponding author. Tel.: þ90 312 210269E-mail address: ebruozgur@gmail.com (E

0360-3199/$ e see front matter Copyright ªdoi:10.1016/j.ijhydene.2010.12.046

a b s t r a c t

Hydrogen is regarded as an ideal energy carrier if it is produced from renewable resources

such as biomass. Sequential operation of dark and photofermentation allows a highly

efficient production of hydrogen from biomass, as maximal conversion of the energy in the

carbohydrates to hydrogen can be achieved. In this study photofermentative hydrogen

production was carried out in a solar panel photobioreactor by Rhodobacter capsulatus wild

type (DSM 1710) and Rhodobacter capsulatus hup� (YO3) strain on the molasses dark

fermentation effluents which were obtained using an extreme thermophile Caldicellusir-

uptor saccharolyticus in the dark fermentation step. Continuous hydrogen production on the

molasses dark fermentation effluents was achieved up to 55 days with R. capsulatus wild

type and 75 days with R. capsulatus hup� in outdoor conditions during summer 2009, in

Ankara Turkey. The maximum hydrogen yield obtained using R. capsulatus hup- was 78%

(of the theoretical maximum) and the maximum hydrogen productivity was 0.67 mmol H2/

Lc.h. The maximum hydrogen productivity and yield of the wild type strain on the

molasses dark fermentation effluents were 0.50 mmol H2/Lc.h and 50%, respectively. The

changes in climatic conditions, particularly daily global solar radiation, affected the

hydrogen productivity and yield.

Copyright ª 2011, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights

reserved.

1. Introduction renewable resources the most common methods to produce

Provision of the energy demand, depletion of the common

energy resources and the global climate change that emerged

with increasing fossil-based CO2 emission constitute the main

challenges ofmankind in this century. From this point of view,

hydrogenwill becomemore andmore important in the coming

decades. As the end product of hydrogen on combustion is

mainly water, hydrogen is invaluable as clean, renewable and

harmless energy carrier of the future [1]. Furthermore, it has

higher energy content (142 MJ/kg) compared to natural gas

(28 MJ/kg) [2].

There are several methods to produce hydrogen from

both renewable and non-renewable resources. Among non-

6; fax: þ90 312 2102600.. Ozgur).2011, Hydrogen Energy P

hydrogen are steam methane reforming, partial oxidation of

hydrocarbons, coal gasification and biomass gasification [3].

However, all non-renewable hydrogen production technologies

also increase CO2 emissions and face with the exhaustion of

resources. Obviously, hydrogen can be an ideal energy carrier if

it is produced from renewable resources. Hence, electrolysis

combined with photovoltaics, wind power, hydropower, and

biological hydrogen production from biomass as clean

hydrogen production technologies form an alternative to the

non-renewable.

Biological hydrogen is produced via direct photolysis by

using microalgae and cyanobacteria, photofermentation

throughphotosynthetic purple bacteria, anddark fermentation

ublications, LLC. Published by Elsevier Ltd. All rights reserved.

Table 1 e The composition of the molasses DFE.

Component Concentration Component Concentration

Ni (mM) 0.68 AA (mM) 65

Co (mM) 3.39 LA (mM) 5

Mn (mM) 12.38 FA (mM) 5

Fe (mM) 25.86 PA (mM) 0

Cu (mM) 0.29 BA (mM) 0

Zn (mM) 21.05 NHþ4 (mM) 1

Mo (mM) 5.92 Sucrose (mM) 2

Ca (mM) 0.61 Ethanol (mM) 5

Mg (mM) 5.26 TOC (mg/L) 1870

B (mM) 0.00 COD (mg/L) 6500

Na (mM) 239.24 TN (mg/L) 420

i n t e r n a t i o n a l j o u r n a l o f h y d r o g e n en e r g y 3 6 ( 2 0 1 1 ) 1 1 3 6 0e1 1 3 6 8 11361

processes with dark fermentative bacteria [4]. Two-step bio-

hydrogen production with dark fermentation in the first step

and photofermentation in the second step is a highly efficient

biohydrogen production process, as it allows for themaximum

conversion of substrates toH2. Indark fermentation, biomass is

converted to hydrogen, CO2 and organic acids by thermophilic

or mesophilic dark fermentative bacteria and in photo-

fermentation organic acids derived fromdark fermentation are

converted to more hydrogen by purple non-sulfur (PNS)

bacteria. Such an approach is being employed by the EU 6th

Framework Integrated Project: HYVOLUTION, which aims to

develop a 2-stage bioprocess for the cost effective production of

pure hydrogen from biomass, ranging from energy crops to

bioresidues from agro-industries [5].

Biomass sources used for biological hydrogen production

can be basically defined as organic matters from wood and

wood wastes and residues, crops, trees, plants, grasses,

aquatic plants and algae, agricultural and forestry wastes,

sludge, animal wastes and residues, organic waste materials

and municipal solid waste [6].

Sugar beet molasses is a by-product of the sugar industry

that is obtained as thick syrup during the crystallization of

sucrose. It contains a high amount of sucrose (ca. 50%), and is

rich in inorganic nitrogen, vitamins and salt, which may

support bacterial growth. It is described as a suitable feed-

stock for dark fermentation under mesophilic conditions

[7e10]. It is reported that sequential operation of thermophilic

DFE of molasses can be used as a substrate for the photo-

fermentative hydrogen production using different PNS

bacterial strains, in batch mode. The highest yield and

productivity is obtained with the uptake hydrogenase defi-

cient (hup-) mutant strain of Rhodobacter capsulatus [11].

The main aim of this study is to investigate the applica-

bility of molasses as feedstock on sequential dark and pho-

tofermentation process. To this purpose the parameters

affecting long term photobiological hydrogen production in

outdoor conditions were determined. A stable photo-

fermentative hydrogen production on the molasses DFE was

achieved for 75 days for R. capsulatus hup- and 55 days for the

wild type strain.

2. Experimental

2.1. Microorganism and media

Rhodobacter capsulatus hup- strain which is a mutant strain of

R. capsulatus MT1131 lacking uptake hydrogenase enzyme [12]

and R. capsulatus DSM 1710 were used in this study.

Molasses DFEswere delivered by DLO-FBR (WageningenUR

Food & Biobased Research Wageningen, The Netherlands).

They were obtained by continuous dark fermentation process

using thermophilic dark fermentative bacteria; Caldicellusir-

uptor saccharolyticus. Composition of the effluent is shown in

Table 1.

2.2. Photobioreactor

The photobioreactors used in this study had an internal cooler

made of Plexiglas sheets with a thickness of 10 mm. The

dimensions of the photobioreactors were 45 cm � 45 cm �2 cm. The working volume of the photobioreactors was 4 L.

The sketch of the experimental setup is given in Fig. 1. PVC

rigid sheet was used as a frame which connected the two

Plexiglas sheets together with stainless steel socket cap bolts

and nuts with plastic insert. Teflon gaskets and stainless steel

washers were used for sealing the bolt holes. Sponge rubber

cord was used as a sealing material between the Plexiglas

sheets and the PVC frame. There were ball valves two on the

top and two on one sidelong of the photobioreactor for

sampling and feeding purposes. Additionally, flexible PVC

tubing was used as cooling coil and it was stacked the Plexi-

glas sheet with suction pads. Output connections of the

cooling coil were made by push-in adaptors. The evolved gas

was collected with polyurethane tubing in a cylindrical gas

column made of Plexiglas.

2.3. Procedure

The photobioreactors were sterilized with 3% hydrogen

peroxide. The molasses DFEs were autoclaved at 121 �C for

20 min before processing. The photobioreactors were started

with 25% (by the volume of the reactor) freshly grown

bacteria (R. capsulatus hup-), 50% the molasses DFE and 25%

sodium carbonate buffer (pH 6.4) containing Fe-citrate and

Na2MoO4$2H2O. At the start-up, buffer, Fe-citrate and

Na2MoO4$2H2O concentrations in the reactor were adjusted

to 5 mM, 0.1 mM and 0.16 mM, respectively. No argon was

flushed into the photobioreactor as the reactor was com-

pletely filled [13]. The temperature of the photobioreactors

was recorded continuously with temperature probes con-

nected to a computer. During day time, the temperature was

kept below 35 �C in the photobioreactor with chilled water

(5 �C) circulating through cooling coils placed inside the

photobioreactors. The photobioreactors were operated in

a fed-batch mode with a daily feeding rate of 10% by the

volume of the reactor. The feeding media contained 2 times

diluted molasses DFE containing Na2CO3 buffer (5 mM, pH

6.4), Fe-citrate (0.1 mM) and Na2MoO4.2H2O (0.16 mM).

2.4. Analytical methods

Bacterial cell concentration, pH, temperature, daily global

solar radiation, organic acid consumption, hydrogen produc-

tion, total carbon, total nitrogen, chemical oxygen demand

Fig. 1 e A sketch of the experimental setup.

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and NH4þ were measured during the experiments. The bacte-

rial cell concentration was measured with Shimadzu UV-1201

spectrophotometer. An optical density of 1 at 660 nm corre-

sponds to a cell density of 0.47 g dry cell weight/Lc for R. cap-

sulatus hup� [12] and 0.54 g dry cell weight/Lc for R. capsulatus

DSM 1710 [14]. Total carbon, total nitrogen, chemical oxygen

demand and ammonia analysis were carried out spectro-

photometrically using Hach-Lange TOC, TNT, COD and

NH4eN kit (DR/2400 HacheLange Spectrophotometer,

Germany). The pH was measured with a pH-meter (Met-

tlereToledo); temperature was monitored with a temperature

probe (Elimko E680 Universal Data Loggers/Scanners, Turkey)

plunged into the photobioreactors. A cooler (Pnoso, PSS Series

Process Water Cooler, Turkey) was used to cool the chilled

water circulating through PVC tube inserted into the photo-

bioreactor (Fig. 1). The organic acid analyses were done via

high performance liquid chromatography (Shimadzu 10A

series) equipped with a UV detector (Shimadzu FCV-10AT)

with absorbance set at 210 nm and Alltech IOA-1000 organic

acid column (300 mm � 7.8 mm). H2SO4 (0.085 M) was used as

the mobile phase pumped at a flow rate of 0.4 ml/min using

a low gradient pump (Shimadzu LC-10AT). The oven temper-

ature was kept constant at 66 �C and 10 ml of samples were

analyzed. Produced gas was analyzed using thermal conduc-

tivity detector by gas chromatography (Agilent Technologies

6890N) using Supelco Carboxen 1010 column. The oven,

injector and detector temperatures were 140, 160 and 170 �C,respectively. Argon gas was used as a carrier gas at a flow rate

of 25 ml/min. Global total solar radiation data for Ankara

region related to the experimental period were taken from the

National Meteorology Institute of Turkey. All measurements

were done in the samples taken before feeding.

Molar hydrogen productivity QH2was estimated by

dividing cumulative molar hydrogen produced ðNH2 Þ by the

working volume of the photobioreactor (VPBR) and the total

illumination time (t) as shown in Eq. (1), below. Daily hydrogen

produced ðNH2 Þ was corrected for the pressure and tempera-

ture recorded at measuring day and time.

QH2¼ ��VH2

=22:4�� PH2

� �273=TH2

���ðVPBR � tÞ (1)

where, VH2is the volume of the evolved hydrogen; PH2

is the

pressure of the evolved hydrogen, which was assumed to be

equivalent to the barometric pressure, i.e. 0.9 atm in Ankara,

Turkey; TH2is the temperature of the evolved hydrogen; VPBR is

the working volume of the photobioreactor; t is the illumina-

tion time during the experimental period.

The molar yield (%YN) of the photofermentation process

was calculated as the percent ratio of the cumulative moles of

hydrogen which was produced ððNH2 ÞexpÞ to the theoretical

moles of hydrogen that can be produced from utilized organic

acids (acetic and lactic acid) ððNH2 ÞtheoÞ as given below;

%YN ¼ �NH2ðexpÞ=NH2ðtheoÞ�� 100 (2)

The stoichiometric equations for the calculation of the

theoretical moles of hydrogen by converting utilized acetic

and lactic acid are given below:

Acetic Acid : C2H4O2 þ 2H2O/4H2 þ 2CO2 (3)

Lactic Acid : C3H6O3 þ 3H2O/6H2 þ 3CO2 (4)

The moles of utilized acetic acid ðNutAA Þ and lactic acid

ðNutLA Þ were estimated as shown in Eqs. (5) and (6),

respectively.

NutAA ¼ CPBRAA� VPBR þ

n� CF �

Xni

CPBRAA

!� VF (5)

NutLA ¼ CPBRLA � VPBR þ n� CF �

Xni

CPBRLA

!� VF (6)

i n t e r n a t i o n a l j o u r n a l o f h y d r o g e n en e r g y 3 6 ( 2 0 1 1 ) 1 1 3 6 0e1 1 3 6 8 11363

where CPBRAAand CPBRLA

are the acetic and lactic acid concen-

trations in the photobioreactor at the end of batch phase, VPBR

is the working volume of the photobioreactor; CF is the acetic/

lactic acid concentration in the feeding media; VF is the

volume of the feeding media; n is the number of days that

acetic and lactic acid were fed; and CPBRAA and CPBRLA are the

concentrations of acetic and lactic acid that was removed

before feeding.

Acetate conversion efficiency (% hAA) for the continuous

operation was calculated by dividing the moles of acetic acid

utilized ðNutAA Þ to that was fed to the photobioreactor ðNinAAÞ

(Eq. (7)).

%hAA ¼ NutAA=NinAA� 100 (7)

3. Results and discussions

3.1. Growth and hydrogen production

Variations in the daily hydrogen production and cell concen-

tration in outdoor continuous photobioreactor run by R. capsu-

latus hup- are shown in Fig. 2. After the inoculation, thebiomass

concentration was 0.16 gdcw/Lc. The continuous feeding was

started on the 11th day where the biomass concentration

reached to 0.45 gdcw/Lc. The cell concentration in the photo-

bioreactor increased till 23th day then it stabilized around

0.9 gdcw/Lc. The hydrogen production started on the 5th day.

Daily hydrogenproduction varied from0.7 to 2.1 L/daybetween

11 and 40th days of the experiment. After 40th day daily

hydrogen production decreased and between 41th and 75th

days it varied from 0.2 to 1.0 L/day. Themaximumproductivity

(0.67 mmol H2/Lc.h) and molar yield (78%) were obtained

between 11 and 40th days. Hydrogen productivity decreased to

0.38 mmol H2/Lc.h and molar yield was 42% of the theoretical

maximum between 41th and 75th days. The produced gas was

90e95% H2, the rest being CO2. The overall acetate conversion

efficiency for continuous operation was 77%.

Fig. 2 e Growth and hydrogen production in outdoor continuous

DFE. Starting date of the experiment was 30.07.09. Feeding was

Fig. 3 illustrates variations in the daily hydrogen production

and biomass in outdoor continuous photobioreactor run by R.

capsulatus DSM 1710. The initial biomass concentration was

0.3 gdcw/Lc. The continuous feedingwas started on the 6th day

where the biomass concentration reached to 0.7 gdcw/Lc. The

cell concentration in thephotobioreactor increased to 1.0 gdcw/

Lc till 18th day then it gradually decreased to 0.7 gdcw/Lcbetween 18th and 49th day. After 49th day cell concentration

started to increase and reached to 0.9 gdcw/Lc at the end of the

experiment. The hydrogen production started on the 5th day.

The hydrogen productivity was 0.22 mmol H2/Lc.h and molar

yield was 10% that were obtained in the first 5 days. Hydrogen

productivity increased to 0.50 mmol H2/Lc.h and molar yield

was 50% of the theoretical maximum between 6th and 41st

days. After 41st day the hydrogen productivity and yield were

0.28mmolH2/Lc.h and 36%, respectively. The produced gaswas

90e95% H2, the rest being CO2. The overall acetate conversion

efficiency for continuous operation was also 77%.

Uptake hydrogenase is a membrane metalloprotein which

physiologically function as H2-uptake hydrogenase. In PNS

bacteria the uptake hydrogenases, mostly belonging to Class I

of [NiFe]hydrogenases, catalyze the conversion of molecular

hydrogen into proton and electron, whereby decreasing the

efficiency of hydrogen production. Deletion of the gene was

shown to improve hydrogen production in several PNS bacteria

including R. capsulatus, R. sphaeroides, Rhodospirillum rubrum

[12,19,20]. In this study, the uptake hydrogenase deficient

strain of R. capsulatus resulted in higher hydrogen productivity

and higher yield which is in agreement with the previous

results obtained in batch cultures on molasses DFE [15].

3.2. Organic acid consumption

In Fig. 4 organic acid concentrations in the photobioreactor

run by R. capsulatus hup- are depicted. The photobioreactor

was started to be fed with 10% daily feeding rate at the 11th

day. During the batch operation 30% of acetate that was

initially fed was consumed. After feeding started the acetate

photobioreactor run by R. capsulatus hup- on the molasses

started at 11th day.

Fig. 3 e Growth and hydrogen production in outdoor continuous photobioreactor run by R. capsulatus DSM 1710 on the

molasses DFE. Starting date of the experiment was 17.08.09. Feeding was started at 6th day.

i n t e rn a t i o n a l j o u r n a l o f h y d r o g e n en e r g y 3 6 ( 2 0 1 1 ) 1 1 3 6 0e1 1 3 6 811364

concentration in the photobioreactor continued to decrease

due to simultaneous hydrogen production and biomass

growth in following 14 days. The acetate concentration in the

photobioreactor continued to decrease even after the feeding

started. After 40th day, acetate accumulated in the photo-

bioreactor and at 75th day its concentration reached to 12mM.

Lactic acid was completely consumed throughout the exper-

iment. Propionic and butyric acid formation was negligible.

However, formic acidwas produced in a considerable amount.

It reached to its maximum value (28 mM) at 26th day and

gradually decreased to 16 mM at 75th day.

The variation in the organic acid concentrations in the

photobioreactor run by R. capsulatus DSM 1710 is illustrated in

Fig. 5. Theacetatewhichwas initially fed to thephotobioreactor

Fig. 4 e Organic acid consumption in outdoor continuous photo

Starting date of the experiment was 30.07.09. Feeding was star

was consumedby 93% in thebatch period. At the 6thday before

feeding started its concentration was 2.7 mM. After 16th day

acetate in the photobioreactor started to accumulate in the

photobioreactor.At theendof theexperiment its concentration

was 10 mM. Lactic acid was completely consumed throughout

the experiment. Propionic and butyric acid formation was

negligible. There was a considerable amount of formic acid

production in this photobioreactor, too. Its concentration

reached to 26 mM at the 19th day. After 25th day its concen-

tration decreased gradually and reached to 14mMat the end of

the experiment.

Differences in consumption of acetate in the batch phase

correlate well with biomass accumulation trends of both

bacteria. R. capsulatus hup-, which exhibited a longer lag phase

bioreactor runs by R. capsulatus hup- on the molasses DFE.

ted at 11th day.

Fig. 5 e Organic acid consumption in outdoor continuous photobioreactor runs by R. capsulatus DSM 1710 on the molasses

DFE. Starting date of the experiment was 17.08.09. Feeding was started at 6th day.

i n t e r n a t i o n a l j o u r n a l o f h y d r o g e n en e r g y 3 6 ( 2 0 1 1 ) 1 1 3 6 0e1 1 3 6 8 11365

of growth, consumed less acetate during batch operation,

while thewild type R. capsulatus, which grew faster, consumed

more acetate during the same period. During fed-batch oper-

ation, biomass was stabilized and acetate was consumed

mainly for hydrogen production. As hydrogen productivity

decreased towards the end of the operation, acetate accu-

mulated. Formate was produced simultaneously with the

increase in biomass, for both strains.

3.3. Impact of solar radiation on hydrogen production

The daily global solar radiation versus daily hydrogen

production in the photobioreactors R. capsulatus hup- and

R. capsulatusDSM 1710 are shown in Figs. 6 and 7, respectively.

Fig. 6 e Daily global solar radiation energy versus daily hydrogen

R. capsulatus hup- on the molasses DFE. Starting date of the exp

The daily global solar radiation energy decreased from 7000 to

1700 Wh/m2 during the experiments. A direct relation

between the hydrogen production and daily global solar

radiation energy was observed for both reactors. As the daily

global solar radiation decreases daily hydrogen production

also decreases.

3.4. Temperature variation in the photobioreactors

The temperature in the photobioreactors was controlled with

chilled water circulating through the internal coil. The photo-

bioreactor temperature was controlled around 35 �C by

running the cooler for 9 h in day-time in the photobioreactor

run by R. capsulatus hup- for 55 days and in the photobioreactor

production in outdoor continuous photobioreactor runs by

eriment was 30.07.09. Feeding was started at 11th day.

Fig. 7 e Daily global solar radiation energy versus daily hydrogen production in outdoor continuous photobioreactor runs by

R. capsulatus DSM 1710 on the molasses DFE. Starting date of the experiment was 17.08.09. Feeding was started at 6th day.

i n t e rn a t i o n a l j o u r n a l o f h y d r o g e n en e r g y 3 6 ( 2 0 1 1 ) 1 1 3 6 0e1 1 3 6 811366

run by R. capsulatus DSM 1710 for 35 days. After 55th day of

photobioreactor run by R. capsulatus hup- and 35th day of

photobioreactor run by R. capsulatus DSM 1710 the photo-

bioreactors were not cooled because of the low air tempera-

ture. Hence, there were fleeting temperature increases above

45 �C in the photobioreactors in several days. The cell

concentration in the photobioreactors was not affected

significantly from these temperature increases. However,

hydrogen productivity and yield decreased. At the same

period, the temperature of the photobioreactors decreased

below 10 �C during nights, due to seasonal reasons. It may be

one of the reasons causing a decrease in hydrogen productivity

and acetate consumption. Extreme temperature variations

were reported to decrease the hydrogen production signifi-

cantly, in batch cultures of R. capsulatus [18]. The optimum

temperature for hydrogen production by R. capsulatus is 30 �C.Hence, in outdoor experiments a control for minimum and

maximum temperature is required during day and night.

3.5. Evaluation of the maximum productivity and yields

Table 2 summarizes TOC, TN, COD and NHþ4 values in the feed

at the first day and in the effluent at the last day of experiments

Table 2 e TOC, TN, COD and NHD4 concentration in the

feed and in the effluent of photobioreactor.

R. capsulatus hup- R. capsulatus DSM 1710

Component Feed Effluent Effluent

TOC (mg/L) 1870 1200 1830

TN (mg/L) 420 42 129

C/N 5 29 14

COD (mg/L) 6500 4890 5880

NHþ4 (mg/L) 1 0.6 0.0

in the photobioreactors. The total organic carbon (TOC) and the

total nitrogen (TN) in the photobioreactors reduced due to

cellular activity. The C/N ratio increased during operation

which is critical for hydrogen production by photosynthetic

bacteria [16]. That might be due to a higher consumption rate

of nitrogen sources present in themolasses DFE such as amino

acids and ammonia. The optimum C/N ratio is suggested as 20

in continuous operations by R. capsulatus hup- [17]. Ammonia

present in the molasses DFE was utilized completely and no

ammonia inhibitionwas observed. In batch operation theDFEs

with low ammonia content are essential for the high hydrogen

productivity since the ammonia is the strong inhibitor of the

nitrogenase enzyme [15].

Table 3 summarizes the productivities, and yields in the

photobioreactors. The maximum hydrogen yield achieved

with R. capsulatus hup- was 78% (of the theoretical maximum)

in continuous operation which was much higher than the

yield obtained by R. capsulatus DSM 1710 (50% of theoretical

maximum) at the same operating conditions. The maximum

hydrogen productivity of R. capsulatus hup- on the molasses

DFE was 0.67 mmol H2/Lc.h which is higher than the produc-

tivity of the wild type strain (0.50 mmol H2/Lc.h).

Table 3 e Maximum molar hydrogen productivity andyields.

Microorganism Process Maximumhydrogen

productivityðQH2

Þmmol H2/(Lc.h)

MaximumMolarYield

(YN) (%)

R. capsulatus

hup-

Batch [15] 1.37 58

Continuous 0.67 78

R. capsulatus

DSM 1710

Batch [15] 0.75 20

Continuous 0.50 50

i n t e r n a t i o n a l j o u r n a l o f h y d r o g e n en e r g y 3 6 ( 2 0 1 1 ) 1 1 3 6 0e1 1 3 6 8 11367

A promising outcome of the continuous operations is that

the yields obtained in continuous cultures of both R. capsulatus

hup- and R. capsulatus DSM 1710 are higher than that obtained

in batch cultures (Table 3).

4. Conclusions

In this study photobiological hydrogen production on dark

fermentor effluent of molasses was achieved at a high rate

and productivity using R. capsulatus hup- and R. capsulatusDSM

1710. The biomass concentration remained stable for 2

months of operation. Hydrogen productivity and yield are

higher at high light intensity; they decrease with decreasing

light intensity due to seasonal changes.

This study showed that, the molasses DFE can be used as

a substrate for long term photofermentative hydrogen produc-

tion in outdoor conditions. During summer the maximum

temperature should not be higher than 38 �C. The temperature

was effectively controlled by placing internal cooling coil into

the panel reactor. For stable operation, controlling the

maximum temperature during the day and the minimum

temperature during the night is strongly recommended.

Acknowledgements

This study was supported by Hyvolution the EU 6th Frame-

work Integrated Project 019825: Non-thermal production of

pure hydrogen from biomass. We thank to Dr. Truus de Vrije

fromWageningen UR, Food and Biobased Research group, The

Netherlands for supplying molasses DFE. We acknowledge

Kerime Guney from Department of Chemical Engineering and

Tamay S‚ eker from Central Laboratory of Middle East Tech-

nical University, Ankara, Turkey, for carrying out elemental

and organic acid analysis.

Symbols

C Concentration, (mM)

Lc Liter culture

N Number of moles, (mole)

n number of days

P Pressure, (atm)

PBR Photobioreactor

PNS Purple non-sulfur bacteria

QH2Molar productivity of hydrogen, (moleH2/Lc.h)

R Rhodobacter

V Volume, (L)

T Temperature, (�C),t Time, (h)

Y Yield (%)

Greek letters

h Efficiency (%)

Subscript

c Culture

exp Experimental

F Feed

i Initial

theo Theoretical

ut Utilized

Abbreviations

AA Acetic acid

BA Butyric acid

C/N Carbon to Nitrogen Ratio

COD Chemical oxygen demand, (mg/L)

DFE Dark Fermentor Effluent

FA Formic acid

GC Gas Chromatography

hup- Uptake hydrogenase deficient (mutant)

HPLC High Performance Liquid Chromatography

LA Lactic acid

PA Propionic acid

TN Total nitrogen, (mg/L)

TOC Total organic carbon, (mg/L)

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