Factors affecting ethanol fermentation using Saccharomyces

cerevisiae BY4742
Yan Lin
a,b,
*, Wei Zhang
a
, Chunjie Li
a
, Kei Sakakibara
b
, Shuzo Tanaka
b
, Hainan Kong
a
a
School of Environmental Science and Engineering, Shanghai Jiao Tong University, Shanghai 200240, PR China
b
Program of Environment and Ecology, Faculty of Science and Engineering, Meisei University, Tokyo 191-8506, Japan
a r t i c l e i n f o
Article history:
Received 9 November 2011
Received in revised form
1 February 2012
Accepted 5 September 2012
Available online xxx
Keywords:
Ethanol
Fermentation
Saccharomyces cerevisiae
Yeast
Thermotolerant ability
Strain optimization
a b s t r a c t
Fermentation of sugar by Saccharomyces cerevisiae BY4742, for production of ethanol in
a batch experiment was conducted to improve the performance of the fermentation
process. The thermotolerant ability of S. cerevisiae to grow and ferment glucose at elevated
temperatures similar to the optima for saccharication was investigated. The inuences of
temperature, substrate concentration and pH on ethanol fermentation were observed. The
yield for ethanol production and changes in the fermentation pathway were compared
under different conditions.
When the temperature was increased to 45

C, the system still showed high cell growth
and ethanol production rates, while it was inhibited at 50

C. The maximum specic
growth rate and the maximum specic ethanol production rate were observed between 30
and 45

C with different initial glucose concentrations. The maximum sugar conversion at
30

C after 72 h incubation was 48.0%, 59.9%, 28.3%, 13.7% and 3.7% for 20, 40, 80, 160 and
300 kg m
3
of glucose concentrations respectively. Increased substrate supply did not
improve the specic ethanol production rate when the pH value was not controlled. pH 4.0
e5.0 was the optimal range for the ethanol production process. The highest specic
ethanol production rate for all the batch experiments was achieved at pH5.0 which is
410 g kg
1
h
1
of suspended solids (SS) which gave an ethanol conversion efciency of
61.93%. The highest specic ethanol production rate at 4.0 was 310 g kg
1
h
1
of SS. A
change in the main fermentation pathway was observed with various pH ranges. Forma-
tion of acetic acid was increased when the pH was below 4.0, while butyric acid was
produced when the pH was higher than 5.0. In the presence of oxygen, the ethanol could be
utilized by the yeast as the carbon source after other nutrients became depleted, this could
not occur however under anaerobic conditions.
2012 Elsevier Ltd. All rights reserved.
1. Introduction
Growing attention has been devoted in the past years to the
conversion of biomass into fuel ethanol, considered the
cleanest liquid fuel alternative to fossil fuels. It is now
understood that it is important to use biomass energy as
a means of providing modern energy to the billions who lack
it, and it may also be a viable alternative energy source to the
worlds ever depleting natural reserves [1].
There are several kinds of raw materials for ethanol
fermentation. For the utilization of cellulose as the raw
material, simultaneous saccharication and fermentation
* Corresponding author. School of Environmental Science and Engineering, Shanghai Jiao Tong University, Hino, Shanghai 200240,
PR China. Tel.: 86 21 54744540; fax: 86 21 54740825.
E-mail address: linyansjtu@126.com (Y. Lin).
Available online at www.sciencedirect.com
ht t p: / / www. el sevi er. com/ l ocat e/ bi ombi oe
b i oma s s a nd b i o e ne r g y x x x ( 2 0 1 2 ) 1 e7
Please cite this article in press as: Lin Y, et al., Factors affecting ethanol fermentation using Saccharomyces cerevisiae BY4742,
Biomass and Bioenergy (2012), http://dx.doi.org/10.1016/j.biombioe.2012.09.019
0961-9534/$ e see front matter 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.biombioe.2012.09.019
(SSF) combines enzymatic hydrolysis of cellulose with simul-
taneous fermentation of the sugars obtained to ethanol, and
gives higher reported ethanol yields. This therefore requires
lower amounts of enzyme because the yeast fermentation
step helps reduce the end products inhibition from cellobiose
and glucose formed during enzymatic hydrolysis [2e4].
However, SSF requires compatible fermentation and
saccharication conditions with a similar pH, temperature
and optimum substrate concentration [5,6]. The main dif-
culty with SSF of cellulose is the different temperature optima
required for saccharication (40e50

C) and fermentation
(20e35

C) [5e7]. In order to preserve the viability of the yeast,
the SSF process is typically operated between 30 and 38

C at
which the cellulose is far below its peak operational level [6].
The use of thermotolerant yeasts capable of fermenting
glucose to ethanol at elevated temperatures, which are closer
to the optima for the activity of the cellulolytic complex, is
therefore advisable when employing SSF processes [7].
Moreover, for the existing or converted fermentable
sugars, it is also important to improve the efciency of the
fermentation system to utilize it with a high ethanol produc-
tion ability. On the other hand, since the distillation cost per
unit amount of ethanol produced is substantially higher at low
ethanol concentrations [8], several investigators have dealt
with the idea of concentrating sugar solutions prior to
fermentation [8e11]. Clearly it is necessary to improve the
ethanol fermentation performance and to solve the problem
between the concentration of ethanol produced and sugar
added if an economically sustainable system is to be created
using this method. During batch fermentation, many inu-
ential parameters, such as pH, pO
2
and temperature, can
greatly inuence the specic rate of growth, and inhibition
can be caused either by product or substrate concentration
[12,13]. The viability of cell populations, the specic rate of
fermentation and the sugar uptake rate are all directly related
to the desired medium condition [14e16].
The purpose of this research was to obtain high ethanol
production with high productivity and to test the thermoto-
lerant ability of Saccharomyces cerevisiae to grow and ferment
glucose at elevated temperatures similar to the optima for
saccharication. The effect of temperature, pH value and
initial glucose concentration on the production of ethanol by
S. cerevisiae was evaluated to improve the ethanol fermenta-
tion performance.
2. Materials and methods
2.1. Yeast strain and culture conditions
S. cerevisiae BY4742 (originally from EUROSCARF, Germany)
[17,18], a strain preserved at the laboratory (Department of
Chemistry, Meisei University, Japan), was used in this study. S.
cerevisiae was maintained at 4

C on agar slants containing (in
kg m
3
): bacto-yeast extract 10; bacto-peptone 20; glucose 20,
and bacto-agar 20. The prepared media was sterilized at 121

C
for 20 min. Pre-cultures were inoculated from agar slants and
grown at 30

C overnight in 250 ml shake-asks with mineral
medium containing (in kg m
3
): bacto-yeast extract 10; bacto-
peptone 20 and glucose 20 while stirring at 1.8 Hz. Pre-cultures
of (10 ml) were used to inoculate 500 ml bafed shake-asks
containing 250 ml of the above media. The inoculum was
grown for approximately 24 h and used to further inoculation.
2.2. Batch fermentation
Batch fermentationexperiments onthe effects of temperature
and initial substrate concentrations under microaerobic
conditions (stirring at 1.8 Hz with a nal dissolved oxygen
concentration of 1.0 g m
3
) were carried out in duplicate twice
using 20e300 kg m
3
of glucose of the initial glucose solution
as the sole carbon source for S. cerevisiae. Experiments were
performed in 500 ml Erlenmeyer asks with 250 ml total liquid
volume and were initiated by transferring 5% of the starter
culture to the prepared media. The experiments were carried
out for 7 days in isothermal conditions at 30

C and monitored
by harvesting 15 ml samples every 24 h for analyses.
Experiments on the effect of pH on ethanol fermentation
were run in a fermentor (MBF-500ME P.A., EYELA) with
a 0.003 m
3
working volume, stirred at 1.3 Hz, and the tempera-
ture was controlled at 30

C. The pH value was controlled by
automatic addition of 2 kmol m
3
NaOH or HCl. The fermentor
was sealed and equipped with a syringe for sample removal. It
was also tted with a CO
2
exhaustion port to allow venting of
carbon dioxide which is a byproduct of the reaction. The
experiments were started by adding the specic amount of
yeast inoculatedintothemediumcontainingdifferent amounts
of glucose. The nal yeast concentration was 2 kg m
3
of SS.
2.3. Analytical methods
Ethanol and Volatile Fatty Acid (VFA) were measured using
a gas chromatograph (GC-8APF, SHIMADZU) equipped with
a 3 m 2.6 mm glass column packed with polyethyleneglycol
(PEG) (Chromosob W 80/100 AW-DMCS, SHIMDZU) (80e100
mesh). The column temperatures for ethanol and VFA
analyzing were 80

C and 140

C, respectively. The injection
port and ame ionization detector temperature for ethanol
was 100

C, while for VFA it was 180

C. Nitrogen, used as the
carrier gas for both ethanol and VFA, was set at gas ow
pressures of 200 and 150 kPa, respectively. Before injection,
analyzed samples were ltered through a 0.20 mm membrane
lter to allow determination of the VFA and ethanol concen-
trations in supernatants. Cell growth was determined by
measuring the optical density (OD) at 600 nm using a spec-
trophotometer (UV-1600, SHIMADZU). The cell dry weight was
obtained using a calibration curve. The cell dry weight was
proportional to cell turbidity and absorbance at 600 nm [19].
Biomass concentration was determined by use of the dry-
weight method for SS [13]. Accordingly, samples were centri-
fuged, and then the settled solids were washed with distilled
water and dried for 2 h at 105

C.
3. Results and discussion
3.1. Inuence of temperature
Competition during ethanol fermentation carried out at
different temperatures may be a way of testing the endurance
b i o ma s s a nd b i oe ne r g y x x x ( 2 0 1 2 ) 1 e7 2
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Biomass and Bioenergy (2012), http://dx.doi.org/10.1016/j.biombioe.2012.09.019
of the strain used in this system. This could then be used as
a method for determining the optimal condition for ethanol
fermentation and also a criterion for rapidly selecting one of
several strains while at the same time studying resistance to
temperature in a controlled situation, i.e. under laboratory
conditions.
In this study, the inuence of temperature on the ethanol
fermentation by S. cerevisiae BY4742 was studied with regard
to biomass and ethanol production. Batch fermentation in
shake asks for ethanol production was carried out in dupli-
cate for one week at various initial glucose concentrations
from20 to 300 kg m
3
and controlled at constant temperatures
of 10, 20, 30, 40, 45 and 50

C. Experimental results revealed the

cells increased exponentially at the beginning of incubation,
then entered a stationary phase after several days incubation,
for all operating temperatures. Higher temperatures made the
exponential growth of the cells shorter (data not shown).
Fig. 1 shows the changes in ethanol concentration at
different temperatures with the initial glucose concentration
of 40 kg m
3
and yeast concentration of 2 kg m
3
of SS over
a one week incubation period. For general ethanol production
by yeast, the maximum fermentation time in batch process
was 72 h [20].
Experimental data in Fig. 1 showed that when the
temperature increased, the maximum fermentation time was
shortened, but a much higher temperature inhibited the
growth of cells and then the fermentation signicantly
declined. In this study, cell growth and ethanol production
declined considerably at 50

C, which showed the inhibition
effect on cell growth at higher temperatures. This phenom-
enon may be explained because the higher temperature
results in changing the transport activity or saturation level of
soluble compounds and solvents in the cells, which might
increase the accumulation of toxins including ethanol inside
cells [20,21]. Moreover, the indirect effect of high temperature
might also be ascribed to the denaturation of ribosomes and
enzymes and problems with the uidity of membranes [20,21].
However, at lower temperatures the cells showed lower
specic growth rates which may be attributed to their low
tolerance to ethanol at lower temperatures [22,23]. The
maximum specic growth rate and the maximum specic
ethanol production rate were observed between 30 and 45

C
withdifferent initial glucose concentrations as showninFig. 3.
It is commonly believed that 20e35

C is the ideal range for
fermentation and at higher temperatures almost all fermen-
tation would be problematic [6,7,12,20,22]. However, as shown
in Figs. 2 and 3 in this study, when the temperature was
increased to 45

C, the system still showed a high cell growth
and ethanol production rates and the lowest m
t
/m
30
at different
glucose concentrations was around 0.8. We also observed
a higher specic ethanol production rate at higher glucose
concentrations when tested at 45

C. With a higher tolerant
fermentation temperature, similar to the optimal temperature
for cellulolytic activity, it may be possible for the SSF process
to improve the nal efciency. Moreover, as shown in Fig. 1,
ethanol yields may further be improved at elevated temper-
atures for a shorter culture time.
In addition, the ethanol concentration was found to peak
and then decline at temperatures above 20

C, and the lower
glucose concentration made the decline time occur earlier.
These biochemical changes may indicate that cells originally
growing on glucose switched from a fermentative metabo-
lism using mainly glycolysis and forming ethanol, to a respi-
ratory metabolism in which the ethanol formed in the earlier
stages of growth was consumed using the tricarboxylic acid,
glyoxylate cycles and mitochondrial electron transport chain
[24]. Results in Fig. 1 also show that ethanol concentration
rose steadily at low temperatures and wont decline within
168 h, possibly because at these lower temperatures the yeast
was not active which is because of the low tolerance to
ethanol [22,23].
0
5
10
15
20
25
0 24 48 72 96 120 144 168
Incubation time(hours)
E
t
h
a
n
o
l

c
o
n
c
e
n
t
r
a
t
i
o
n

(
k
g

m
-
3
)
10C 20C
30C 40C
45C 50C
Fig. 1 e Changes in ethanol concentration at different
temperatures with an initial glucose concentration of
40 kg m
L3
over a one week incubation period.
0.2
0.4
0.6
0.8
1.0
1.2
1.4
1.6
1.8
2.0
10 20 30 40 50
Temperature (C)

t
/

3
0
20 kg m-3
40 kg m-3
80 kg m-3
160 kg m-3
300 kg m-3
Fig. 2 e Ratio of specic growth rate (m) at different
controlled temperatures to that at 30

C with different
initial sugar concentrations after 72 h incubation (m
t
/m
30
is
a ratio of specic growth rate at t

C to m at 30

C).
b i oma s s a nd b i o e ne r g y x x x ( 2 0 1 2 ) 1 e7 3
Please cite this article in press as: Lin Y, et al., Factors affecting ethanol fermentation using Saccharomyces cerevisiae BY4742,
Biomass and Bioenergy (2012), http://dx.doi.org/10.1016/j.biombioe.2012.09.019
3.2. Inuence of substrate concentration
The batch experiment was performed with various glucose
concentrations to develop ethanol production. The initial
glucose concentrations in the batch experiments were 20, 40,
80, 160 and 300 kg m
3
tested at 30

C. The experimental
conditions and the results summarized in Fig. 4 show the
changes in ethanol concentrations at different initial glucose
concentrations over a one week incubation period, while Fig. 5
demonstrates the specic ethanol production rate and
ethanol conversion rate at different initial glucose concen-
trations after 48 and 72 hours incubation.
The production of ethanol was affected by the substrate
concentration between 20 and 300 kg m
3
. As shown in Fig. 4,
higher substrate concentrations may achieve higher ethanol
production, but a longer incubation time was required for
higher initial glucose concentrations above 80 kg m
3
at
a temperature of 30

C when the pH was not controlled. More-
over, higher initial glucose concentrations, such as 300 kg m
3
,
may have actually decreased the ethanol conversionefciency
when the pH value was not controlled, since the higher
substrate and production concentrations may have inhibited
the process of ethanol fermentation (as shown in Fig. 5).
Fig. 5 shows the specic ethanol production rates and
ethanol conversion efciency at different initial sugar
concentrations after 48 and 72 hours incubation at 30

C. The
data above illustrates that higher initial glucose concentration
may decrease the ethanol conversion efciency. The
maximum sugar conversion after 72 hours incubation was
observed at 48.0%, 59.9%, 28.3%, 13.7%, and 3.7% for 20, 40, 80,
160 and 300 kg m
3
of glucose, respectively. More substrate did
not improve the specic ethanol production rate when the pH
value was not controlled.
3.3. Inuence of pH
Improved ethanol fermentation activity can be achieved by
controlling various parameters. In addition to temperature
and substrate concentration, pH is also a key factor that
affects ethanol fermentation [13]. In this study changes in
ethanol and VFAs were investigated to estimate the activity of
the ethanol production ability with changes in pH. This was
examined at pHs 3.0, 4.0, 5.0, 5.5 and 6.0 in an anaerobic Jar
Fermentor.
Fig. 6 shows the results of the batch test used to investigate
the effect of pH on ethanol production. When the pH was
lower than 4.0, the incubation time for maximum ethanol
Fig. 3 e Specic ethanol production rates at different initial
sugar concentrations with different controlled
temperatures after 72 h incubation. * Specic ethanol
production rates were calculated as milligrams of ethanol
produced per grams of cell mass per hour (g kg
L1
h
L1
of SS).
0
5
10
15
20
25
30
35
0 24 48 72 96 120 144
Incubation time(hours)
E
t
h
a
n
o
l

c
o
n
c
e
n
t
r
a
t
i
o
n

(
k
g

m
-
3
)
20 kg m-3
40 kg m-3
80 kg m-3
160 kg m-3
300 kg m-3
Fig. 4 e Changes in ethanol concentration under different
glucose concentrations after 6 days incubation at 30

C.
0
20
40
60
80
100
120
140
160
180
200
0 50 100 150 200 250 300
Initial glucose concentration(kg m
-3
)
S
p
e
c
i
f
i
c

E
t
h
a
n
o
l

P
r
o
d
u
c
t
i
o
n

R
a
t
e
(
g

k
g
-
1

h
-
1
)

a
n
d
E
t
h
a
n
o
l

C
o
n
v
e
r
s
i
o
n

R
a
t
e

(
%
)
48h-SEPR 72h-SEPR
48h-ECR 72h-ECR
Fig. 5 e Specic ethanol production rates and ethanol
conversion efciency at different initial sugar
concentrations after 48 and 72 h incubation at 30

C.
b i o ma s s a nd b i oe ne r g y x x x ( 2 0 1 2 ) 1 e7 4
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concentration was prolonged, but the maximum concentra-
tion was not very low. When the pH value was above 5.0, the
quantity of ethanol produced substantially decreased.
Therefore a pH range of 4.0e5.0 may be regarded as the
operational limit for the anaerobic ethanol production
process. The highest specic ethanol production rate for all
the batch experiments was achieved at pH 5.0 which is
410 g kg
1
h
1
of SS, with an ethanol conversion efciency of
61.93%. The specic ethanol production rate at pH4.0 was
310 g kg
1
h
1
of SS, which is not signicantly lower than the
value obtained at pH5.0. Therefore, considering the chemical
requirement for pHadjustment, pH4.0 may be regarded as the
operational limit for the ethanol production process.
In addition, in Fig. 6 the ethanol concentration did not
decrease after the nutrient was consumed as in Figs. 1 and 4.
This may indicate that the ethanol could not be utilized as the
carbon source under anaerobic condition.
Our experimental results could indicate that pH plays
an important role in determining the fermentation pathway
used in anaerobic ethanol production processes. Table 1
shows competition for the substrate, glucose, by the
microorganisms, and may suggest a change in the main
fermentation pathway at various pHranges. The above results
show that the main products were ethanol and butyrate
between pH 5.5e6.0 at 30

C with the initial glucose concen-
tration of 40 kg m
3
. When the pH value was lower than 5.0,
acetic acid was the main product. These results suggest that
the mechanism of ethanol production may include the reac-
tions as follows:
4C
6
H
12
O
6
/2CH
3
COOH3CH
3
CH
2

2
COOH8H
2
8CO
2
(1)
C
6
H
12
O
6
H
2
O/C
2
H
5
OHCH
3
COOH2H
2
2CO
2
(2)
Eqs. (1) and (2) refer to Moat and Gaudy and Gaudy,
respectively [25,26]. As indicated in Eq. (1), 4 mol of glucose
were converted to 2 mol of acetic acid and 3 mol of butyrate
acid. Here, although the butyrate acid formation is not popu-
larly discussed in yeast metabolism, this phenomena was
observed in this study and also agreed with the results of
Teresa and Carmen [27].
There are also many examples of an alteration of the fatty
acid prole in the yeast [28,29]. During this process, with the
pH value higher than 5.0, much glucose was consumed and
0
5
10
15
20
25
0 24 48 72 96 120 144 168
Incubation time(hours)
E
t
h
a
n
o
l

c
o
n
c
e
n
t
r
a
t
i
o
n

(
k
g

m
-
3
)
pH=5.0 pH not controlled
pH=4.0 pH=6.0
pH=5.5 pH=3.0
Fig. 6 e Changes in ethanol concentration with an initial
glucose concentration of 40 kg m
L3
over one weeks
incubation with different pH values.
Table 1 e The ratio of ethanol and VFAs to total products
a
after 48 and 72 h incubation at 30

C with different pH values.
pH value Ratio of ethanol and VFAs after 48 h incubation (%) Ratio of ethanol and VFAs after 72 h incubation (%)
Ethanol Acetic acid Butyric acid Ethanol Acetic acid Butyric acid
No control 65.36 1.41 0.15 65.55 1.48 0.10
3.0 65.15 2.21 0.07 65.15 2.51 0.07
4.0 65.54 1.32 0.09 65.54 1.28 0.05
5.0 65.54 1.63 0.02 65.54 0.86 1.51
5.5 56.49 6.01 9.18 56.49 3.01 15.74
6.0 48.80 9.00 17.05 48.80 6.05 14.80
a Total products include ethanol, VFAs, CO
2
and glucose in terms of carbon.
0
50
100
150
200
250
300
350
400
450
500
0 24 48 72 96 120 144 168
Incubation time(hours)
S
p
e
c
i
f
i
c

e
t
h
a
n
o
l

p
r
o
d
u
c
t
i
o
n

r
a
t
e

(
g

k
g

-
1

h

-
1
)
300kg m-3-pH not controlled
300kg m-3-pH4
160kg m-3-pH not controlled
160kg m-3-pH4
Fig. 7 e Comparison of the specic ethanol production
rates between pH values set at 4.0 and uncontrolled pH
with initial sugar concentrations of 160 and 300 kg m
L3
after one weeks incubation at 30

C.
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converted to by-products, so the ethanol conversion efciency
was greatly decreased (as shown in Figs. 7 and 8).
In the case of Eq. (2), 1 mol of glucose was converted into
1 mol of ethanol and 1 mol of acetic acid. Although there was
some ethanol produced, the ethanol fermentation yield was
still reduced by the acetic acid production. Table 1 shows that
when the pH was 4.0 and 5.0, the quantity of by-products was
less than that observed under other conditions.
Results in section 3.2 show that a higher substrate
concentration may prevent the ethanol fermentation process
occurring. One of the reasons may be the accumulation of
high concentrations of ethanol and by-products which make
the pH change. So if the pH was set at a suitable value, the
efciency might be somewhat increased. Compared with
fermentation where the pH was not controlled, when the pH
was controlled at 4.0, the VFAs in the nal product were
reduced and the specic ethanol production rate and the
ethanol fermentation efciency were signicantly improved
(as shown in Figs. 7 and 8). After pH adjustment, the ethanol
fermentation yields were increased to the range of 99.8% and
97.4% of the maximum theoretical value with the initial
glucose concentrations of 160 and 300 kg m
3
, respectively.
The highest specic ethanol production rates at pH 4.0 were
340 and 260 g kg
1
h
1
of SS, respectively.
4. Conclusions
The present work tested the thermotolerant ability of S. cer-
evisiae to grow and ferment glucose at elevated temperatures
and examined the inuences of temperature, initial substrate
concentration and pH value on ethanol fermentation.
The maximum specic ethanol production rates were
observed between 30 and 45

C with different initial glucose
concentrations. Whenthe temperature was increased to 45

C,
the system still showed higher cell growth and ethanol
production rates and the lowest m
t
/m
30
at different initial
glucose concentrations was 0.8.
Higher substrate concentration could achieve higher
ethanol production, but a longer incubation time was required
for initial glucose concentrations above 80 kg m
3
at 30

C
when the pH was not controlled.
The changes in the operational pH in the ethanol produc-
tion process may have induced a change in the main
fermentation pathway. Thus it is important to control pH
value in the range of 4.0e5.0. Beyond this range, the formation
of by-products, such as acetic acid and butyric acid may have
consumed some of the substrate and reduced the efciency of
ethanol fermentation.
Acknowledgments
The research project was sponsored by Major Science and
Technology Program for Water Pollution Control and Treat-
ment (2009ZX07101-015-003) and State Key Laboratory of
Pollution Control and Resource Reuse Foundation
(PCRRF09002).
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0
10
20
30
40
50
60
70
80
90
100
0 24 48 72 96 120 144 168
Incubation time(hours)
E
t
h
a
n
o
l

c
o
n
v
e
r
s
i
o
n

e
f
f
i
c
i
e
n
c
y

(
%
)
300 kg m-3-no control
300 kg m-3-pH4
160 kg m-3-no control
160 kg m-3-pH4
Theoretical value
Fig. 8 e Comparison of ethanol conversion efciency
between pH set at 4.0 and uncontrolled pH with initial
sugar concentrations of 160 and 300 kg m
L3
after one
weeks incubation at 30

C. * The maximum theoretical
ethanol fermentation yield was expressed by carbon and
calculated according to the equation of
C
6
H
12
O
6
/2C
2
H
5
OH D 2CO
2
.
b i o ma s s a nd b i oe ne r g y x x x ( 2 0 1 2 ) 1 e7 6
Please cite this article in press as: Lin Y, et al., Factors affecting ethanol fermentation using Saccharomyces cerevisiae BY4742,
Biomass and Bioenergy (2012), http://dx.doi.org/10.1016/j.biombioe.2012.09.019
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Yan Lin holds a Ph.D. (2005) in environmental engineering from
Shanghai Jiao Tong University, China. She has worked as a post-
doctoral researcher in the Asian Center for Environmental
Research at Meisei University, Japan since 2005. She is now an
associate professor in Shanghai Jiao Tong University. Her current
project is Environmental conservation technology and utilization
of biomass in Asia (environmental technology) e ethanol
fermentation, nanced by the Ministry of Education, Culture,
Sports, Science and Technology, Japan.
b i oma s s a nd b i o e ne r g y x x x ( 2 0 1 2 ) 1 e7 7
Please cite this article in press as: Lin Y, et al., Factors affecting ethanol fermentation using Saccharomyces cerevisiae BY4742,
Biomass and Bioenergy (2012), http://dx.doi.org/10.1016/j.biombioe.2012.09.019