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initiation of cell growth) lengthens as salt concentration increases. (ii) Utilization of the primary carbon and energy source is reduced. (iii)
Change in concentration of metabolic products:
(a) there is a decrease in the production of
ethanol as salt content increases and (b) there is
an increase in the concentration of other fermentation products (such as glycerol, acetaldehyde,
etc.) as salt content increases.
room
temperature.
I'o-
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-4.0
0If
0 8.0-o
wz 6.0
>_x
-5.0
a3
70
60
50
40
30
20
FERMENTATION PERIOD, hr
Effect of NaCl levels on yeast biomass in
10
FIG. 1.
semisolid growth cultures of S. cerevisiae in ca. 10%
glucose. All points represent data points.
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I_
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u
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u
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--
10
20
40
30
50
60
70
FERMENTATION PERIOD, hr
10
50
60
40
30
20
FERMENTATION PERIOD, hr
70
FIG. 4. Effect of NaCl levels on glycerol production in semisolid growth cultures of S. cerevisiae in ca.
10% glucose.
lytes in several yeasts promote high accumulations of polyhydric organic compounds, such as
glycerol, 2,3-butanediol, arabitol, and erythritol.
Tajima and Yoshizumi (10) have found that
Saccharomyces formosensis Nakazawa in 0.25
to 1.00 M NaCl converted much of the sugar
substrate into glycerol, 2,3-butanediol, mannitol, erythritol, organic acids, vicinal diketone,
acetaldehyde, and CO2.
Our gelatin study results corroborate previous
reports on the effect of salt in stimulating the
production of glycerol during the fermentation
of glucose by S. cerevisiae in liquid culture (Fig.
4). However, it should be noted that the control
(0.0% NaCl) curves, after reaching a plateau,
subsequently show a gradual drop in glycerol
beginning at about 24 h, at which time the
glucose substrate has been exhausted (Fig. 2),
suggesting the shift to glycerol as substrate for
the yeast cells. This suggestion is reinforced by
the diauxie curves (Fig. 1) for low salt levels.
The same sequence of events is seen in the 1.0,
2.0, and 3.0% NaCl curves, with the reduction in
glycerol beginning after 30 to 35 h. The concentration of glycerol continued to build up in the
4.0 and 5.0% NaCl cultures in which (Fig. 2)
glucose was not exhausted during the length of
%NaCI
.- 0.0
0-2.0
- 3.0
- 40 _
A- 5.0
o
4
10
20
30
60
50
40
FERMENTATION PERIOD, hr
70
the run.
Brown (2) has discussed evidence for considering the polyols synthesized by S. cerevisiae as
fulfilling several physiological functions in the
yeast, including the role of food reserves.
The acetaldehyde curves (Fig. 5) corroborate
the previously observed (10, 20) increasing
buildup of acetaldehyde in liquid cultures at high
NaCl levels, but only if measurements are made
between 10 and 17 h or after 40 h. All of the
curves, including the control curve, reach a
maximum and then fall off at variable rates.
Tempest et al. (17) have reported that the
presence of 4% (ca. 0.67 M) NaCl S. cerevisiae-
CD
%NoCl
-0.0
3.0
0 -4.0
A - 5.0
x- o.o
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a-
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O
0-2.0
i 0.4 _-3.0
4
0-4.0
*-5.0
5.0
10
20
30
40
50
60
70
FERMENTATION PERIOD, hr
FIG. 5. Effect of NaCi levels on acetaldehyde pro-
0-
U0.5
0.4
o.3
70
1.4
2.0
Es4
0
_-
0.6
60
1.2
1.0
8-3.0
0-4.0
' 0.7
liquid culture.
Mechanisms proposed to explain the inhibitory
effects of NaCI on yeast fermentations. Umemoto
et al. (19) have suggested a partial explanation of
the inhibitory effects of electrotytes in terms of
the inhibition of yeast (pyruvate) carboxylase,
the enzyme that catalyzes the decarboxylation
of pyruvate to acetaldehyde. Obviously, if acetaldehyde production is slowed down, the production of ethanol is reduced proportionately.
They concluded that in the absence of acetaldehyde, if the fermentation was to continue, a
hydrogen acceptor other than acetaldehyde
must become available to oxidize the NADH
-0.0
1.0
20
30
40
50
FERMENTATION PERIOD, hr
x -0.0
3i
10
FIG. 7. Effect of NaCl levels on the pH of semisolid growth cultures of S. cerevisiae in ca. 10% glucose.
ON
%NOCI
0.8
0.6
0.4
0.2
14
0.2
0
0 .3
0.1
o.
U)
'0
10
20
30
40
50
60
70
FERMENTATION PERIOD, hr
6. Effect of NaCl levels on intracellular lysine
FIG.
production in semisolid growth cultures of S. cerevisiae in ca. 10%o glucose.
.11
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parameter indicates greater alcohol dehydrogenase decay with time as the NaCl level increases.
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ACKNOWLEDGMENTS
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LITERATURE CITED
stress
recvery
isreducedand
acetaldehyde
ethanol).
to ethaol).
acetldehydeto
This would
Ths
would
explaiin the
181-242. In A. H.
Laboratories.
4a.Hodge, J. E.,
reducing
and
Determination of
and carbohydrates. Methods Carbohydr.
Chem. 1:386-388.
5.
mination of ethanol,
p.
45:2554-2560.
7.Norkrans,
B. 1966.
growth related
to
Studies
on
marine-occurring yeasts:
that e thanol
production is reduced
,thanolproducion
p.
shown in Fig. 8.
forme d
in eukaryotic microorganisms,
and
tempera-
54:374-392.
p.
1-42. In D. J. D. Hockenhull
yeast
Technol. 50:764-769.
11. Tajima, K., and H. Yoshizumi. 1975. Mechanisms of
abnormal fermentation of distiller's yeast in salted media
(such
NAD(P) redox
slThe inhibition
of fermentation by the
highly concentrated
salts in molasses. J. Ferment. Technol. 44:77-84.
in
enzyme
and
New York.
R. D.,
time e mnzyme activities becomes even more pronounc:ed when the area of hysteresis curve (another measure
measure of the difference bbetween
een
upper
lo
trajectorifferenes)is graphed
functiion
is
Fig.
9.
This
D.
C.-J.
and
on
J.
the intracel-
as
new
Tech.
Biotech.
fermentation
process
for
A
producing both ethanol and
This study developed from the United States-Taiwan Cooperative Science Program, which included the United StatesRepublic of China Seminar on Fermentation Engineering held
at the University of Pennsylvania on 30 May to 1 June 1978
and the Fermentation Engineering Research visit to Taiwan
between 6 and 21 June 1979 under the sponsorship of the
Division of International Programs of the National Science
Foundation (Project no. FCV-147).
S. Y. Huang and C. H. Lin collaborated in this effort. H. H.
Wang contributed significantly to the definition of the study
described in this report.
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