Yeast and fermentation: the optimal pH
Vika Shimanskaya, Karin Bartels
Dominicus College, The Netherlands
14 April 2010
Summary
The ability of yeast to convert sugar into ethanol has been used since ancient times in baking
and brewing. The exhaustion of oil resources has stimulated interest in researches after the
alternative sources of energy. The production of ethanol by yeast fermentation is one of the
most common ways of getting bio-ethanol. The yeast fermentation occurs in an oxygen free
environment and raises the question of what the optimal pH will be in the conversion of
glucose or other sugars to ethanol by yeast cells, Saccharomyces cerevisiae. The fermentation
process was followed at pH’s 2,8; 4,4; 6,0; 8,4 by measuring the release of CO2. This resulted
in an optimal pH of 2,8. But it also raised the question like what is the influence of ethanol on
yeast cells. We found an amount of CO2 that was less than optimal.
Introduction
Ethanol fuel is widely used in lots of
countries as a transport fuel. World ethanol
production tripled during the last seven
years. Ethanol is usually produced by the
reaction with ethene (g) and water (g):
CH2=CH2 + H2OC2H5OH
The source of ethene is crude oil. But as
the crude oil runs short, a major problem
arises: the shortage of ethanol. That’s why
the scientists have been searching for an
alternative way of production of ethanol.
This knowledge has stimulated interest in
the fermentation of sugars. It is usually a
microbial fermentation, as it will only
work directly with the sugar.
The sugars which are used for the
fermentation are obtained from two
components of a plant: starch and
cellulose. The energy for the reaction is
usually provided by sun using
photosynthesis.
There is only one more thing you need to
transform sugar to ethanol: enzymes, or, in
our case, yeast cells.
Yeast cells are eukaryotic micro-organisms
and are classified in the kingdom of Fungi.
There are about 1500 species described
and these organisms dominate the fungal
diversity in the oceans. Yeasts do not
require light to grow, but do use sugar as a
source of energy.
If oxygen is present, yeast cells will use it
and break down sugars to CO2 and H2O. In
an oxygen free environment, yeast will use
an alternative pathway. This pathway
doesn’t require oxygen. The end products
are CO2 and C2H5OH (ethanol).
All in all there are three pathways for yeast
fermentation in a glucose solution:
1. The fermentation, which occurs at high
glucose concentration or in an oxygen free
environment. The equation for this reaction
is:
C6H12O62C2H5OH + 2CO2
2. The oxidation of glucose, which occurs
when the glucose concentration is low
(below 50 mg/L) in an aerobic
environment. The equation for the
oxidation is:
C6H12O6 + 6O26CO2 + 6H2O
3. The oxidation of ethanol, which occurs
when you have a very limited supply of a
yeast culture. The equation of this reaction
is:
C2H5OH + 3O22CO2 + 3H2O
In our research the first pathway is the
most interesting, because we use an
oxygen free environment and a high
concentration of glucose in the solution.
This means that the first reaction will take
place:
C6H12O62C2H5OH + 2CO2
Some researches indicate that the optimal
pH for an anaerobic yeast fermentation is
an acidic pH. This raises the question:
Which pH will be optimal for the
production of ethanol and CO2 as the end
products of yeast fermentation?
We expect that the optimal pH for yeast
fermentation is lower than 5,0. Yeast cells
have enzymes, and enzymes work best at
(slightly) acidic pH’s.
Experimental design and approach
We prepared 2,837 L of a 20% sugar
solution in distilled water. Then 8
erlenmeyer flasks of 0,5 L each, were filled
with the sugar solution for 80% full. The
other 20% was acid. Two bottles were
labelled with pH 2,8; two bottles with pH
4,4 and so on. We added 5 grams of
baker’s yeast to each flask. After that we
weighed and labelled all the balloons. Then
each balloon was fit over the neck of a
flask.
were averaged and the deviation
determined and graphically presented.
Results
After two hours we observed that the
balloons were beginning to fill in with
CO2. Fourteen days later all the balloons
were puffed up.
Table 1 presents, in duplicate, the mass (in
grams) of the released CO2 gas at different
pH’s (2,8; 4,4; 6,0 and 8,4). The table
presents the averaged masses of the
released CO2 gas in grams and its
deviations at the various pH’s.
pH
Balloon 1,
mass CO2
2,8
0,04
Balloon 2,
mass CO2
(g)
0,03
4,4
6,0
8,4
0,03
-0,01
0,02
0,03
0,03
0,02
(g)
Averaged
mass CO2 (g)
0,035 ±
0,005
0,03 ± 0,0
0,01 ± 0,02
0,02 ± 0,0
Table 1: Release of CO2 (in grams) and averaged
release of CO2 (in grams) at pH 2,8; 4,4; 6,0 and
8,4.
Figure 1 shows the averaged measured
release of CO2 (in grams) when yeast cells
grow at the pH 2,8; 4,4; 6,0; 8,4.
0,04
0,035
Averaged mass (g)
0,03
0,025
0,02
0,015
0,01
0,005
0
0
2
4
6
8
pH
We left the flasks with the balloons for 14
days in a room temperature. After 14 days
we tied off the balloons and reweighed
them. The difference in masses within each
set of balloons after and before
fermentation was calculated. For each pH
the masses pH released CO2 (in grams)
Figure 1: Averaged measured release of CO2 (in
grams) versus pH.
Data analysis
As presented in table 1, the average
maximum amount of released CO2 (in
grams) in the fermentation process is
10
0,035g. This is equal to
0,035/44,01=7,95*10-4 mol CO2.
Discussion and conclusion
The observation that all the balloons grow
and puff up after a couple of hours
indicates that in all flasks baker’s yeast
culture was growing and produced CO2
gas.
Table 1 shows that the averaged
production of CO2 gas was highest at pH
2,8. So this is the optimal pH for yeast
fermentation.
Looking critically at our experimental
procedure and approach we see that in all
sets of experiments we considered the
same independent and dependent variables
and we kept the same variables constant.
So, the problem lies in the possibility that
we couldn’t keep all the control variables
constant during 14 days.
Looking at our approach we can’t say for
sure that our measuring instruments were
very good and reliable. One of the results
was impossible. The weight of the release
of CO2 at pH 6 balloon 1 was negative.
This means that the available measuring
instruments were not reliable.
Evaluation
During our research we had a few
difficulties with our set up. The first
problem was, when we fit the balloon over
the neck of the flask there was a possibility
for the air to escape. The second problem
was, that we couldn’t fit all the balloons
over the neck of the flask at the same time.
The last problem was, that some of the
carbon dioxide gas was still in the flask
when we tied off the balloons. This means
that not all the gas was measured.
One possible improvement could be using
better and more reliable measuring
instruments.
As we were doing this research, a couple
of questions arose:
Is it necessary to keep the temperature of
the environment constant?
Or, does the produced ethanol have a
negative effect on the growth of yeast
cells?
Bibliography
1) Slaa, J., Gnode, M., & Else, H. (2009).
Yeast and fermentation: the optimal
temperature. Journal of Organic
Chemistry: Chem. Dut. Aspects 134
2) http://en.wikipedia.org/wiki/Ethanol_fu
el
3) http://en.wikipedia.org/wiki/Yeast
4) http://www.yobrew.co.uk/fermentation.
php
5) http://wiki.answers.com/Q/How_does_
pH_effect_yeast_fermentation