# Yeast and fermentation: the optimal pH balance

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```					   Yeast and fermentation: the optimal pH value
written by: Rosanne de Roo and Nicole Zomerhuis
Submission date: April 2010
Ichthus Lyceum, Driehuis, The Netherlands

Summary
We depend on oil. Oil and other fossil fuels are running out. We need to find a solution to this
problem. Now the focus is on bio-ethanol. This substance can be extracted from glucose if it is done
using the correct method.
Glucose can be fermented into ethanol and carbon dioxide by yeast cells if the reaction chamber is
oxygen free. In our inquiry we used leture yeast cells from dr Oetker. We wanted to find out what
the optimal value of pH is for the fermentation reaction. We filled twelve flasks with 18 grams of
granulated sugar. We used solutions with a pH 3,5 , 4,5 , 5,5 , 6,5 ,7 and 7,5. Then we let the yeast
do its work for 3 days in an environment of 35 ºC. We expected the optimal ethanol levels to be at
pH 7. We got very different results out of our experiment. We saw that pH 7 was in fact very low. We
think this is because of mistakes. All the flasks were in duplicate.

Introduction                                          Not only oxygen can have an effect on the
If you want to create bio-ethanol you need to         amount of ethanol that can be produced. Also
use yeast. Yeast is a kind of fungi that helps        things like pH of the solution, temperature,
the glucose react into ethanol and a gas. This        kind of glucose, amount of glucose and
gas is carbon dioxide. In the following               amount of yeast have an effect.
chemical equation you see the reaction that
the yeast helps to do.

When you look more closely you see that the
reaction is completely without oxygen. It is
important to let the yeast react in an oxygen         Figure 1: Structure of a glucose molecule
free environment. If you do let oxygen slip
into the reaction chamber the following will          In our experiment we want to investigate
happen:                                               what the optimal pH is to let the yeast react
in. Our research question is:
What is the optimal pH for a solution with 18%
The glucose will react with the oxygen and will       glucose (sugar) and leture yeast cells (dr.
be converted into carbon dioxide and water.           Oetker) in 35 ºC in an oxygen free
The ethanol that has been formed will also             environment ?
react into water and carbon dioxide.                  We think that the optimal pH for the reaction
will be pH 7. We think this because pH 7 is
neutral. When the pH is lower than pH 7 the
It is essential that the reaction in this             solution is an acid. When the pH is higher than
experiment is in a completely oxygen free             7 the solution is a base. In acid and base
environment . If not, there won’t be any bio-         solutions the ions are very active. They jump
ethanol formed and you will have useless              from atom to atom. We think that this isn’t
substances or less ethanol.                           beneficial for the yeast cells. We think that
this will affect the cells and the yeast will not
work in the optimal way. Less ethanol will be
produced. When the solution is pH 7 there will
not be any active ions so the yeast
cells will not be affected by the ions. At this
pH the optimal amount of ethanol will be
formed because the yeast can work in his
optimal fashion.                                     Figure 2: Distillation apparatus

Experimental design                                  In the second part we started the distillation
In an experiment it is very important to keep        process. We poured 100 mls out of the flasks.
all variables constant. To investigate if one of     Then we heated 100ml up to 100°C. When the
the variables can be changed to get the              temperature had reached this point we had to
optimal amount of ethanol out of the solution,       stop the distillation process. If you go on past
you need to change one of the variables. In          100°C, you will get water into your distillate.
our experiment we changed the pH of the              That is not what you want. You only want the
solution which contained the yeast and               ethanol to come out of the solution. After that
glucose. The other important variables are:          we let the ethanol pour into a volumetric
temperature, kind of glucose, amount of              flask. We filled it up to 10 ml with water.
glucose and amount of yeast. We kept these           Because of this you could calculate the
variables constant. We also made the                 amount of ethanol in mls.
experiment as oxygen free as possible.
Because we kept all the important variables
First we chose to fill twelve flasks each with       constant the experiment seems quite
18,00 grams of sugar. Then we put exactly            accurate. The significant of the results is quite
7,00 grams of leture yeast cells from dr.            high. It is 3 significant numbers.
Oetker in the solution.
The bottles were filled with different solutions     Results
of pH values. We used solutions with a pH 3,5        Now we are going to give our results. We will
, 4,5 , 5,5 , 6,5 ,7 and 7,5. The bottles were       discuss the observations and the results.
filled to 271,5 mls. Then balloons were put
over the solutions to keep most of the oxygen        Observations
out. Still there was a bit of oxygen in the flask.   When we did the first part of our experiment
We decided to do this because the                    there were not very many observations. We
fermentation reaction makes the substance            put the yeast and sugar into the solution. You
rise. If we had filled up the flask to its           couldn’t see anything happening the
maximum, the solution would have poured              substances just lay there.
into the balloon and our experiment would            In the second part of our experiment there
have been inaccurate. After that we shook            were more observations. When we got our
each flask for one minute.                           bottles they smelt of unbaked bread. It really
Every pH solution flask was done in duplicate.       was a hideous smell. All twelve mixtures
This means that we had two flasks with the           looked similar, they had a pretty dark brown
same pH value. In total we had twelve flasks.        colour. Yet there were a few notable
We labeled all the flasks and put them into a        differences. In the bottle with a pH of 3,5
bath that had a constant temperature of 35           there was no mixture. The yeast was on the
°C. We waited for three days and then we             bottom of the bottle and the rest of the mix
started the second part of our experiment.           was floating above it. The bottles with a pH
between 6,5 and 7,5 were much lighter in
colour and the smell was worse than in the
other bottles.
Data and data analysis
In the following table you can see how much
ethanol was formed.
pH   Grams   Volume Massa %
while we understood the yeast was dead. The
of      %                                 low pH had killed the yeast cells.
ethanol                                   Because the smell was much worse in the
bottles with a pH between 6,5 and 7,5 we
3,5 6,7         8,38       6,79                think that in those bottles the yeast
fermented the best. If you look at the results
4,5 5,5         6,88       5,56                you can see that true.

5,5 9,5         11,88      9,68                Another striking thing is the reduction at pH
7, we believe that this is possibly an error. It
6,5 9,2         11,5       9,37                does not coincide with the trend in the graph.
7,0 5,7         7,13       5,77                our test again, especially the pH 7. This way
we will know if it was our mistake , or if it is
7,5 12,75       12,75      10,41               really like this.
The answer to our experiment question is that
7,5 pH is the optimal pH to let a fermentation
As you can see the lowest pH values (3,5 and    process do its work.
4,5) have the lowest amount of pH. Then pH 7    We believe that our experiment was a
has a very low number of ethanol. The most      success.
ethanol was formed at 7,5 pH.
Evaluation
As you can see in our results the amounts of
ethanol are weird. It varies a lot and that
think a lot has gone wrong.
First we think that the solutions were not
oxygen free enough. Because of this, a lot of
water and carbon dioxide were formed
instead of ethanol and carbon dioxide.
Secondly we think the distillation process had
its faults. We think we heated the solutions
too much and because of this, water was
distillated. In the distillate we got ethanol but
also water which we absolutely did not want
to happen.
Thirdly we think we did not clean the
In the following graph you can see the amount   distillation apparatus good enough so water
of ethanol that was formed.                     and ethanol were still in parts of the
apparatus of our previous experiments.
Discussion and conclusion                       If we make sure that in future experiments we
When we did our research on the internet, we    fill the flasks up to the top and try to find
saw that the best pH for fermentation was       another way to make sure nothing comes out
3,0. We thought that was wrong and still        of the flasks, we have solved one problem.
believed that the optimal pH was 7 . Because    The second problem we can solve just by
the school said a pH of 3,0 was too low, we     being more accurate and stopping the process
had to use a pH of 3,5. When we saw the         at the right time. The last problem can be
results we first did not understand. After a    solved easily. We just have to clean the
apparatus and wait till all the fluids have run
out.
We can improve our inquiry in many different
ways. If we do that we can say what the
optimal pH value is to let the fermentation
process do its work. If you know what the
optimal pH is, you can also think about setting
up new inquiries. Maybe you can find out
what the best kind of glucose is, or the best
temperature, or the best amount of yeast that
can be used.

If you know all the answers to these
questions, you can get the optimal amount of
ethanol that can be formed by yeast.

Bibliography
250px-Simple_chem_distillation.PNG

mmons/a/a5/Alpha-d-glucose.png

3.http://www.yobrew.co.uk/fermentation.p
hp

mole00/mole00195.htm

5.http://www.chemheritage.org/educational
services/pharm/antibiot/activity/yeast.htm

6.http://herbarium.usu.edu/fungi/FunFacts/
Fermentation.htm
Experimental set-up   ICD7_Rosanne_Nicole

Rosanne:
rosanne_love47@hotmail.com

Nicole : turn_puma@hotmail.com

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