Sri Ram Dayal Khemka Vivekanandha

Vidhyalaya
Junior College

NAME: SAKTHIVELAYUDHAM. G
CLASS: XII A
ROLL NO:

CHEMISTRY INVESTIGATORY PROJECT

RATE OF FERMENTATION OF FRUIT JUICES
CONTENTS:
1) INTRODUCTION
2) MAIN USES OF FERNTATION
3) LIMITATIONS
4) PASTEUR SALT
5) PRODUCTION OF CITRIC ACID
6) EFFECT OF OXYGEN
7) ADDITION OF YEAST
8) EXPERIMENT
9) OBSERVATION
10) RESULT
 INTRODUCTION
Fermentation is typically the conversion of carbohydrates to alcohols
and carbondioxide or organic acids using yeasts, bacteria, or a
combination thereof, under anaerobic conditions (absence of oxygen)
by the action of enzymes. Enzymes are complex organic
compounds, generally proteins. They are highly specific with
regard to their substrates. Fermentation in simple terms is the
chemical conversion of sugars into ethanol. Ethanol
fermentation, also referred to as alcoholic fermentation is the
biological process in which sugars such as glucose, fructose, and
sucrose are converted into cellular energy and thereby produce
ethanol and carbon dioxide as metabolic waste products. All
ethanol contained in alcoholic beverages is produced by means of
fermentation induced by yeast. Wine is produced by fermentation of
the natural sugars present in grapes and other kinds of fruit .Ethanol
fermentation occurs in the production of alcoholic beverages and
ethanol fuel, and in the leavening of bread dough. Fermentation is used
in preservation techniques and in production of foods such as yogurt,
cottage cheese (paneer), dhokla, idli, chocolates, cheese etc.
‘Fermentation’ has been derived from the Latin word fervor,
which means ‘to boil’, as during fermentation, there is a lot
of frothing in the liquid due to evolution of carbon dioxide.
This gives it the appearance as if it is boiling! Yeasts are unicellular
eukaryotic microorganisms classified in the kingdom Fungi, Yeast size
can vary greatly depending on the species, typically measuring
3-4 μm in diameter, although some yeasts can reach over 40 μm. Most
yeasts reproduce asexually by mitosis, and many do so by an
Asymmetric division process called budding. Yeasts do not form a single
taxonomic or phylogenetic grouping. The term yeast is often
taken as a synonym for Saccharomyces cerevisiae. Natural
fermentation precedes human history. The earliest evidence of
winemaking dates from eight thousand years ago, in Georgia,
in the Caucasus area. Seven-thousand-year- old jars containing the
remains of wine have been excavated in the Zagros Mountains
in Iran. There is strong evidence that people were fermenting
beverages in Babylon circa 3000 BC, ancient Egypt circa 3150 BC,
pre-Hispanic Mexico circa 2000 BC, and Sudan circa 1500 BC.
Ancient fermented food processes were developed long before man
had any knowledge of the existence of the microorganisms involved.
When studying the fermentation of sugar to alcohol by yeast, Louis
Pasteur concluded that the fermentation was catalyzed by a
vital force, called “ferments”, within the yeast cells. The “ferments”
were thought to function only within the yeast cells. The “ferments”
were thought to function only within living organisms. Nevertheless,
it was known that yeast extracts Yeast extract is the name
given to processed yeast products made by extracting the cell
contents (removing the cell walls)) can ferment sugar even in the
absence of living yeast cells. While studying this process in 1897,
Eduard Buchner found that sugar was fermented even when
there were no living yeast cells in the mixture; by a yeast secretion that
he termed zymase, i.e., fermenting activity of yeast is due to
active catalyst of biochemical origin. In 1907 he received the Nobel
Prize in Chemistry for his research and discovery of “cell-free
fermentation.

MAIN USES OF FERMENTATION

The primary benefit of fermentation is the conversion of sugars and
other carbohydrates, e.g., converting juice into wine, grains into
beer ,carbohydrates into carbon dioxide to leaven bread, and sugars in
vegetables into preservative organic acids. Food fermentation has been
said to serve five main purposes:
1) Enrichment of the diet through development of a diversity of
flavors, aromas , and textures in food substrates.
2) Preservation of substantial amounts of foods through lactic
acid, alcohol, acetic acid, and alkaline fermentations
3) Biological enrichment of food substrates with protein, essential
amino acids, essential fatty acids, and vitamins Elimination of anti-
nutrients
4) A decrease in cooking time and fuel requirement

Limitations
0ne of the limitations of fermentation as a process is its requirement
for multiple reagents. Secondly, in many cases the time taken is quite
long and this creates a need for catalyst. Without catalysts, the reaction
is extremely slow. The limitation of our project is the slight error in the
result and the project is limited to the fermentation of the juices with
Baker’s yeast and not under normal conditions i.e. without adding
Baker’s yeast. Owing to the different criterion on which the rate of
fermentation depends, if the experiment is not carried out in the
optimal temperature range, the rates will turn out to be different than
the actual rates of the juices that have been taken. It is not possible
to get the exact theoretically estimated value due to impurities
in the reagents as well as the compounds. Another point to be noted is
that the rates calculated from this experiment is just one case and this
can’t actually access the rate of fermentation of the fruit. An average
needs to be taken to access its actual value.

Pasteur’s Salt
Pasteur’s salt solution is prepared by dissolving ammonium tartrate,
10.0 g; potassium phosphate, 2.0 g; calcium phosphate, 0.2 g;
and magnesium sulphate, 0.2 g dissolved in 860 ml of water. The
Pasteur’s salts in solution act as a buffer to any acids the yeast may
create. Since yeast only converts sugar (most likely sucrose or glucose)
to ethanol under anaerobic conditions, and it is unreasonable to
assume that there will be no oxygen present in the laboratory, some
acetic acid is created as a result. The Pasteur salts act as buffers to the
acidity so that the proteins in the yeast do not become denatured.

Production of Citric acid

Citric acid is a useful product in both the food and
pharmaceutical industries; it is used in food as a preservative and to
produce an acidic, sour taste in soft drinks and other beverages. In the
pharmaceutical industry it can be used as buffering agent and to clean
equipment. Citric acid is formed by the fermentation of a molasses
substrate by the fungus Aspergillus Niger.The biochemical pathway
involved includes the production of pyruvate in glycolysis, followed by
its conversion to citric acid via the condensation of acetyl co-enzyme A
and oxaloacetate.

Effect of oxygen
If oxygen is present, some species of yeast will oxidize pyruvate
completely to carbon dioxide and water. Thus, these species of
yeast will produce ethanol only in an anaerobic environment.
However, many yeasts such as the baker’s yeast Saccharomyces
cerevisiae , or fission yeast Schizo saccharomyces pombe prefer
fermentation to respiration. These yeasts will produce ethanol even
under aerobic conditions. Hence the rate of fermentation varies. The
fermentation process is not only complex but always in a state of flux.
Process, we are therefore in a situation to always be adaptive and
reactive to these changes so that throughout the fermentation process
we are always sustaining the conditions in a narrow window of
optimal fermentation conditions. In order to help us do this we need
to know fermentation kinetics. When we talk about fermentation
kinetics we are talking about fermentation models. Kinetics and
modellings are very useful to us as tools to make fermentation
predictions and enhancing our experimental designs to be more
focused to the specific problems such as the rate limiting steps or
product inhibition. The study of fermentation kinetics helps us by
providing clear quantitative data for us to understand the process and
improve the process accordingly. Peering into observation ports
might be good advertising gimmick for fermentation technology
but do not really help much in understanding the process or even to
control and predict the fermentation outcome. Subjective observations
will rarely help in producing optimum fermentation process and
thus affect profitability studies and making decisions.
Its numbers that count!
Thus the importance of the study of fermentation kinetics or models.
The first step in the study of fermentation kinetics is to
understand the various processes involved in the whole process. Such
questions such as inputs and outputs, the metabolic pathways involved
and type of products or side products formed. The various individual
reactions involved and what factors control the metabolite levels. Then
only after all the relevant data are obtained do we start formulating the
models.

Addition of yeast
In wine making, yeast is normally already present on grape
skins. Fermentation can be done with this endogenous “wild
yeast,” but this procedure gives unpredictable results, which depend
upon the exact types of yeast species present. For this reason, a pure
yeast culture is usually added, this yeast quickly dominates the
fermentation. Baker’s yeast is the common name for the strains of
yeast commonly used as a leavening agent in baking bread and bakery
products, where it converts the fermentable sugars present in the
dough into carbon dioxide and ethanol. Baker’s yeast is of the species
Saccharomyces cerevisiae , which is the same species commonly
used in alcoholic fermentation, and so is also called brewer’s yeast.

Experiment
AIM:
To study rate of fermentation between a fruit juice and vegetable juice.
Requirements:
Conical flasks (250 ml), test tubes and water bath, Apple juice and
Fehling’s solution.
Theory:
Louis Pasteur in 1860 demonstrated that fermentation is a purely
physiological process carried out by living micro-organism like yeast.
This view was abandoned in 1897 when Buchner demonstrated that
yeast extract could bring about alcoholic fermentation in the absence
of any yeast cells. He proposed that fermenting activity of yeast is due
to active catalysts of biochemical origin. These biochemical catalyst are
called enzymes. Enzymes are highly specific. A given enzyme acts on a
specific compound or a closely related group of compounds.
Fermentation has been utilized for many years in the preparation of
beverages. Materials from Egyptian tombs demonstrate the procedures
used in making beer and leavened bread. The history of fermentation,
whereby sugar is converted to ethanol by action of yeast, is also a
history of chemistry. Van Helmont coined the word ig salt in 1610 to
describe the bubbles produced in fermentation. Leeuwenhoek
observed and described the cells of yeast with his newly invented
microscope in 1680. The fruit and vegetable juices contain sugar such
as sucrose, glucose and fructose. These sugars on fermentation in the
presence of the enzymes invertase and zymase give with the evolution
of carbon dioxide. Maltose is converted to glucose by enzyme maltose.
Glucose is converted to ethanol by another enzyme zymase.
Invertase:
C12H22O11 + H2O C6H12O6 + C6H12O6
Sucrose glucose fructose

Zymase:
C6H12O6 + C6H12O6  2C2H5OH+2CO2
Glucose fructose ethanol

Diastase:
2(C6H1005)n + nH20 nC12H22O11
Starch maltose

Glucose is a reducing sugar and gives red colored precipitates with

Fehling’s solution, when warmed. When the fermentation is complete,
the reaction mixture stops giving any red color or precipitate with
Fehling solution.

Procedure:
1. Take 5.0 ml of any fruit juice and vegetable juice in a clean seperate
250 ml conical flask and dilute it with 50 ml of distilled water.
2. Add 2.0 gram of Baker’s yeast and 5.0 ml of solution of Pasteur’s
salts to the above conical flask.
3. Shake well the contents of the flask and maintain the temperature of
the reaction mixture between 35-40°C.
4. After 10minutes take 5 drops of the reaction mixture from the flask
and add to a test tube containing 2 ml of Fehling reagent. Place the test
tube in the boiling water bath for about 2 minutes and note the colour
of the solution or precipitate.
5. Repeat the step 4 after every 10 minutes when the reaction mixture
stops giving any red color or precipitate.
6. Note the time taken for completion of fermentation

Observation:
Volume of fruit juice taken =
Volume of distilled water added =
Weight of Baker’s yeast added =
Volume of solution of Pasteur’s salts =
TIME(IN MINUTES) COLOUR OF COLOUR OF
REACTION MIXTURE REACTION MIXTURE
WITH FEHLINGS WITH FEHLINGS
SOLUTION IN CASE OF SOLUTION IN CASE OF
FRUITJUICE VEGETABLE JUICE
10
20
30
40
50
60

Result:
Rate of fermentation of ___________ juice is greater
than___________ juice.

Bibliography:
1) Pradeep’s chemistry
2) NCERT chemistry
3) www. Icbse.com