Fermentation Investigation

* Carbon dioxide gas is soluble in water - some is not included in the results

* Surface tension of meniscus pressure - carbon dioxide gas may have been compressed

* Growth of yeast culture - the no. of yeast cells are not the same

* Intoxication by ethanol - some yeast cells may be intoxicated and die

* As more CO2 is collected into the tube, there is less yeast inside it

* The end of the Durham tube is hemispherical - no linear relation of rate of reaction at the start

* Measurement of the length of the meniscus - a ruler was used, and it was not in contact with the Durham tube

* The tubes have to be lifted out of the water bath to take measurements - the environmental temperature could have affected the rate of fermentation every time measurements are taken

Some of these errors can be improved if the experiment was to be carried out again. The Durham tube could be substituted with similar equipment that has flat end, instead of being hemispherical, so the change in height of the meniscus would go in linear proportion. Perhaps a scaled glass rod could be used to instead of a ruler. With this, the readings would be taken adjacently to the tube, rather than outside the test tube itself

If a further investigation is to be carried out, the temperature of the environment and the types of the sugar used are the interesting fields to head forth towards.

Background knowledge

FERMENTATION is the breakdown of sugars by bacteria and yeast using a method of respiration without oxygen (anaerobic respiration). It involves a culture of yeast and a solution of sugar, producing ethanol and carbon dioxide with the aid of the enzymes. This is an 8-10 step process requiring different enzymes each time, but it can be simplified like this.

This process can be slowed down by DENATURATION of the enzymes at a certain temperature.

All the ENZYMES are protein chains of amino acids. They exist in the form of ?-helix structure with hydrogen bonds holding the pitches together. On the amino acid molecules, there is R a group. They react with each other to form peptide bonds, transforming the chain into a 3-dimensional structure. Along the chain there are active sites where interaction between the enzyme and the substrate happens. These sites are sensitive to heat, like the hydrogen bonds that hold the 3D molecule together. When heat is applied to the enzyme, energy is given into the molecule. The active sites deform and the hydrogen bonds break, denaturing this enzyme. It would not be able to function as usual, and this is not reversible. This is called DENATURATION. The 3D ?-helix structure would breakdown and the active sites would change in shape; they would not be able to accommodate the substrate any more. The analogy of this is to compare a key to a keyhole. If the keyhole has changed, the same key would not fit in any more, and the lock would not be unlocked. The same thing happens here, and fermentation could not continue after this has occurred. Also when the temperature is too low, the enzymes would not work because there is not enough energy for activities to happen.

Concentration of the solutions should make the yield of fermentation higher as they become more concentrated. This would be because the sugar molecules are more abundant as the concentration increases, so the probability that the enzymes come into contact with the molecules is higher. However, more concentrated solutions mean less osmotic pressure.

OSMOSIS is the movement of solvent such as water through a semi-permeable membrane separating solutions of different concentrations. The solvent passes from the more dilute solution to the more concentrated solution until the two concentrations are equal. Many cell membranes behave as semi-permeable membrane, and osmosis is vital mechanism in the transport of fluids in living organisms - for example, in the transport of water from the roots up the stems of plants. The region that has less percentage of water is said to have less OSMOTIC PRESSURE, and vice versa. Water permeates the cell wall into the region with low water potential (osmotic pressure) until the osmotic pressure is the same on both sides.

Water is vital to living organisms, and having too high concentration of sugar risks osmosis problem, and vice versa, as a cell cannot work properly when there is too much water in it.

Analysis

First of all, it is apparent that fermentation is a process that occurs in a linear fashion: the lines in which the reactions occur on are straight. The beginning of the best fit lines are omitted because they are not in linear relation to each other, as the equipment used was not in linear proportion of surface are to volume. Conspicuously, the solution with 100% sugar concentration has worked best of out of the three. A comparison of these can be made as the gradients of these lines are calculated. They would represent the rate in which the reactions are taking place. To make the rates comparable, the gradients are taken from the same point of the graph - between 10 and 20 minutes. Thus it is as follow:

Conc. Rate

200 0.92

50 1.38

00 1.70

The rate is the movement of the meniscus in mm per minute. There seems to be a proportion that establishes a connection between them. If more data of other concentration are acquired, perhaps a formula could be derived from the results.

There are explanations for this. At 200% concentration, the osmotic pressure inside the cells of the yeast is higher in comparison to the sugar solution outside, so water is lost into the solution - through osmosis. The cells now contain less water potential. Although the sugar content in the solution is high, increasing the probability of interaction between the enzymes and the sugar molecules, the yeast cells are now less capable at carrying out reactions, because it has less water potential than before. They become less efficient at fermentation. After all it is the osmotic pressure that is important. It was a misfortune that there was not enough time for the 50% concentration fermentation to be done, but it is evident that 100% concentration would work best. This concentration is quite likely to have equal osmotic pressure, or water potential, in the mixture. Although 50% concentration solution would have much higher water potential in the cells, the abundance of sugar molecules would not be high enough to bear as good productivity as the 100% one.

The vital key point here is that the osmotic pressure has to reach equilibrium, which is closer to 100% concentration, of 3g of sugar per 100ml of water, than any others.

The results obtained are similar to the prediction. If the concentration theory is to be considered, it is a failure, because osmotic pressure actually takes major part in this experiment. Apart from that, all went well in accordance with the secondary prediction.

Results of fermentation with different concentration of sugar solution

200% concentration (6 g per 100 ml) Temp. - 30 C

Time in Distance extended by the meniscus in mm

minutes 1st 2nd 3rd average

0 0.0 0.0 0.0 0.0

5 12.0 11.0 10.0 11.0

0 15.0 15.0 14.0 14.7

5 20.0 20.0 19.0 19.7

20 25.0 26.0 24.0 25.0

25 30.0 30.0 30.0 30.0

30 34.0 35.0 33.0 34.0

50% concentration (4.5 g per 100 ml) Temp. - 30C

Time in Distance extended by the meniscus in mm

minutes 1st 2nd 3rd average

0 0.0 0.0 0.0 0.0

5 11.0 10.0 11.0 10.7

0 16.0 16.0 17.0 16.3

5 23.0 24.0 23.0 23.3

20 30.0 30.0 30.0 30.0

25 37.0 37.0 36.0 36.7

00% concentration (3 g per 100 ml) Temp. - 30C

Time in Distance extended by the meniscus in mm

minutes 1st 2nd 3rd average

0 0.0 0.0 0.0 0.0

5 12.0 12.0 12.0 12.0

0 20.0 18.0 19.0 19.0

5 28.0 27.0 28.0 27.7

20 37.0 37.0 36.0 36.7

Rate of fermentation

Concentration (%) Rate in mm/minute

200 0.92

50 1.38

00 1.70

Planning

Aim

To fine whether different type of sugar solution effect the rate of fermentation. How does it effect the rate and which type of concentration works best.

Factors affecting the rate of fermentation

* Temperature

* Type of sugar

* Concentration of sugar

* Amount of sugar

The factor I have chosen is to vary is the concentration of sugar. The concentration will be 200%, 150%, 100% and 50%.

Safety precaution

* Hot bath is dangerous and you might boil you hand therefore care must be taken.

* Yeast culture is not every good to your eye so care must be taken. Therefore safety glass might need to be worn.

* Any equipment that is broken is dangerous for example broken glasses because it could cut you.

Prediction

What I have predicted the out come of the result will be is that the amount of carbon dioxide given off will increase as the concentration of sugar increase. The rate of fermentation will increase proportionally as concentration increase but it will eventually stop.

Apparatus

* Test tubes - for containing some of the yeast culture for the experiment.

* Durham tubes - for finding the amount of carbon dioxide give off.

* Pipette - for putting the yeast culture into test tubes and Durham tubes.

* Yeast culture - for testing the rate of fermentation.

* Ruler - for measuring the volume of carbon dioxide given off.

Planning

Methods

. Transfer 2-3 cm3 of the 200% sugar concentration culture into each test tube.

2. A small Durham tube has to be filled with yeast culture of the same concentration and inverted in each test tube.

3. The test tubes are then put into a hot bath of 30???C.

4. The time at which each Durham tube is set up is note down. At timed, (every five minutes), the length of the bubble collecting inside each tube is measured.

5. The results are taken for 20 minutes.

6. Repeat steps one to five but this time with different concentration of sugar solution, which are 150%, 100% and 50%. But as the time is not available I could only do 200%, 150% and 100%

7. Each experiment of different concentration will repeated three times and find the average results.

Background knowledge

Fermentation

The experiments I have done are based on fermentation. Fermentation is the process of making alcohol and making bread. In fermentation the two important substances are the yeast and sugar in the form of glucose. The process turns glucose in to ethanol, alcohol, and carbon dioxide. It can be written as:

Glucose ethanol + carbon dioxide

C6H12O6 (aq) 2C2H5OH (aq) + 2CO2 (g)

The whole takes about 8 or 10 steps long for it to complete. In each of the steps there is a need of ENZYMES. Enzymes have a 3D structure of ??helix. In every layer, pitches, they are joins by hydrogen bonds, which can be easily broken down. These enzymes are used for breaking down the sugar molecules. This will release energy, which is necessary for the yeast cells to use for multiplying. The reaction could be written as:

Substrate + enzyme product + enzyme

The substrate stand for sugar solution and the products are ethanol, carbon dioxide and energy. All of the enzymes are form of protein and protein molecules are amino acids. Therefore as the temperature increases the rate also increase. The rate of reaction could be calculated by:

The increase in movement of meniscus per 5 min

The increase in time per 5 min

On each enzyme there are active sites which can accommodate certain substrate. Active sites break down substrate into small pieces. When energy is produced it breaks down the structure of enzymes thus making them unable to break down the substrate. This is called DENATURATION.

Evaluation

From the experiment I have done I did the best I could to produce the out come of the results to be perfect enough of the conclusion. Though out the experiment the out come of the results have been well enough and I think that the way I did the experiment is the best possible that I could do.

The results collected have come out to be well more than I expected. When the results are plot on the graph it could be seen clearly that not much is wrong with them. Also the method I have carried out for the experiments I think it is the best way that I could come out with. This is because I use the method of repeating the experiments three time to get as accurate results as possible by taking the average result form the repeated ones.

Errors

The errors I think that might occurs during the experiment could be of that:

. The Durham tubes used has on the end a dome shape, which is an uncontrolled variable. The product will cause a great increase but as the more of cross section increase down the tube the product will be less varied. Therefore there is a non-linear relationship between volume of gas produced and height increase.

2. The yeast culture was not shaken at the beginning therefore the yeast might not be spread out through out the culture.

3. The surface tension of Durham tube might cause some change because as the meniscus descend it might stick to the side of the tube and when the pressure builds up and trying to drag it down the meniscus will not come down steadily. Therefore the result might not have come out well.

4. The carbon dioxide might have dissolve in the solution causing the amount of carbon dioxide to be lesser.

5. The tube was taken out of the water bath to measure therefore the temperature will not be constant.

After all what I have in mind of improving the experiment is that to change the shape of the Durham tube from hemispherical shape end to flat end. This is because the results that are collected can be even more accurate. Also what I would do more for my experiment is that to do the same but change the factor from sugar solution to change in temperature and also both of the factors together.

Analysis

Form the graphs what I have found out was that on the comparison of volume of CO2 produced of different concentration of sugar solution the concentration that has the most CO2 produced is 100% and the least is 200%. This is because that supposing the more concentrated the solution is the more ethanol and CO2 will be produced because more can be react with the enzymes. But in fact it is not because the osmotic pressure which is that the water will flow from less concentrated solution into more concentrated solution. This result for the yeast cells to loose water to the solution, which makes the yeast less efficient.

For the rate of fermentation the one that has the fastest rate is 100% and the least is 200%. What I could also noticed from this graph is that the rate increases proportionally as concentration increase. The rate of reaction, in this case, is the movement of the meniscus per minute. The rate of reaction depended on the surrounding temperature and the concentration of the solution therefore the more concentrated the solution is the faster the rate. For this graph though it is not so accurate because that the rate in the beginning of the experiment is ever so fast that it changes the look of the graph so much. Therefore I ignore the first volume of the result because if I have included it into the line the gradient of the graph will not be accurate enough and this will result in the change of the rate of reaction. The rate of reaction table could be seen in the obtaining evidence along with the other results. The formula for calculating the rate of the reaction is:

The change in volume of carbon dioxide (y)

The change in time per minute (x)

It is a shame that I did not have enough time to do the rest of the experiment such as the sugar concentration of 50% because I do not have enough time to complete it. Though I have not done the other experiments I still have enough of the results to conclude the experiment.

Although I did not do 50% sugar concentration I could still say that the best sugar concentration will still be 100% because it has the nearest osmotic pressure on both the solution and the yeast cells. The main conclusion here is that the most important factor is the osmotic pressure that effects the whole experiment.

Due to my prediction what I have predicted was nearly all right except on which was that the amount of carbon dioxide given off will increase as the concentration of sugar increase was wrong because that it is exactly the opposite. Except that the other prediction is the same as the result.

00% concerntration with 3 g per 100 mil. Temp 30 C

Time (min) Movement of meniscus (cm)

st 2nd 3rd Average

0 0 0 0 0

5 11 12 12 11.7

0 20 20 20 20

5 27 28 28 27.7

20 36 37 37 36.7

Rate of fermentation on different concentration of sugar

Concentration (%) Rate (mm/min)

200 1.34

50 1.61

00 1.87