Yeast, which is a fungus, respires anaerobically and anaerobic respiration involves energy release in the absence of oxygen, it therefore requires a hydrogen acceptor which in this case is Methylene Blue, an artificial hydrogen acceptor that accepts the Hydrogen from the sugar solution. Methylene Blue turns colourless when exposed to a reducing agent, therefore if it turns colourless it is implied that the yeast cells are actively respiring.
Hypothesis:
Assuming that yeast cells are alive at 35˚C the optimum temperature can be predicted to be higher than this therefore the lowest temperature at which all the yeast cells are killed can be predicted to be in the range of 50˚C - 60˚C.
In humans, most enzymes have an optimum temperature at roughly 35˚C. Yeast is a fungus so it is not implied that it has the same optimum temperature as human enzymes although some types of fungi are believed to work up until 70˚C, therefore we can assume that yeast respires at 35˚C.
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Apparatus:
Method:
A 10cm3 sample of yeast was taken and put into a boiling tube. Equal volume of sugar solution (sucrose) was added as yeast uses this as a fermentation substrate. This was then left to ferment whilst a water bath was set up for temperatures in the range 40˚- 60˚C. The boiling tube was then added to the water bath and left for 5 minutes in order for the yeast to become accustomed to its new environment. 0.2cm3 of Methylene Blue was added to a test tube and also placed in the water bath with the boiling tube for 5mintutes. After 5 minutes, the Methylene blue was added to the yeast and the colour change was observed. A sample of this solution was then taken using a glass rod and placed on a spotting slide with a glass cover slip. This was then placed under a Light microscope under high power. The number of dead cells and living cells were counted in one field of view and then repeated for four other fields of view. These values were then converted into a percentage and if there was any colour change with the Methylene blue, this was noted. The formula used to work out the percentage for example for living cells would be: number of living cells / total number of cells counted * 100.
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Fair Test/Variables
- A control experiment was set up in order to see the blue colour the Methylene blue will change to when all the yeast cells are killed. This was done by boiling the yeast using a Bunsen burner
- Also, the same was done at 30˚C when the yeast was alive to see what the solution looks like when it turns colourless.
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The temperature had to be kept constant and as there was a 5 minute interval, the temperature may fall therefore would be topped up by hot water from the kettle
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The concentration of Methylene Blue (0.2cm3) added to the boiling tube had to be kept the same and was measured out using a syringe.
Preliminary Results
A preliminary experiment was carried out using the method above.
Safety Precautions
The following safety precautions were taken:
- An overall had to be worn as Methylene Blue stains permanently
- Goggles - as a precaution because of boiling water and Methylene Blue.
- Hair had to be tied back – a precaution for when the Bunsen burner is in use
Extra Care had to be taken as the experiment involves hot water which could result in scalding. Also the equipment being used was made of glass therefore it was important to take necessary safety precautions.
Modifications
The following changes were made to the method after the preliminary results:
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Instead of 10cm3 of yeast and sucrose being used, these amounts could be halved and instead 5cm3 of both solutions could be used.
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Rather than 0.2cm3 Methylene blue being used, 0.1cm3 of Methylene should be used as it is assumed that with a stronger concentration it will take longer to see a colour change. The range of temperatures used should begin from 40˚C-60˚C However once 50˚C is reached, then more accurate temperatures such as 54˚C and 58˚C should be used in order to determine the lowest temperature that yeast cells are killed at more accurately.
The results table for the actual experiment would look like this:
Using the results a graph can be plotted:
This graph could be used to estimate the temperature at which all yeast cells are dead. The following graph shows a predicted curve and shows how this could be obtained.
Bibliography
Biology 1 Textbook - Cambridge
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