Affect of sucrose concentration on the rate of respiration.

02/05/07

Biology coursework

Investigation- Affect of sucrose concentration on the rate of respiration.

Planning

Aim and Background information

The aim of this investigation is to find out how the affect of changing the sucrose concentration affects the rate of respiration of yeast. The reaction can be measured by the amount of carbon dioxide given of by yeast and ethanol is also produced as a result of the reaction.

Yeast (Saccharomyces cerevisiae) is a unicellular fungus, which is frequently used in baking. The precise classification is a field that uses the characteristics of the cell, ascospore and colony. Physiological characteristics are also used to identify species. One of the better-known characteristics is the ability to ferment sugars for the production of ethanol. Budding yeasts are true fungi of the phylum Ascomycetes, class Hemiascomycetes. The true yeasts are separated into one main order Saccharomycetales. Yeasts multiply as single cells that divide by budding (eg Saccharomyces) or direct division (fission, eg. Schizosaccharomyces), or they may grow as simple irregular filaments (mycelium). In sexual reproduction most yeasts form asci, which contain up to eight haploid ascospores. These ascospores may fuse with adjoining nuclei and multiply through vegetative division or, as with certain yeasts, fuse with other ascospores. The awesome power of yeast genetics is partially due to the ability to quickly map a phenotype-producing gene to a region of the S. cerevisiae genome. For the past two decades S. cerevisiae has been the model system for much of molecular genetic research because the basic cellular mechanics of replication, recombination, cell division and metabolism are generally conserved between yeast and larger eukaryotes, including mammals.
The yeast's function in baking is to ferment sugars present in the flour or added to the dough. This fermentation gives off carbon dioxide and ethanol. The carbon dioxide is trapped within tiny bubbles and results in the dough expanding, or rising. Yeast are unicellular fungi that are versatile laboratory micro-organisms. They grow rapidly and have simple nutritional requirements. When yeast degrades nutrients in the absence of oxygen they use the process of glycolysis to produce energy in the form of ATP.
Below you can see a diagram of a yeast cell,

Prediction

I predict that as the concentration of sucrose is increased the yeasts respiratory rate will also increase producing more carbon dioxide as a result. But this will only be true up to a certain point after that the level of carbon dioxide produced will begin to level off and gradually decrease. Below is a sketch graph interpreting my prediction and what my results may look like.

Explanation of prediction

My reason for the above prediction is that yeast is a unicellular fungus, which feeds saprophytically and can respire both aerobically and anaerobically. In the experiment the yeast will be respiring anaerobically and breaking down the glucose stored in it as a waste product of this process it will also form CO2 and alcohol in the form of ethanol. This respiration process is called fermentation. The yeast breaks down the glucose using a series of enzymes. I deduce from this that the more glucose that is present in the yeast the more will be broken down and therefore more CO2 and ethanol will be produced as waste products at a faster rate. At lower concentrations, the active sites on most of the enzyme molecules are not filled because there is not much substrate. Higher concentrations cause more collisions between the molecules. With more molecules and collisions, enzymes are more likely to encounter molecules of reactant. The maximum velocity of a reaction is reached when the active sites are almost continuously filled. Increased substrate concentration after this point will not increase the rate. Reaction rate therefore increases as substrate concentration is increased but this is only true up to a certain point and after this the level of products formed e.g. carbon dioxide will begin to level off and gradually decrease which indicates that the rate of reaction has decreased.

The product formed as a result of the reaction, which is carbon dioxide accumulates as a waste product of fermentation in yeast and cellular respiration. Fermentation releases two molecules of the gas from the anaerobic degradation of glucose, as well as two molecules of ethanol plus usable energy for cell function. Cellular respiration, an aerobic process, liberates six molecules of carbon dioxide as well as several water molecules and energy. More energy is released by cellular respiration than by fermentation because glucose is completely oxidized in the process. Thus, carbon dioxide is a waste product of the energy-releasing mechanisms of the cell. Carbon dioxide is an indicator of the rate of substrate degradation. More carbon dioxide will be released as the rate of cellular respiration increases.

Enzymes catalyse reactions by lowering the activation energy for the reaction. Enzymes are made from protein and are specific as to which substrate they work on. In order for this to happen the enzyme forms a complex with the substance or substrates of the reaction. An enzyme molecule is globular and very large but only a small part of it, the active site, is involved in the reaction. In order for this to happen the shape of the active site has to match that of the substrate molecule. The reaction then takes place as the substrate molecule fits in and is then held there until the reaction is complete. The product can then be released and the enzyme is then ready to take part in another reaction. This is known as the lock and key hypothesis. The active site has a distinct Shape e.g. like a lock. Just as only the right 'key' will fit a lock, so only the right substrate has the right shape to fit into the active site.

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The enzyme and substrate slot together to form a complex, as a key fits into a lock. In this complex the substrate is enabled to react at lower activation energy. Catalysts associate with reactant substrates and cause a redistribution of electron densities in the reacting substrates. The bonds that need to be broken in the reaction are weakened by the association with the catalyst. This makes the reaction occur faster because the weakened bonds are easier to break. The enzyme is usually very large compared to the reacting substrate. Enzymes have folds and creases and the reactant molecules fit into a definite location in these folds and creases. This may be due to bonds within it being deformed and stressed in the complex, so making them more likely to react. Once the reaction has been catalysed, the products are no longer the right shape to stay in the active site and the complex breaks up, releasing the products and freeing the enzyme for further catalytic action.

A zymase-complex enzyme will only bind with a glucose molecule to produce the ferments carbon dioxide and ethanol, which brings about the fermentation in my experiment. This ties in with the Induced Fit theory which states that the substrate cannot bring about catalysis and the reaction itself, but the active site, when it comes in to contact with the substrate slightly changes its shape to form an effective fit and arrangement of catalytic groups on its surface which brings about the catalysis reaction.
Yeast after activation creates the ferments carbon dioxide and ethanol by secreting the enzyme zymase (a complex of 12 enzymes) in the Yeast, which acts on the sucrose. Yeast have to make energy, stored as ATP to carry out all cellular functions. To do this they can respire both aerobically when there is plenty of oxygen, but where oxygen is short, they respire anaerobically doing this they are called partial anaerobes. This produces less energy, but keeps the Yeast alive. Pyruvic acid has to be broken down in respiration when formed by breaking down of glucose molecules, this can’t be done in the same way as it is aerobically when respiring anaerobically which is how the carbon dioxide and ethanol is formed through the zymase. Here is the equation for anaerobic respiration:

Enzymes in cytoplasm

(Zymase complex)

Glucose -----------------> ethanol + carbon dioxide +energy

C6H12O6 2C2H5OH CO2 210 Kj/mole

There is 2ATP from each respired glucose molecule - in aerobic there is 38ATP.

Explanation of Glycolysis

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This is a preview of the whole essay

Enzymes in cytoplasm

(Zymase complex)

Glucose -----------------> ethanol + carbon dioxide +energy

C6H12O6 2C2H5OH CO2 210 Kj/mole

There is 2ATP from each respired glucose molecule - in aerobic there is 38ATP.

Explanation of Glycolysis

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Glycolysis is a nine step process, with carbohydrate intermediates at each step, that breaks a glucose molecule into two molecules of pyruvic acid, and produces two molecules of ATP for use by the cell via substrate level phosphorylation and two molecules of NADH, a high energy electron carrier. Normally, NADH would drop its two electrons and H+ ion at the electron transport chain, but this process of chemiosmosis does not occur during fermentation. Chemiosmosis requires O2 as the final electron carrier in the electron transport chain. The O2 is broken in half and is reduced with 2 electrons and 2 H+ ions to form H2O, water. Without O2, yeast will not proceed to chemiosmosis or even the Kreb Cycle, a cycle that produces even more high-energy electron carriers. Instead, it converts the pyruvic acid and 2 NADH that have been produced to 2 molecules of ethanol, a two-carbon compound. In this way, NADH is oxidized to NAD+ and is able to be reused. Two molecules of CO2, one for each ethanol that is made, are released as by-products.

I also think that after the level of carbon dioxide begins to level off it will also begin gradually decrease. I think this due to the outer solution being more concentrated then the internal concentration of the yeast cell. Osmosis is known as a special case of diffusion. Osmosis can be defined as the passage of water molecules from a region where they are in higher concentration to a region where they are in a lower concentration. Outside a cell, there is a low concentration of solute molecules and a high concentration of water molecules known as a hypotonic solution. Inside the cell, there is a high concentration of solute molecules and a low concentration of water molecules known as a hypertonic solution.

So as the outer solution is hypertonic compared to the inner yeast cells which are hypotonic water will diffuse from inside the cells in to the outer surrounding solution. So less carbon dioxide is going to be produced because if the yeast cell loses water less enzymes are going to be activated which would result in less active sites being available. So the decrease in carbon dioxide is going to be due to less enzymes would be occupied by the substrate molecule.
As enzymes only work in aqueous solution if the concentration is increased even more water will move out of the yeast cell as the substrate concentration is being increased because less enzymes are being activated, as there are less active sites available. So as the decrease in less active sites being occupied by the substrate molecules increases the amount of carbon dioxide being produced will decrease as a result of this.

Factors affecting the rate of reaction
As stated in my aim the factor which I am going to investigate is the substrate concentration, but there are also other factors which could affect the rate of respiration of yeast and these are stated below.

Temperature – enzymes work best at an optimum temperature. Below this, an increase in temperature provides more kinetic energy to the molecules involved. As the temperature increases the kinetic energy of the substrate and enzyme molecules increases and so they move faster. The faster these molecules move the more often they collide with one another and the greater the rate of reaction. As the temperature increases, the more atoms, which make up the enzyme molecules vibrate. This breaks the hydrogen bonds and other forces that hold the molecules in their precise shape. After about 40’C they begin to unravel and denature as the tertiary and quaternary structures change. So after this point the rate of reaction will decrease as the enzyme begins to denature.

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pH – As with temperature, enzymes have an optimum pH. If the pH changes much from the optimum, the chemical nature of the amino acids can change. This may result in a change in the bonds and so the tertiary structure may break down. The active site will be disrupted and the enzyme will be denatured. This is then going to reduce the rate of reaction resulting in less carbon dioxide production.

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Enzyme concentration – At low enzyme concentration there is great competition for the active sites and the rate of reaction is low. As the enzyme concentration increases, there are more active sites and the reaction can proceed at a faster rate. Eventually, increasing the enzyme concentration beyond a certain point has no effect because the substrate concentration becomes the limiting factor. So the higher the concentration the higher the rate of reaction due to there being more active sites for the substrate molecules to bind with.

Substrate concentration – at a low substrate concentration there are many active sites that are not occupied. This means that the reaction rate is low. When more substrate molecules are added, more enzyme-substrate complexes can be formed. As there are more active sites, and the rate of reaction increases. Eventually, increasing the substrate concentration yet further will have no effect. The active sites will be saturated so no more enzyme-substrate complexes can be formed. This is due to the enzyme concentration being the limiting factor as there are not enough active sites for the substrate molecules to bind with.

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Method

Below is a detailed list of the procedures that I am going to carry out in order to obtain reliable and accurate results.
To begin the experiment, firstly wash and clean all the equipment.
As the yeast has to be activated for 15minutes, this should be done first.
So a large beaker should be filled with hot water, its temperature continuously monitored and kept at 40’C,
Measure out 100cm3 of water in the measuring cylinder and pour it in to the conical flask,(Make sure that the bottom of the water’s meniscus sat horizontal with the required gradient, when looking from a 90’ angle from the side)
Weigh out accurately 3grams of yeast. (This is going to be constant for each mass of sucrose used)
Then weigh out the mass of sucrose that is going to be used. (Making sure that the balance is set to zero with the weighing boat on there,)
The masses of sucrose that I am going to use are as follows, 0,2,4,6,8,10.
Once the temperature of inside the conical flask is 40’c the buffer solution should be added. (This will keep the pH constant)
Then the sucrose can be added, however much is being investigated, making sure that the weighing boat is angled correctly so that all the sucrose is added to the flask to prevent any falling out of the flask.
It should be correctly stirred and if necessary large pieces should be broken up using the glass rod.
Monitoring the temperature keeping it at 40’C adding hot water if the temperature falls add the yeast to the solution.
Making sure that the solution is correctly stirred and no water is spilt. Once the yeast has been added, the stopwatch should be started and timed for 15 minutes.
Whilst the yeast is in the process of activation the rest of the apparatus should be set, making sure the clamp stand and boiling tube are correctly placed.
Fill the boiling tube upside down and place it underneath the water level in the water bowl. (Make sure the water level in the boiling tube is below the curve, a mark should be made on the test tube)
Once the yeast has been activated the temperature should be extrapolated to 37’C.
Now the experiment can begin placing the bung firmly in the flask.
The experiment should be timed for 3 minutes and the measurement of the amount of water displaced should be noted.

Below are the table headings of how I will record my results, and how they will be interpreted.

Once the results have been recorded, the equipment should all be washed and set up again.
Each mass of sucrose should be repeated twice, in order to increase the accuracy. The same method should be followed to investigate each mass of sucrose.

Looking at the method that I have chosen, I think it will enable me to produce accurate results, as the equipment and range of tests which I have chosen to do are appropriate. The equipment which is going to be used will also prevent errors, as the equipment will be laid out safely insuring that all the apparatus are connected to each other securely.

Apparatus
Below is a list of all the apparatus that I will use in order to carry out my experiment.

Conical flask (250cm3)
Beaker (500 cm3 )
Beaker (1dm3 )
Boiling tube
2 thermometers
Measuring cylinder (100cm3)
Syringe(5 cm3)
Glass rod
Spatula
Clamp Stand
Collecting bath(plastic container)
Capillary tube (connecting)
Stop watch

Reason for chosen apparatus
Within my chosen apparatus I have chosen a 250cm3 conical flask, the reason why I have chosen this over a test tube is that it can hold a greater volume than a test tube, so therefore it will allow the yeast to mix with sucrose solution more easily. It will also mean that the temperature of the solution within the flask will stay constant, as the solution will firmly be shaken allowing the heat to conduct through.
A large beaker will also be used in order for it to act as a water bath. This beaker will enable the flask to sit firmly and allow all the solution to be within the watermark, which will mean that all the solution will be kept at a constant temperature.
I also propose to use a measuring cylinder for measurements at will allow me to correctly see that the level is correct as the meniscus will not be over the mark. The stopwatch will be used for me to correctly time each experiment and exclude the use of estimation or guessing of how long each experiment has been going on for.
In order to keep the conditions of experiment constant a buffer solution is going to have to be used, for this a syringe will be used to measure it as this is the most accurate way and will prevent any errors occurring in the measurements.
In order to measure the displacement of each reaction a measuring cylinder is going to be used as well as a collecting bath. The reason why I have chose to use a boiling tube is that it is a more accurate way as from the measurements that I take I am going to carry out many statistical calculations.

Diagram Of Apparatus
Below you can see a diagram of how my experiment is going to be set up.

Preliminary work

In order to carry out my experiment correctly and investigate my chosen factor I carried out some preliminary work. This enabled me to chose appropriate scales for the amount of yeast and sucrose to use. Also in my preliminary work my task was also to work out the amount of time the yeast should be left to be activated. In the experiments that I carried out, I used the same method designed for the original experiment, as this will ensure that the experiment is carried out correctly and safely.
I carried out the experiment to notice if there was a significant change in using different masses of yeast.
The result that I obtained are displayed below,

As you can see above all the raw data from both experiments is displayed in the above table and the average has also been worked out as well as the volume of carbon dioxide produced in cm3. This was done using the following formula,

Volume= πr2h

The radius of the boiling tube is represented by ‘r’. The radius is the internal diameter divided by two. 2.16/2= 1.08cm.

In the formula the ‘h’ represents the height of the water displaced in cm’s.

π= 3.1421592654

In my preliminary experiments the conditions which I followed were, using 5grams of sucrose for each experiment, (kept constant throughout) temperature at 40’C for the activation period which was 20 minutes and then extrapolate the temperature to 37’C for the experiment, 150 cm3 of water was used and the experiment was done for 5 minutes.
From my results straight away by looking at my table I can see the masses of yeast that I should consider for my real experiment, as you can see that if the yeast mass is above 4 the reaction is to fast and cannot be measured using my equipment. When masses of 6 and 8 grams of yeast where used the bung on the flask had to be removed due to much froth being produced which forced the bung to pop off due the pressure build up. Also all the water in the boiling tube had been displaced so the reaction could not be measured. I think this may have been due to the experiment been timed for to long and all the water has been displaced.

Also taking my preliminary work a step further I noticed that the amount of water used could have caused the problem of the build of to much pressure as the volume of the solution was quiet high. But another cause of the problem could have been the activation time, as I noticed that after 20 minutes large amounts of froth had been produced by the yeast.

From my preliminary work I can conclude that the mass of yeast that I should use in my experiment should be 3grams. As this will produce reliable results and the results will be measurable. But the amount of water I use will be reduced to 100 cm3 as this will not make the solution overflow as the volume is not as large. As well as making these changes I am also going to reduce the activation time of the yeast to 10-15 minutes. I think this is more appropriate as I noticed that the formation of froth began after 5 minutes in my preliminary experiments. Another major change which I am going to make is that the amount of time the experiment is timed for, I am going to measure the amount of carbon dioxide produced for 3 minutes as I think this is more appropriate and when higher masses of sucrose are used the bung is not forced of due the pressure build up.

Fair Testing

In order to carry out my investigation correctly and obtain accurate result there are a number of factors which will have to be kept constant other than the variable that I am investigating. All the quantities will have to be carefully measured since small variations in the amount of yeast used can make significant variations in the results. I have chosen to repeat the experiment 3 times because it therefore allows me to calculate an average time. This will ensure that there are no abnormal results and it will increase accuracy.

The amount of yeast is crucial, more yeast means more sucrose will be respired and more products created. An imbalance will upset the results. The amount of yeast will be weighed out on an accurate top-pan balance each time.
The temperature is also a important variable which will have to be kept constant as if the temperature is to high the enzymes will be denatured and if is to low the reaction will not take place as readily as it should. So the temperature will be kept at 40’C for the yeast to be activated and then when I begin the experiment I will extrapolate the temperature down to 37’C. Here it will be kept at this temperature through out the experiment.
The pH of the solution will also be constant at a pH of 7. This will be achieved by using a buffer solution.
The stopwatch will be used in order to ensure accurate timing and that one experiment is not left longer than stated which is 3mins.
To ensure the accuracy and keep all the experiments fair the weighing of all the sucrose and yeast should be weighed out accurately to prevent any errors in the experiment.
Also the amount of water that is going to make up the solution should be constant through out all the experiments as this could affect the rate at which sucrose dissolves in it.

Safety precautions

Safety is an important issue to follow in my investigation as if the experiment is not carried out safely and in a correct manor it could affect the results that are obtained, which could be unreliable. One precaution that could be taken is to avoid the parallax error and always take measurements at eye level.
In my investigation all of the substances that are going to be used are of a low risk but I am still going to take precautions in using all the equipment. The yeast want active may be irritant to skin if any spillages occur so it should be immediately cleaned up and all skin most be thoroughly washed. Also as there is a lot of equipment which is made out of glass extreme precautions should be taken when handling the apparatus to prevent any breakages.

Bibliography

The following textbooks were used, for reading background information on the major topics relevant to my investigation.

Understanding Biology -Glenn and Susan Toole.
Advanced Biology -Ann Fullick.
Biology Principles and processes - Michael Roberts and Michael Reiss,
Biology 1, (Chapter 3) - Mary Jones.
Biology 2, (Chapter 5) - Mary Jones.

The following books stated above were a good source of information as it described the major topics in detail but also reinforced the information with diagrams that explained the information more clearly. I think this was the major advantage of using this source of information. The disadvantage of using this source of information was that the information had to be manually searched for which was time consuming.

I also used the following website to research my investigation

I found using the Internet to research my investigation very interesting, as the advantages of using the Internet were that I was able to search a wide range of topics related to my investigation and this allowed me to get a wide overview of the whole problem. Disadvantages of this source of information was that a lot of irrelevant data was also displayed with the main topics which meant that a large amount of information had to be read through in order to understand the topic that I searched for.

Another source of information that I used was an information CD, Encarta ’99.

Advantages of this were that it allowed me to search exactly information on the topics that were relevant to me and included movie clips in order to explain diagrams.
But the disadvantages of this source of information were that it only displayed brief definitions on the topics and does not explain in detail on the main points of the topic.

Evaluating all the secondary sources I used I found the best source being the textbooks. This was due to a number of reasons but manly the books included a great depth in to the topic that I searched for and increased my understanding as they also included very useful diagrams which interpreted the text for increased understanding.

Results

In order for the results to be obtained in their final form, a calculation is going to be carried out in order to work out the volume in cm3. The following formula can be used.

Volume= πr2h

The radius of the boiling tube is represented by ‘r’. The radius is the internal diameter divided by two. 2.16/2= 1.08cm.

In the formula the ‘h’ represents the height of the water displaced in cm’s.

π= 3.1421592654

So in order to work out the volume for 2 grams the following calculation will be carried out, π * 1.082 * 2.75 = 10.08

In order to take the investigation a step further from these results I am going to work out the concentration of sucrose from the mass of it used. This is most suitable for my investigation as I am investigating how the concentration affects the rate of reaction. Further statistical calculations that I carried out are shown below.

The concentration can be worked out using the following formula,

Concentration = moles ÷ Volume (dm3)

In order to work out the concentration of sucrose, firstly the moles have to be worked out using the following formula.

Moles = Mass ÷ Mr

The Mr of sucrose is equal to 342.

Substituting in the values the concentration and moles can be worked out as below, this is an example of 10g.

Moles = 10 ÷ 342 = 0.029 moles

Concentration = 0.029 ÷ 0.105 = 0.28

Volume = Total volume of solution = 105 ÷1000 = 0.105dm3

Analysis

From the analysis of my data in my results table the volume of gas in 1 minute for 1 gram was worked out and plotted on the graph against the mass of sucrose used. I can conclude that as a general trend, as the mass of sucrose is increased the amount of gas given off is also increased up to a certain point from where the amount of gas collected gradually begins to decrease and level off. So what I stated in my prediction was correct and the diagram I used to interpret my prediction with was also correct.

Looking at my results I can see that when no sucrose was added there was still some carbon dioxide collected even though it was a small amount it shows that the yeast cells have been respiring glycogen in order for it to go in to glycolysis. I think that a very small amount of carbon dioxide was produced because the yeast had a store of glycogen as they were bought in a dormant state. At a mass of 0 grams the volume of gas produced was 0.02 cm3. So the rate was very slow as there was no respiratory substrate added so the rate at which glycolysis occurred was very low which meant that the metabolic activity of the yeast cells was very low.

Form a mass of 0 grams to 6 grams the volume of carbon dioxide collected increased, This is due to not all the active sites being occupied by substrate molecules and as the mass of sucrose was increased the active sites began to be occupied by the substrate molecules which as a result means that more carbon dioxide is going to be produced.

At a mass of 7 grams and above the level of carbon dioxide begins to level off it also gradually begins to decrease. At 7 grams the volume of carbon dioxide produced was 2.36 cm3. I think this is due to the outer solution being more concentrated then the internal concentration of the yeast cell. So osmosis will result in the movement of water from a high water potential to a region of a low water potential. Outside a cell, there is a low concentration of solute molecules and a high concentration of water molecules known as a hypotonic solution. Inside the cell, there is a high concentration of solute molecules and a low concentration of water molecules known as a hypertonic solution. So as the outer solution is hypertonic compared to the inner yeast cells which are hypotonic water will diffuse from inside the cells in to the outer surrounding solution. So less carbon dioxide has been produced because if the yeast cell loses water less enzymes are going to be activated which would result in less active sites being available. So the decrease in carbon dioxide is due to less enzymes being occupied by the substrate molecule. During the intermediate values of the mass of sucrose used the graph begins to show a gradual levelling off, this is due to the enzymes working at a constant rate as all the active sites of the enzymes are being occupied by the substrate molecules, so a increase in substrate molecules has no effect. This shows that the limiting factor here is the amount of enzymes.As enzymes only work in aqueous solution if the concentration is increased even more water will move out of the yeast cell as the substrate concentration is being increased because less enzymes are being activated, as there are less active sites available. So as the decrease in less active sites being occupied by the substrate molecules increases the amount of carbon dioxide being produced will decrease as a result of this. So this will also mean that the rate at which glycolysis occurs will also decrease.

Another reason to why I think the level of carbon dioxide began to decrease is due anaerobic respiration. As I stated in my prediction the products of this are carbon dioxide and ethanol, which is a toxic substance and may inhibit the enzymes which would mean they would not function correctly but also inhibit anaerobic respiration. So if the concentration of the substrate molecules were increased it would mean that the rate of anaerobic respiration would have increased as well as glycolysis increasing its rate too. This would result in the products to increase so more ethanol would be produced. So looking at my results after a mass of 6 grams the volume of carbon dioxide collected began to decrease and this may have been due to the ethanol inhibiting the enzymes and there for reducing the rate of carbon dioxide given off.

Evaluation

Looking at the results I have obtained in my investigation they support my hypothesis that as the concentration of the substrate is increased the rate of respiration will also increase. This did happen as the concentration of sucrose was increased the amount of carbon dioxide given off also increased up to a point where the amount of gas gradually began to decrease and level off. So looking at my results they all fit in to the general trend and there don’t seem to be any major anomalies in the results.

Looking at the result obtained for 8 grams of sucrose, the volume of gas collected seems just slightly of the general curve. This can be seen on the graph. This slight difference in the volume of gas collected may be due to an error when carrying out the experiment. What could have happened was that when the sucrose was being weighed out and placed in the weighing boat I noticed there were lumps of sucrose together as well as the single grains. So this means that the surface area of the sucrose had been reduced, which could mean that it took longer to dissolve properly in the water and this could of have affected the activation of the yeast as there may have been less substrate molecules. So this could have caused the result to be slightly inaccurate, as the sucrose may not have completely dissolved resulting in less substrate molecules for the enzymes to bind with, therefore producing less carbon dioxide.

Looking at the experiment as a whole I think there might have been one source of error which could have affected the results as a whole. This could have been due to heat loss in the experiment. As a wind draft could of caused slight variations in the temperature which may have altered the rate at which enzymes work causing fluctuations.
I think an improvement that could be to the investigation as a whole is to use an electronic water bath. So this will be an advantage as the temperature can be set and if it goes to cold the water will be heated back to that temperature. I think this is more accurate as it would avoid and fluctuations in the temperature and it will mean that the temperature will stay constant. But overall I have ensured that my results are accurate by controlling all the variables stated in my plan. I also took care when using the equipment in order to retain continuity throughout the experiment. For this I checked everything was set up correctly at each reading and prepared my solution each time in the same way.

To obtain more reliable results I would want complete continuity with preparations, arranging sets of substances to create multiple solutions beforehand or preparing them but not actually activating the yeast, so as to prevent any getting a head start over the others. This would ensure that all the preparations are the same and would give continuity. I would also want to be more strict and thorough with preparing solutions and mixing them up. I would want each one to be thoroughly acclimatised to the surroundings and had the same amount of yeast added, same activating time and mixing time. This would enable me to obtain more reliable results throughout. If I were to further investigate this experiment and my results I would probably calculate the point at which where the enzymes begin to denature for respiration in yeast. I could also examine validity and continuity, also may be included them in more intricate calculations which include log.
Looking at my investigation as a whole I don’t think there are any major limitations which may have affected my results but I think a limitation might have been in the results that I have taken. As I think that it would have been more appropriate to investigate the masses of sucrose up to 15 grams. As this would of shown definitely whether or not the graph would level off or if it would decrease more and then level off. So I think this was a limitation in my investigation as the trend stated in my prediction could have been reinforced if the graph fully showed the trend explained in my prediction.

More generally, my results are fairly consistent, as well as being consistent with the prediction, which is an indication that they are reliable, as the primary quantitative prediction as to the rate of respiration increasing as the concentration of sucrose increases appears to be conclusively and reliably proven.

Ibrar Ajaz