The amount of yeast will be controlled by making up a suspension of 5g of freeze dried yeast in 50cm3 of water, and frequently stirring the suspension. The yeast will be measured out with a 1cm3 syringe accurate to 0. 05cm3. The apparatus will be kept the same for each solution, and the boiling tube will be rinsed and dried between each reaction. The oxygen will be collected over water for 15 seconds each time, timed using a stopwatch. Six different concentrations of hydrogen peroxide will be used:.
More readings are being taken at the higher concentrations to enable the asymptote of the graph to be more easily observed. Three close results (within 10% of each other) will be obtained and averaged to ensure reliable results. Anomalous results (those not which do not fit the general trend and are more than 10% different from the other set) will be repeated until three close results are found, and anomalies will be excluded from the average. The most accurate available and practical equipment will be used in order to ensure precision in measurement. Dilutions Percentage concentration of hydrogen peroxide
Volume of 20% Hydrogen Peroxide (cm3) Volume of distilled water (cm3)Safety: 20% Hydrogen peroxide is corrosive and causes burns, lower concentrations are irritant, so safety goggles and protective gloves should be worn when handling it. If it is swallowed, wash out mouth, give a glass or two of water and seek medical attention, since it can cause serious internal damage due to the release of oxygen. If the liquid gets in the eyes, flood the eye with gently running tap water for ten minutes and seek medical attention.
If it is spilt on the skin or clothes, flood the area with plenty of water, remove contaminated clothing, and seek medical attention if a large area is affected or blistering occurs. In the case of spillage, alert the teacher. Apparatus List: Two 10cm3 syringes for hydrogen peroxide and water for dilutions – accurate to 0. 2cm3 1cm3 syringe for yeast – accurate to 0. 02cm3 Rubber bung with two holes and hollow needle. 200cm3 beaker for water bath, Boiling tube, Delivery tube, Trough to collect gas over water, Burette – accurate to 0. 05cm3 Stopwatch – accounting for human error, accurate to 1s.
This represents the most accurate equipment available to carry out this experiment. Collecting the gas over water, and using a hollow needle to insert the yeast means that no gas is lost since it is a closed system. Syringes enable quick and easy measurement of the small volumes with fairly good accuracy. Collecting the gas in an inverted burette is simple to carry out, and is very accurate. Method (diagram as for pilot) 1. The apparatus was set up as shown in the diagram 2. 10cm3 of the solution of hydrogen peroxide was measured out according to the dilution table above. 3.
The burette level was observed and recorded 4. The yeast was stirred, and 1cm3 was measured and placed onto the hollow needle. 5. The stopwatch was started, as the yeast was injected through the hollow needle. 6. When 15s had elapsed, the burette was removed from over the delivery tube, but the burette end was kept in the water to prevent the water from rushing out, and the burette level was observed and recorded. 7. The equipment was washed up and the experiment was repeated to obtain three close sets of results for volume of oxygen given off by the six concentrations of hydrogen peroxide.
From the average volume of gas produced, the rate of reaction was obtained using the formula explained in the background theory. Analysis A graph was plotted of the concentration of hydrogen peroxide against the rate of reaction and a best fit line was drawn. A clear trend could be seen, that as concentration of hydrogen peroxide increases, so does the rate of reaction, as was predicted. At 5% H2O2, the rate of reaction is 35cm3min-1, whereas at 20% the rate is 110. 34 cm3min-1. The slower increase in reaction rate predicted at the higher concentrations is seen.
The gradient of the curve at 5cm3 is 10, whereas at 15cm3 it is much less steep – 3. 78. However, the asymptote predicted is not seen on the graph, because the concentrations do not get high enough. So to obtain values for Vmax and Km the graph was extrapolated by eye to estimate the location of the asymptote – it is at approximately 117cm3min-1.
It produced reliable results from which a confident conclusion can be drawn which supports the original hypothesis. This has been ensured by several methods. All the variables which could affect the rate of reaction were controlled. Some variables were easy to control; others were harder. The variables controlled were: temperature, volume of hydrogen peroxide, amount of yeast, apparatus and time The table below shows how I controlled each variable and how successful this was.
For more detail on how they were controlled see the main plan. Variable How controlled? How easy/successful? Temperature at which the reaction took place The water bath was filled with tap water, adding hot or cold to adjust the temperature to 15? C. This was relatively simple to control, since water is a good thermal buffer. It should not have been a large source of error. Volume of Hydrogen Peroxide Solution This was measured out using two syringes to make up the right dilutions This was also simple to control and should not have caused very much error. Amount of Yeast
This was weighed out on a top pan balance, and stirred into the water to form a suspension. I think this was a major source of error, as the yeast would settle out after being mixed up. There also biological variation as to the amount of catalase in each yeast cell, and the yeast produces bubbles of carbon dioxide which form a foam which makes measuring precisely 1cm3 difficult. Apparatus The same apparatus was used each time, and it was washed between each experiment. This should not have been a major source of error Time 30 seconds was timed for each reading using a stopwatch.
This was a little awkward – as handling a stopwatch whilst manouvring a burette or syringe and hollow needle is not ideal, but it should not have caused a great degree of error. A fair test is one in which all variables are controlled except the one being investigated. In this experiment the variables were quite well controlled. The anomalous results were probably caused as a result of not stirring up the yeast properly, or because of biological variation in the yeast. This could be improved by stirring more often, and by making up a fresh batch of yeast mixture more frequently.
I repeated my experiment until I had two sets of close results. If any results were very different to the rest for that concentration, I excluded them from the average and repeated the experiment again. These results are ringed in red. This could have been caused by any of the errors accounted for in the table above; particularly the difficulty in controlling the amount of yeast. The standard deviations of my results are shown below. % Concentration Hydrogen Peroxide Rate of reaction (cm3min-1) Standard Deviation Set 1 Set 2 Set 3 Average These standard deviations are fairly small.. This shows that the averages I obtained were quite reliable. My graph shows a clear trend which enables me to draw an accurate conclusion, but it does not extend far enough to see the asymptote which is predicted by Michaelis-Menton kinetics.
There would be two ways of obtaining more accurate values for Vmax and hence Km, either by extending the range to make the appearance of the asymptote clearer, or by using a Lineweaver-Burke plot: which is a graph of 1/[S] against 1/rate. The Michaelis Menton equation can be rearranged to show that the y intercept of the Lineweaver-Burke plot is 1/Vmax, and the gradient of the straight line is Km/Vmax. This is much more accurate than trying to obtain values by determining the value of the asymptote by eye.
Better values for the initial rates of reaction could have been obtained if more time were available by tracking the progress of each reaction and drawing a graph of volume of oxygen produced against time, and then taking the gradient at t = 0 for each reaction – the true initial rate.
Bibliography Notes from teachers, Biology 1 – Cambridge University Press, Biological Science 1&2, Taylor, Green and Stout – Cambridge University Press http://www. iupac. org/publications/analytical_compendium/Cha19sec22. pdf, http://koning. ecsu. ctstateu. edu/Plant_Physiology/enzyme1. html.