Research Question: To what extent does altering the saturation of hydrogen peroxide affect the rate of reaction with catalase found in yeast?Aim: To investigate how changing the saturation will cause the rate of reaction to change by using different amounts of hydrogen peroxide to see how much oxygen is produced within two minutes to determine the rate of reaction. Hypothesis: I hypothesise that as hydrogen peroxide saturation increases, the rate of reaction will also increase due to the increased probability of substrate present to react with the enzyme. Personal Engagement and Background Research An enzyme is a globular protein that is a found in living organisms that acts as a biological catalyst, which speeds up reactions that occur in the body without being used up and using ATP. This means that an enzyme can be used over and over indefinitely (“Enzyme & Substrate | Bioninja”). Enzymes react with the substrate by having an ‘active site,’ which is the site where a specific substrate attaches to the enzyme. After attaching to the substrate, an enzyme-substrate complex is formed, whereby the enzyme catalyses the reaction to form the enzyme-product complex. The product is then released from the enzyme, and the enzyme can continue to catalyse reactions (“Enzyme Catalysis | Bioninja”). Enzymes are substrate-specific; this means that only a specific enzyme can catalyse a specific substrate. Enzymes are incredibly important for biological functioning, as they are involved in many processes throughout the body to ensure survival. For example, enzymes are heavily involved in the digestive process, where enzymes such as amylase, protease and lipase break down starch, proteins and fats respectively, into smaller molecules in order for absorption to occur. Without these enzymes, we would not receive the nutrition we need for survival (“Chemical Digestion | Bioninja”). Yeast contains the enzyme catalase, which is produced in the liver of the body and breaks down hydrogen peroxide. Hydrogen peroxide is a waste product of metabolism and is toxic to the body. In order for the body to rid itself of hydrogen peroxide, catalase is produced to break down hydrogen peroxide into water oxygen (“The Health Benefits Of Catalase”). Temperature, pH level and substrate are factors that are essential for enzyme function; at the optimum level, the enzyme will function at its quickest. If the optimum is passed, the enzyme denatures rapidly, whereby the proteins found within the enzyme are altered and substrates are no longer able to bind with the enzyme, causing the rate of reaction to decline rapidly. Temperature allows for an increase in kinetic energy available for the enzyme to move and collide with substrate molecules. High temperatures cause the enzyme to destabilise and disrupts the hydrogen bonds within the enzymes, thus denaturing the enzyme. pH alters protein solubility and changes the overall shape of the enzyme, so the enzyme must be in a solution with the correct pH, otherwise it will not be able to catalyse the reaction. Substrate concentration increases the rate of reaction due to the increased number of molecules of substrate for the enzyme to collide with. Rate of reaction will increase until the enzyme reaches the saturation point, where it is working at its optimum level (“Enzyme Activity | Bioninja”). 2.0: ProcedureVariables Table 1: Table to Depicting Variables of the Experiment VariableType of Variable Reason Saturation of Hydrogen PeroxideIndependent Amount of peroxide changed to investigate its effects on the rate of reactionVolume of Oxygen ProducedDependentChanges based on the saturation of catalaseTemperatureControlEnsures that temperature is not causing change in the rate of reaction; ensures enzymes do not denature if temperature is too highpHControlAllows for the maintenance of of protein structure within the enzyme; ensures rate of reaction is not altered by pHConcentration of CatalaseControlMaintains enzyme concentration ApparatusHydrogen Peroxide (6%)Yeast Gas Syringe with TubePipetteConical FlaskTimerStopperMetal ClampDistilled Water1 measuring cylinder (50 mL)Safety GogglesSafety Precautions Table 2: Hydrogen Peroxide HazardSymptomsPreventionWhat To Do If ExposedSkinCorrosive, Redness, Skin BurnsWear gloves and coat whilst handling Rinse with waterEyesRedness, Pain, Blurred Vision Safety GogglesRinse with waterIngestionSore throat, vomiting, nauseaDo not eat in the labRinse with water, do not ingest further(“CDC – HYDROGEN PEROXIDE (>60% SOLUTION IN WATER) – International Chemical Safety Cards – NIOSH”)No further ethical considerations needed to be made with the experiment. MethodActivate the yeast by adding warm distilled water and sugar according to the instructions and quantities on the label.Set up a conical flask with a stopper connected to a gas syringe.Measure out 15 mL of yeast and pour in the the conical flask.Using a pipette, drop 5 drops of 6% of hydrogen peroxide into the conical flask.Place the stopper on the conical flask to stop the gas from escaping and start the timer for two minutes.When the time is done, record the amount of gas released on the gas syringe.Repeat steps 3 to 6 5 times to gain five sets of data for the number of drops.Repeat steps 1 to 7 for 5, 7, 10, 13 and 15 drops of hydrogen peroxide.5.0: ResultsQualitative Data When hydrogen peroxide was added to the yeast, effervescence occurred. As more hydrogen peroxide was added, bubbling increased rapidly and strongly. Additionally, foam was produced, although the foam was not permanent and eventually faded. As there was the hydrogen peroxide was increased in quantity, more foam was produced. The substrate also changed to a slight brown colour as the yeast was added.Quantitative Data Table 3: Table to Show the Volume of Oxygen Collected in the SyringeOxygen Collected in Syringe (mL ±0.25cm3)Hydrogen Peroxide (# of drops)Trial 1Trial 2Trial 3Trial 4Trial 55 13191615187 172520211910252628262413 838580838615909510010097Processed DataTable 4: Table to Show the Average Volume of Oxygen CollectedHydrogen Peroxide (# of drops)Average (mL ±0.25cm3)516.2720.41025.81383.41596.4Chart 1: Chart to Show the Average Volume of Oxygen CollectedTable 5: Table to Show Rate of Reaction of Catalase against Hydrogen PeroxideHydrogen Peroxide (# of drops)Rate of Reaction (cm3s-1)50.13570.170100.215130.695150.803Rate of Reaction=amount of oxygen produced (mL)unit of time (seconds) Sample Calculation for 5 drops of hydrogen peroxide Rate of Reaction=16.2120=0.135 cm3s-1Calculating the Uncertainty of Rate of Reaction Relative Uncertainty of VolumePercentage Uncertainty= absolute uncertaintylowest measured value 100=0.2513100=1.92% Relative Uncertainty of TimePercentage Uncertainty=0.5120100=0.42% Relative Uncertainty of Rate of ReactionPercentage Uncertainty=Uncertainty of Time (%) + Uncertainty of Volume (%)=1.92%+0.42% Total Uncertainty= 2.34%Chart 2: Chart to Show the Rate of Reaction Between Catalase and Hydrogen Peroxide6.0: ConclusionGraph 4 shows a positive correlation between volume of oxygen and the increase of hydrogen peroxide; as the amount of substrate increases, the more oxygen is produced from the enzymatic reaction of hydrogen peroxide. For example, when five drops of hydrogen peroxide was added, it can be seen that 16.2mL of oxygen collected in the gas syringe, whilst when 15 drops were added, 96.4mL of oxygen was indicated, strongly supporting the original hypothesis, which was that as more hydrogen peroxide was added to the yeast, the higher the rate of reaction. Graph 5 further supports the original hypothesis, as it demonstrates the change the rate of reaction according increased volume of hydrogen peroxide, whereby the fastest rate of reaction occurred at the highest number of drops added to the solution. The graph seems to plateau at 15 drops, potentially indicating the substrate to enzyme ratio was nearing saturation. The graph shows that there is a gradual increase from 0 drops to 10 drops of hydrogen peroxide, where the rate of reaction goes from 0cm3s-1 to 0.215cm3s-1. Following this, there is a rapid increase in the rate of reaction from 10 drops of hydrogen peroxide to 15 drops, where the rate changed from 0.215cm3s-1 to 0.803cm3s-1; an increase of nearly 4 times that occurred in a range of 5 drops, as opposed the 0.215 change occuring in a span of 10 drops, indicating that as hydrogen peroxide presence increased, rate of reaction increased rapidly. The results show support for the original hypothesis; therefore the hypothesis is accepted and it can be said that as substrate saturation increases, the rate of reaction also increases.As the presence of hydrogen peroxide increase, the rate of reaction is also able to increase due to the increased frequency of collisions between enzyme and substrate. As there is more substrate, the likelihood of a collision between enzyme and substrate increases, allowing for the breaking down of hydrogen peroxide due to the formation of enzyme-substrate complex. As more hydrogen peroxide is broken down by the enzyme catalase, more oxygen is produced as a product of the enzymatic reaction, and thus more oxygen gas is collected in the gas syringe, thus achieving the results demonstrated within the investigation.Catalase is an important enzyme needed for living organisms; it catalyses hydrogen peroxide, a toxic waste product, into water and oxygen, keeping the body healthy and functioning. Without catalase and enzymes in general, the body will not be able to function at an efficient rate. 7.0: Evaluation Strengths of the ExperimentHighly controlled variables, ensures that hydrogen peroxide saturation is the only limiting factor of the experimentLow percentage uncertainty increases reliability of data, meaning results can be applied to the conclusion accuratelyLow absolute uncertainty of equipment allows increased accuracy in measuring dependent variableLimitations of the Experiment WeaknessImpactImprovements Limited Testing Lowered reliability of averageIncrease repeated testing to increase reliability of data Inaccurate method of extracting catalaseUnsure of how much catalase is actually present in the yeast solution, so differing drops will have differing amounts of catalase, furthering Use material with measured concentration of catalase to ensure concentration of catalase is known to manipulate the variable is betterFurther Investigation For further investigation, this experiment could be done the other way around to see how enzyme concentration can affect the rate of reaction as opposed to substrate concentration. Additionally, factors such as temperature or differing pH levels could be investigated to see their effects on rate of reaction. Additionally, it may also be worth investigating how hydrogen peroxide reacts with different types of enzymes instead of just catalase, and see how the acid affects their rate of reaction. Furthermore, in regards to this investigation, more values of hydrogen peroxide have been investigated to see at what point the substrate-enzyme solution is fully saturated.