If 10 Ml of of potato catalase is added to 30Ml of hydrogen peroxide and 4 different stresses are present we think the one which has a hotter environment will produce the most amount of oxygen. With what we have learned the heat will make particles move faster so reactions will occur quickly. This will allow the largest window for oxygen to produce. We think around 110-140 Ml of oxygen will be produced. In second place i think the one which is unchanged will have the 2nd most amount of oxygen produced. We also think the other stresses have large negative downsides which will disrupt the function of the enzymes. The osmotic stress will produce the 3rd highest amount of oxygen as the effect will only delay the time it takes the oxygen to produce but after a few moments we believe the salt will only dissolve and the hydrogen peroxide catalase reaction will continue. At 4th place will be the cold stress. The cold temperature will slow down all reactions and the average will be around 60-75 Ml. At last place will be the metal ion stress which we believe wont produce any oxygen as the metal ions are very good Inhibitors. We think at most 5 ml of oxygen will be produced.

Enzymes are proteins found in all living organism. They speed up chemical reactions by lowering activation energy but are never used up in a reaction. Each enzyme has a unique shape, the tertiary structure which covers the active site(place where reactions occur). The structure can shape shift demonstrated by the induced fit model which explains that the shape may change depending on its substrate. This allows for binding to be very effective. enzyme catalase is found in human cells and breaks down waste products (hydrogen peroxide) into water and oxygen. However enzyme catalase is also found in potatoes and is essential for digestion and cellular repair.

Function of an enzyme is based of its 3 dimensional shape which is linked by amino acids. Under extreme conditions structures may be deformed also known as denaturation, which prevents binding.

Enzymes face many stresses, one of those being temperature stress. Anything above 105 degrees F can make molecular collisions and kinetic energy so high that it will cause shape of the structure to denaturation. Lower temperatures do the opposite. They make the enzyme slow down and chemical reactions also become slower. However, they do not break the shape of the enzyme like high temperatures

Metal ions are another stress. In this experiment we use copper sulfate. The copper or Cu2+ is a positively charged ion. These ions bind on to the the enzyme and block the active site. This blocks the way of the hydrogen peroxide to enter and the copper ion acts as an inhibitor. The sulfate is a spectator and adds no other benefits or disasters.

Osmotic stress occurs when there is a difference between solute concentration between a cell and everything else. When salt or sodium chloride is added to a reactions, liquids around the salt move away and get displaced. This creates a hypertonic environment. The liquid tries to fix the difference in concentration but this can lead to unstable environments. It may disrupt reactions and can partially destroy the enzymes structure. Hydrogen and ionic bonds may also be disrupted which can denature the enzyme.

Manipulated 4 different types of stresses Hot-105+ Degrees F Cold- Below 65 Degrees F Osmotic- 1/8 tsp salt chemical- 1/8 tsp copper sulfate

Responding Oxygen produced in ML over a 1 minute and 30 seconds time frame.

Controlled

• Enzyme Type- Catalase from a potato
• Enzyme concentration- 10 ML 
• Hydrogen peroxide concentration-  30 ML
• Reaction Time- 1 Minute and 30 seconds
• Mixing Method- Mixing will not be done and reactions should occur on their own
• Equipment- All equipment and the set up of the apparatus will remain the same and all equipment and materials will be cleaned with soap and water and dried using a paper towel.
• How diluted the enzyme concentration was(Around 20 Ml per peeled potato which created around 80-100 ML. The total volume was 100-120 ML and if we divide these numbers be the 20 ML of water you get a concentration of anywhere between 80%-83.3%

Uncontrolled

• Volume of reaction to the chemical and osmotic stress will be a little higher(2 ML higher) because we are adding an additional compound. 
• The temperature for the hot and cold stresses won't be consistent throughout the entire test and may increase or decrease over the period of time due to thermal equilibration. 
• As we are getting the enzyme from several potatoes, purity and age of the enzyme may be different in each separate test.
• Delays between starting the experiment and the timer.

Catalase preparation 1- Cut and peel 2 potatoes. 2-Blend with 40Ml of water. 3-Belnd until a bubbly liquid forms. 4-Strain out chunks. 4- Measure 10Ml out in a measuring cup.

Basic Reaction 1- Set up apparatus (Connect tubing to the syringe and the stopper which is plugged in the flask). 2- Measure 30 ML of hydrogen peroxide and put into the flask with your 10 ML of catalase. 3-Seal flask with the stopper and start the timer. 4- After 1 minute and 30 seconds detach tube from syringe. 5-Oxygen should have pushed back the syringe allowing you to record your results. 6- repeat until 3 trials are complete.

Hot and Cold 1- When apparatus is set up fill a container with your water at the set temperature and place the flask in the water. 2- Repeat steps 2-6 from the basic reaction.

Osmotic and Chemical 1-On step from the basic reaction when you mix all the chemicals add your additional compound (salt or copper sulfate). 2- Repeat steps until three trails are complete.

Trail 1 Unchanged- We can see that the solution started rising almost immediately. AT the start bubbles formed fairly quick and reactions occurred quickly, proven by how fast the oxygen was moving the syringe. At around the 30 second mark it was at 25 Ml which was very quick. However there was a significant drop after this.  For the next minute nothing happened in the solution, and oxygen only raised up till 12 Ml  ending at 37 ML.

Cold- The solution did not react for several seconds, we measured the temperature to be at 59.4 degrees F. The solution rose about a cm but nothing else occurred. It produced no oxygen at 0 ML.

Hot-The reaction started at around the first 10-15 second, but the the syringe only started moving after around 50 seconds. The temperature was 108 degrees F It moved quick for the next 35 seconds but the last 15 seconds it showed very slow movement. It ended at 69 Ml.

Osmotic-bubbles were formed and the solution raised in the very first 10 seconds. The syringe also moved very quickly, 20 seconds into the experiment.  For the next 10 seconds the syringe was pushed back insanely quick. When there was around 50 seconds left the oxygen production lowered at a more consistent speed and remained like this until the end. It ended at 87 ML.

Chemical-As soon as the copper sulfate was poured into the flask the color changed to a light blue color. Not a single bubble was formed and the solution did not rise at all. No oxygen was produced.

Result- The osmotic stress had produced the most amount of oxygen, at 2nd was the hot temperature, and at third was then unchanged one. The most surprising result to us was the unchanged reaction which produced very little oxygen compared to what we learned. It should have produced at least 90 ML but fell very short to what amount we learned it would produce.  Both copper sulfate and the cold temperatures got 0 ML which did not surprise us to much but we did believe they would produce at least 10 ML.

Trail 2 Unchanged-The reaction started to rise at around the 30 second mark. 40 seconds into the experiment was when the syringe started to move. For the next 20 seconds the solution had produced a significant amount of bubbles but the oxygen production died out quick after a minute passed by. The last 10 seconds had little to no movement in the syringe where it ended at 50 ML.

Cold- The temperature was measured at 63.6 degrees F. 15 seconds into the experiment was when the movement of the syringe started. The movement was super quick and after about 30 seconds it had already reached 40 ML and was maintaining oxygen production reaching 85 ml 15 seconds later.  When there was 40 seconds remaining the oxygen production dropped hard. However it continued to move and went over 150 Ml with 10 seconds still remaining, but oxygen production almost immediately stopped and it finished at 160 Ml.

Hot-the temperature was measured at 107.7 degrees F. The reaction started at around 25 seconds into the experiment but no oxygen was produced. At around 15 seconds remaining oxygen was finally being produced. It reached 15 Ml but showed pretty much no movement with the last 8 seconds. It ended at 19 Ml.

Osmotic- Reaction started to rise as soon as everything was poured in. The syringe moved 20 seconds into the experiment but wasn't moving to quickly.  A minute remained an it was at around 20 Ml. It had a sudden increase in speed but the oxygen produced slowed down again. It had a consistent speed after this and reached 65 Ml with 30 seconds left. Oxygen produced remained stable but with around 15 seconds left it died out a little. It ended at 91 ML

Chemical-Just as trail one as soon everything was poured in the color turned blue. There was no change at all in how much the solution rose and no oxygen was produced either.

Results- In this trial the cold stress was the leader in oxygen produced, in 2nd was osmotic and at third was the unchanged reaction. The most surprising result was the cold temperature which in trial 1, had no oxygen produced but this time it obliterated every other test in terms of oxygen produced. Unchanged reaction and the osmotic reaction were still pretty consistent but the hot temperature had a significant decrease compared to the first trial.  The chemical reaction had pretty much an identical test in both trials.

Trail 3 Unchanged- Reaction started almost instantaneously as the chemicals were dropped. Syringe started moving 10 seconds in at an incredible speed. 30 seconds into the experiment it had already reached 65 Ml and was still moving very quickly. Around 40 seconds in is when the speed of the oxygen produced decreased but was still moving at a very impressive rate. With 30 seconds still remaining it had reached 140 Ml and showed no signs of stopping.  With 10 seconds still remaining it had capped out the syringe and 4 seconds later you can see oxygen building up tightly in the tube and flask. Due to how much oxygen was pushing against the syringe it had almost popped off if we didn't stop it. From what we estimated it would reached 180 Ml.

Cold- The temperature was measured at 64.1 degrees F. The reaction quickly grew in size and around 20 seconds in was when the oxygen started producing. It moved surprisingly fast and 30 seconds in it was already at 35 Ml. The speed at which oxygen was being produced remained stable, only slightly decreasing in speed. With around 10 seconds remaining when it reached 130 Ml was when you could see the big difference in speed ending at around 148 Ml.

Hot-The temperature was measured at 106.4 degrees F. 15 seconds into the experiment was when oxygen started moving the syringe. In 30 seconds it had already reached 30 Ml and remind at a steady speed. 40 seconds into the experiment was when the speed dropped by a huge margin. The syringe had slowed down and with 30 seconds remaining it had reached 75 Ml. However, for the next 30 seconds the syringe barely moved and ended at 99 Ml.

Osmotic- Syringe started moving after about 15 seconds but solution did not rise much. At around the first 30 seconds was when it caught some speed and solution began to rise rapidly. within the first 30 seconds it had only gotten to 25 Ml but sped up significantly.  It maintained this speed for 10 seconds but started to slightly decrease in the rate at which oxygen was being produced. It reached around 80 Ml with 30 seconds remaining. It ended at 116 Ml.

Chemical- N/A, we can conclude it would have similar results to trials 1 and 2 and would most likely end at 0 Ml.

Results- Unchanged came in at first, in 2nd was the cold temperature and at 3rd was osmotic. Almost every result was surprising as all of them had significant increases in oxygen production compared to trails 1 and 2. Notably the unchanged reaction almost 4 times its previous trial. Osmotic increased from trials 1 and 2 but still remained consistent. The cold temperature reaction slight decrease in oxygen production from the previous trial. The hot temperature reaction also increased by 80 Ml. In the observations we made we noticed that trail 3 had many inflated numbers and many tests had way better results then the previous trials.

Across all three trials are results were slightly inconsistent. A major think we noticed is that on each separate trial, results started to inflate in there numbers. This could have been caused by our lack of experience at the beginning and overtime we started to get better and sealed the flask much quicker. Since, when we look at the average and see the increase in number as the trial number changed we can conclude or techniques got slightly better. Now this is very important because in our experiment we believe averaging the results would be a much better way of showing our experiment instead of viewing each trial separately because of the incontinency in our trials. Across all three trails the environmental stresses had major impacts and wee see this in the form of oxygen rate lowering.

Firstly, Temperature stress clearly demonstrated affecting the enzyme but not very consistently. In Trial 1 we see that hot produced 69 Ml and cold produced 0. In Trail 2 cold jumped up to 160 Ml and hot dropped to 19. Cold had a sudden increase and outperformed every other test. If we don't include the chemical stress in trial 2, hot ends up being in dead last for oxygen produced. In trail 3 cold slightly dropped down and hot ended up producing 99 Ml, its highest so far. In terms of average Cold was in first place and hot was in second last. We can conclude a few things from these results. In trail 1 cold produced no oxygen but its next 2 trials it ended with outstanding numbers, We think the reason is that in trial 1 the temperature was 59.4 degrees F and increased by 4 degrees F over the next 2 trails. Because the temperature was so low in the first trial we can safely assume it was the reason for the poor performance. the final 2 trials for cold both had consistent numbers and fairly close temperatures, this proving our point more. If we look at the hot temperature its much harder to assume the causes of inconsistency. Trail 3 did end up having the closest temperature to the optimal temperature but only by a slight margin. However, we can conclude a few things. Because the enzyme did still end up producing some oxygen we know that it was only partially denatured. We also know that hot temperatures in the environment can actually affect enzymes if we compare the base test to the average of the hot stress.

Secondly, the osmotic stress showed high consistency over all three trails. It was the second most consistent variable and only had a 29 Ml gap between the worst trial and best trial. It still showed the same pattern that I mentioned before. Trial 1 had 87 Ml, trial 2 had 91 Ml, and trail 3 had 116 Ml. That oxygen produced increased every single trial. We can conclude a few things. First, because the consistency was so high and because it had the second best average off all the tests we can assume that osmosis doesn't affect enzyme production very much. As mentioned in the background research, osmosis doesn't directly harm and enzymes structure unlike other stresses. This is one explanation for the great performance but there's one more, Salt is pretty large particle and has a high surface area, especially in a group. When we added the salt we increased the total surface area of the reaction. If the surface area is higher there's more contact between the enzyme and substrate making the oxygen production faster.

lastly, the chemical stress produced 0 Ml of oxygen across all three trials. This strongly shows that copper sulfate is an inhibitor. We can very safely assume that the copper ions interfered with the active site. We weren't able to complete the third trial but the evidence from the first 2 trails was so strong that we are 99 percent sure 0 oxygen would have been produced.

Inconclusion, enzymes are very sensitive to environmental conditions. Chemical can extremely affect enzyme functions. Even with our inconsistency in the cold temperatures we still were able to figure out even the smaller increase or decrease in a temperature affects enzymes. Osmosis was a temporary stress and acted more as a positive then a negative for the enzyme. From our inconstancies in results we show the importance of repeating the experimentation multiple times to study enzyme behavior.

The purpose of this experiment was to see how enzymes behave and respond to natural environment. We used 3 different stresses to recreate the environment which enzymes have to constantly battle. We used a temperature stress including both a hot and cold temperature. We used an osmotic stress by adding larger particles into the solution to create water displacement. Lastly we used a metal ion to create a chemical stress. Our hypothesis was incorrect. We believed because the hotter temperature would allow the reactions to occur quicker that it would end up producing the most amount of oxygen . Instead it fell to 4th place. In 2nd we thought it was going to be the one which was unchanged. It fell down to 3rd place. In 3rd our hypothesis was the osmotic stress which surprisingly rose up 2nd place. In 4th place we stated that it would be the cold temperature which sook us the most because it ended up in first/ The only one we got correct was that the Chemical stress would end up in last place which was what ended up occurring. Even though our hypothesis ended up being very different from our results, the outcomes still made some sense. The cold temperature didn't hurt the enzymes structure, it just slowed the movement down, unlike the other stresses which directly harmed the tertiary structure. Due to this reason it may be the reason it won. The osmotic stress which came in second only had a temporary problem, which was the water displacement caused in the first 30-40 seconds. After this moment everything was stable. The metal ion and hot temperature reactions harmed the tertiary structure the entire reaction which may have been the reason of there poor performances. In Conclusion, this experiment shows how different stresses do affect enzyme activity but they all do it in very different ways. Some were easier for the enzyme to counteract while others were very difficult.

Our topic has a very important application in the real world. Catalase is found in almost every living organism.Enzymes like catalase are in all living organisms because they are speeding up the chemical reaction that happens.Catalase breaks down hydrogen peroxide into water and oxygen,which is protecting the cells from damage. The stresses we added shows us how catalase reacts to changes in the environment and helps us better understand how enzymes actually function,this information is great help in human health and medicine. The results from our experiment help us explain why certain stresses are more effective but we also see which conditions can improve the activity. Hot temperatures can damage the enzyme but the cold doesn't. This can be viewed in 2 ways. High fevers in humans can cause denaturation of the enzymes in your body. Low temperatures may slow the reaction. In agriculture salt can interfere with plant enzymes so osmotic stress should be considered.  We also see how small amounts of metal ions can greatly reduce enzyme activity and act as inhibitors. Copper sulfate in agriculture is also used as a pesticide so high concentration can be very harmful for plants. Scientists also benefit from this information as they see how pollution, climate change,  and environmental changes can cause harm to all life, not just us. 

-First mistake didn't actually affect the results but the timer on the first video is set to 5 minutes. We still ended it after 1 minute and 30 seconds so we will only consider it as a visual error. -We weren't able to complete the third trial for Copper sulfate because we ran out of hydrogen peroxide. Even though we weren't able to conduct it, we concluded it would've ended up at 0ML anyways because of how strong the effect of it really was. We used the info we had and put it to use. When I wanted to test copper sulfate we knew the results would be horrible. We  wanted to show how even the slightest amount could change the course of the enzyme.  -Another mistake we made which did mess up our results was that some tests ended up going over 150 ML. There was an additional gap without any numbers on our syringe and using some measurement we concluded that it was 170 ML in total but on our 3rd experiment for the one that was unchanged ended up going over 170 ML with still 5-6 seconds remaining which we were not expecting to happen. If we were to give an estimate I would say around 180 Ml. There was a total of 90 seconds but it reached capacity at 85-84 seconds so there was 2 ML-2.02ML of oxygen produced every second(2ML/S-2.02ML/S). Using this info we accounted for the missing 5-6 seconds so 2 times 90 and 2.02 times 90 which gave us 180ml-182 ML .The last mistake was we didn't have any gloves at our house which really put me and my moms safety in danger due to the chemicals we were handling.

Biology Online. (n.d.). Enzyme. https://www.biologyonline.com/dictionary/enzyme Cleveland Clinic. (n.d.). Enzymes. https://my.clevelandclinic.org/health/articles/21532-enzymes Creative Enzymes. (n.d.). Effect of temperature on enzymatic reaction. https://www.creative-enzymes.com/resource/effect-of-temperature-on-enzymatic-reaction_50.html Khan Academy. (n.d.). Catalysts. https://www.khanacademy.org/science/ap-chemistry-beta/x2eef969c74e0d802:kinetics/x2eef969c74e0d802:catalysis/v/catalysts Khan Academy. (n.d.). Introduction to enzymes. https://www.khanacademy.org/science/biology/energy-and-enzymes/introduction-to-enzymes Khan Academy. (n.d.). Introduction to proteins and amino acids. https://www.khanacademy.org/test-prep/mcat/biomolecules/amino-acids-and-proteins1/a/introduction-to-proteins-and-amino-acids LibreTexts. (n.d.). Introduction to amino acids and proteins. https://chem.libretexts.org/Bookshelves/Organic_Chemistry/Book%3A_Organic_Chemistry_with_a_Biological_Emphasis_v2.0_(Soderberg)/01%3A_Introduction_to_Organic_Structure_and_Bonding_I/1.04%3A_Structures_of_some_important_biomolecules/1.4.01%3A_Introduction_to_amino_acids_and_proteins National Center for Biotechnology Information. (n.d.). Enzymes. https://www.ncbi.nlm.nih.gov/books/NBK9921/ National Center for Biotechnology Information. (2014). [Article on enzymes]. https://pmc.ncbi.nlm.nih.gov/articles/PMC3988381/ ScienceDirect. (n.d.). [Article on enzymatic activity].https://www.sciencedirect.com/science/article/abs/pii/S101113441500007X

We want to Acknowledge a few people on our science fair. Firstly, We want to acknowledge the friend who was responsible for giving us this topic. He gave us a basic version of this topic so we refined his but we wouldn't be in CYSF if it wasn't for him giving us the idea. Secondly, I want to show thanks to the teachers of Khalsa School Calgary. Our science teacher gave us much help and feedback so our project was the best it could be. other teachers also judged and assessed our project. We also want to acknowledge our parents who spent money on providing us with the equipment and materials we need. They also aided us in our experiment and supervised us so nothing bad occurred.