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Abstract:
The purpose of this experiment was to gauge the effect that temperature had on enzyme activity and the substrate concentration. The rate of enzyme activity had a direct correlation to amount of concentration of the enzyme or substrate. If the concentration of substrate was halved, the enzyme activity rate was also halved. If the concentration of substrate was doubled, the enzyme activity rate was also doubled. The use of environments with fixed temperature acted as an inhibitor or assistance to enzyme activation. The independent variables were the temperature, concentration of substrate, and concentration of enzyme. The sole dependent variable was the reaction rate. My colleagues and I tested the absorbance levels of the enzymes for twenty seconds intervals for a total of two minutes using a spectrophotometer. Results showed that enzyme activity rate had a steady increase until the temperature of 60 degrees Celsius caused denaturation in the protein which showed a lower activation rate than the coldest temperature measured.
Introduction:
Enzymes are macromolecules called proteins. Enzymes act as catalysts within living cells, thus speeding up biochemical reactions by lowering the activation energy required. Each individual enzyme has a structure that correlates to a specific function and an area of activation called the activation site. An activation site is a concave area until a substrate combines with it by filling the gap. Within this specific experiment there were many enzymes (including peroxidase) but we can conclude that peroxidase was the key enzyme used because only a specific enzyme can have a specific reaction with a specific substrate. This makes sense because the definition of substrate is “a substance or layer that underlies something, or on which some process occurs, in particular” the key word being “particular”.
The perfect analogy to explain this process is called the Lock and Key. Imagine you get off from work and attempt put your car keys into your 2003 Cadillac CTS. As your try to insert your key you notice it stops halfway. Although your car is a 2003 Cadillac CTS the car you’re attempting to unlock is not yours. An everyday example is when rennin (enzyme) is added to liquid milk proteins (substrate) that produce coagulated milk solids known as curd (product). The main takeaway from these examples above is that only a compatible enzyme can work with a substrate. In this experiment we used an enzyme extract of horseradish called peroxidase which is acquired by homogenization in buffer. These broken cells release many enzymes including peroxidase. To test for peroxidase we had to mix the extract with the compound guaiacol. Guiacol is normally colorless but becomes brown after oxidation. Peroxidase Combined with 25 ml of 0.1M Citrate Phosphate buffer and the substrate hydrogen peroxide (H2O2).
There are many factors that can increase enzymes but in this lab we only focus on temperature. Similarly to people, enzymes work optimally in a desirable temperature. More so heat is used to increase the kinetic energy of the surrounding molecules which result in a greater number of collisions between said molecules and reactions to happen quicker. In colloquial terms a higher temperature can increase reaction rates while a lower temperature will decrease reaction rates. Because enzymes are proteins they are affected by denaturation which causes a protein to breakdown.
Temperature has a measurable effect on enzyme activity rates within the spectrophotometer. As temperature decreases so does activity rates. As temperature increases activity rates increases. This holds true until the high enough temperature creates an unsatisfactory for an enzyme and causes denaturation. Enzymes are similar to humans in this case. Human beings work optimal at 34 degrees Celsius (body temperature), but a lower or high body temperature can affect output negatively.
Description of experiment:
The production of this experiment began with the extraction of peroxidase from a horseradish, 25 mL 0.1M cirate-phosphate buffer, and a blender jar. We cut and weighed 1.0 gram of the horseradish and placed it in the blender for twenty seconds. After it was well blended we used a double layer cheesecloth to pour the contents into a beaker (except for the chunks of horseradish). For the next steps we labeled the enzyme concentration and a graduated cylinder with the word “buffer” that held citrate-phosphate ph 5 and then used two pump dispensers of hydrogen peroxide and guaiacol solutions into 9 labeled test tubes. Test tubes labeled #2 and #3 were placed in an incubation chamber of 5 degrees Celsius. Test tubes labeled #4 and #5 were placed in incubation chambers of 25 degrees Celsius. Test tubes labeled #6 and #7 were placed in incubation chambers of 34 degrees Celsius. The final test tubes labeled #8 and #9 were placed in an incubation chamber of 64 degrees Celsius. Once the test tubes reached an incubation time of ten minutes they were put through the spectrophotometer to calculate absorbance rate. Before calculating the absorbance rate the spectrophotometer was set to a wavelength of 500. We began to write down absorbance rate for twenty second intervals as soon as the enzyme and substrate were mixed for a total time of two minutes. After writing the absorbance rate for each twenty second interval.

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Abstract. (2019, Mar 22). Retrieved January 17, 2021, from https://midwestcri.org/abstract-4/

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