BIO161 Lab Report #2 PDF

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Lab Report #2
Biological Catalysts: Enzymes
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Lab Report #2 Biological Catalysts: Enzymes

Overview: Biological Significance: The biological significance of this lab is that enzymes play an important role that help chemical reactions in the cell happen and that they can be altered by changes in either temperature or pH. This can helpful by changing the enzymes temperature or pH to either inhibit it from reacting or to make the enzymes perform more efficiently and rapidly.

Hypothesis: Due to a change in temperature or pH, an enzyme’s conformation and reaction rate is altered, thus making the enzyme to become denatured. By increasing the temperature, the enzyme’s reaction rate increases and by increasing or decreasing the pH, the enzyme’s reaction rate decreases.

Methods: (Describe how the biological materials and lab tools are used to answer questions) Peroxidase experiment: In the lab an enzyme, peroxidase, was extracted from a turnip, for the purpose of determining the effect of temperature and pH on the activity of the enzyme. This was done so by exposing the chemical reaction H2O2 + guaiacol- ⟶ H2O + guaiacol, aided by peroxidase, to different temperatures and pH values and recording the absorbance of the sample using a spectrophotometer at every 20 seconds for 120 seconds. For the temperature portion, a total of 11 test tubes were used and each tube contained a certain amount of buffer, H2O2, peroxidase extract, and guaiacol. The first tube was the blank, and the following tubes varied in temperatures, from 4°C,  23°C, 32°C, 48°C, and 100°C (boiled). In addition for the pH portion, 9 test tubes were used, each one also containing a certain amount of solution. Similarly, the first test tube was the blank, and the rest varied in pH values, from pH of 3, 5, 7 and 9. The enzyme’s reaction rate was assessed by calculating the slope for each temperature and pH value with its absorbance over the amount of time in seconds.

Popcorn experiment: In lab Clintest tablets were used to determine if saliva contains enzymes that convert starch that is found in popcorn into sugars. Clintest tablets are used for the fact that change color in the presence of simple sugars. An iodine solution (I2KI) that changes color when introduced to starch was used to make sure that the popcorns had starch on them, Five paper cups were used to test every component of the test, One of the paper cup contained a mL of distilled water, crushed up popcorn and

drops of iodine solution this cup was to make sure that starch is present in the popcorn. The second cup contained mL distilled water and a clintest tablet to make sure that the water did not have sugar in it. The third cup test for sugar in the saliva provide by containing a mL of saliva and Clintest tablet. The fourth cup tested to see if the popcorn had sugar. This was done by crushing up a popcorn and adding one mL of distilled water and one clintest tablet. The fifth and final cup was the actual test to check if saliva had an enzyme that converts starch into simple sugars, which was accomplished by the cup containing a popcorn that was chewed for one minute and one Clintest tablet.

Results: - see report booklet (the 4 graphs are separate)

Discussion/Conclusions: Part 1: ( type discussion/conclusions concerning effect of temperature (including boiling) and pH on enzymes.) Enzymes are proteins that are critical to catalyzing reactions. They react with certain substrates in order to speed up the rate of reaction within the cell. These catalysts are produced by the cells in very small amounts and are not completely consumed during a chemical reaction. Without enzymes, reactions would be significantly slower and basic functions such as breathing or digesting food, would not be possible. In this experiment, the variables in which denature an enzyme were observed and recorded. Denaturation in an enzyme occurs when conditions in the environment, like temperature, pH and ionic concentration leads to a change in its conformation and enzyme activity. This experiment focuses on the influence temperature and pH have on the reaction rate of an enzyme. As the temperature increases, so does the rate of reaction and in extremely high temperatures the enzyme becomes denatured. The enzyme’s shape changes and the substrate no longer fits into the active site. An enzyme which has become denatured is permanently inactive and will no longer react. Based  off the results on graph 3, this is exactly what happens. At 100°C, the reaction rate of the enzyme is zero meaning it is not active and it is not taking part of the chemical reaction. The results support the hypothesis made regarding that with temperature changes, the enzyme’s functionality is also changed; high temperature increases the rate of the enzyme’s reaction and at really high temperatures (above 32°C)  the enzyme’s conformation is completely transformed that it no longer reacts. Additionally, pH is a measure of the concentration of hydrogen ions in a solution. The more hydrogen ions present, the lower the pH of the solution. Enzymes function efficiently over a specific pH range. A change in pH, whether it is above or below this range, decreases the enzyme reaction rate. It is evident in the results collected (graph 4) that a pH of 3 and 9 reduces its reaction rate. Therefore concluding that extremely  high or low pH values result in a loss of enzyme activity.

Part 2: ( type discussion/conclusions concerning identification of enzyme(s) in saliva – explain role of each cup (#1-5) and if your results allow you to make any conclusions on presence/absence of a salivary enzyme.) Clintest tablets are used to determine whether or not sugars are present. Whenever there are

sugars present, the tablets will turn the mixture into a color that is different from a dark blue. In the first cup, a kernel popcorn, distilled water and iodine was mixed together. The purpose of this cup was for the cup to act as a positive control for starch. The second cup consisted of distilled water and a Clintest tablet. The tablet dissolved turning the water a dark blue color which concluded that starch and saliva were not present in the cup. The third cup was made up of saliva and a Clintest tablet. This cup’s role was to be a negative control for starch. In the fourth cup, a kernel popcorn, distilled water, and a Clintest tablet was combined. The purpose of this cup was to prove that water does not play a factor in the conversion of starch to simple sugars. Lastly, cup five consisted of a chewed kernel popcorn and a Clintest tablet. When the tablet was added, the mixture immediately turned a dark golden color which shows that the saliva did play a role in starch conversion to simple sugars. Therefore, our hypothesis regarding saliva containing an enzyme that changes starch into simple sugars was supported.

Responses to Review Questions:

4) Offer an explanation of why we use refrigerators. In other words, explain why a partially eaten hamburger gets moldy after a week if left on the kitchen counter, but takes much, much longer to get moldy if kept in a refrigerator. Note that the temperature of most refrigerators is 4°C.

The reason a half eaten hamburger left on the counter gets moldy after a week is because it is exposed to the environment, where if any of its ingredients that can oxidize will and any bacteria that made it way the the hamburger ingest. When the hamburger is placed inside the refrigerator the bacteria and more importantly the enzymes in the cells that cause the burger to mold at a higher rate are placed in a temperature that inhibits them to cause the reaction of it becoming moldy to slow down a substantial amount. This is due to the fact that enzymes can only function within a range of temperature.

5) If you cut an avocado, plant cell defense reactions lead to the production of certain compounds. These compounds impart a brown color. Explain why adding lemon juice (has a very low pH) to guacamole prevents the guacamole from browning.

The reason that avocados turn a brown color is that plants cells defense reacts to the exposure with the air , this process it aided by an enzyme. When lemon juice is added it hinders the enzymes, this is due to the enzymes having regions in which they can function relating to pH and the the lemon juice alters the

pH of the avocados and the cells within. The low pH (acidic) of the lemon juice slows down or possibly denatures the plant cell’s defense that avocados have, thus preventing it from browning. The same can be achieved if the avocado was place in a temperature where the enzymes are out of the range in which they can function properly.

9) What conclusion could be made if the popcorn plus water test resulted in the same Clinitest reaction products as that with the popcorn plus saliva?

If the popcorn plus water test resulted in the same Clinitest reaction products as that with the popcorn plus saliva test, it would reject the hypothesis. The hypothesis states that since saliva contains an enzyme that converts starch into simple sugars, simple sugars will only form in the presence of saliva. If the Clintest reaction products were the same in the water test, it can be concluded that water also contains an enzyme that converts starch into simple sugars....