October 29, 2013
This experiment it designed to assist one in becoming acknowledged with enzymes. They will then learn why these enzymes are important and why it is used often in organic systems. The lab will also give them the chance to look at the activity rates of the enzymes and determine how it can be affected by environmental conditions. These conditions include temperature, pH, and substrate concentration.
The process of metabolism can be interesting yet very complex. It takes certain substances like enzymes to get them going. As catalysts, enzymes facilitate reactions that normally cannot occur under biological conditions. They increase the speed of reactions and or lower their activation levels. Several factors can influence the shape of an enzyme. These include the pH, temperature, and substrate concentration. The temperature in an enzymatic reaction dictates the enzymes rate of reaction. It was noted that the 100° C water bath after every five minutes gave the same results; starch was present.
This is because when the temperature increases, the enzymes denatured making the starch presentable. Since enzymes thrive at optimal pH ranges of 6-8, increasing these values can also cause them to denature. When the concentration in a solution is low the enzymatic activity ends up slowing down causing the conversion of starch to sugar occur at a slow rate. All in all this lab lets one see that without enzymes, more energy would be required to power the breaking down and building up of foods.
Experiment and Observation (Data Collection)
Enzyme and Temperature
12 mL of distilled water was added to alpha-amylase powder to make a 1% solution. Three water baths at 0ᵒ C, 37ᵒC, and 100ᵒC were made. Using a strip of tape, the 96-well microplate was labeled 1-3 on the short side to correlate with the test tubes. The long side contained the numbers 0, 5, 10…30, 60, and 90 to represent minutes. Three pipettes were label 1-3 to correspond with the test tubes. A drop of IKI indicator (iodine), with a clean pipette, was placed into each microplate well marked 0 to 30 minutes and 1-3 sides, which also coincided with the test tubes and covered with a plastic wrap. Three test tubes were obtained and labeled 1-3. These tubes, with a marker pencil and a metric ruler, became identified with 1 cm and 6 cm from the bottom. The amylase solution, along with the pipette filled the 1 cm mark on each test tube.
Test tube 1, along with its corresponding pipette, was incubated in the 0°C water bath. Two was incubated in the 37°C bath and three in the bath marked 100°C. After five minutes, another 1% solution of starch was added to each test tube to the 6 cm mark. Once mixed, two drops of the solution was placed into the row marked “0” for zero minutes, and the 1, 2, 3 columns. Immediately, the color change was observed. The tubes were returned back into the original water baths and after five minutes two more drops were taken out and put into the corresponding columns and the 5 minute row. Once again the color change was noted.
This step was repeated every five minutes for 30 minutes until the drops from the test tube solution turned amber. Once amber, which ever test tube yielded the no starch present was discontinued. However, after 30 minutes if the tube did not change to amber, test tube 3, it was placed into the 37°C water bath for 30 minutes. The presence of starch was tested by placing one drop of IKI indicator and two drops of the solution in the test tube in the related 60-minute well row. The color was again recorded. If the testing revealed that the tube still contained starch, another 1 cm of amylase was added and allowed to sit in the 37° water bath for another 30 minutes. This was retested in the 90-minute well row.
Enzyme and Substrate Concentration
The same steps were taken for this testing except there were five test tubes, each with