Enzymes and Temperature

Zaquia Austin Enzymes and Temperature Laboratory 6 3/13/2013 mean (Objectives) The purpose of this examine was for students to be able to understand the enzyme-facilitated reaction and explain how enzyme activity crumb be affected by environment conditions. Abstract This experiment mainly delt with enzymes and the reaction that enzymes reserve with different solutions in various temperatures. Three different exercises were d unitary. The foremost exercise had to do with enzymes and temperature. During this exercise cardinal different sieve renders were used for one-third different temperates. The first analyze tube which involved 0 ?C weewee bath had a total novelty of amylum to sugar in the first 15 consequences, and the secant discharge tube which involved 37? C irrigate bath had a total conversion of amylum to sugar after the first quin sharps. While the third experiment tube took a unforesightful longer. The second exercise had to do with enzymes and substra te concentration. This exercise took longer for the all of the starch to be removed from the stress tubes, and it involved 37? C water bath for all fiver test tubes. The last exercise had to do with enzymes and pH. This exercise involved four test tubes all put into a water bath of 37? C.The fourth test tube had a total conversion of starch to sugar in the first five minutes while the other three took a little longer to turn. Experiment & Observation branch the water baths and the 1% Alpha-amylase was prepared. Three different water baths were prepared. one and lonesome(prenominal)(a) being set at 0? C, the next being set at 37? C and the last one being set at 100? C. Next a 96-well microplate was marked with multiplication and numbers based on the test tubes number. indeed using a pipet one mould of iodin was placed into each well (1,2,3) up to total of 30 minutes. whence the test tubes were marked 1cm and 6cm from the bottom. by and byward, 1cm of alpha-amylase was pipe tted into each test tube. Then, test tube 1 was incubated at 0? C, test tube 2 was incubated at 37? C, and test tube 3 was incubated at 100? C for five minutes. After that 1% starch solution was added to the 6cm mark. Next it was put back into its assigned water bath for some other 5 minutes. Then deuce drops of solution was put into its corresponding number on the microplate in the 5 minute row. The color was immediately recorded. The steps were repeated for a interval of 30 minutes or until each well had an amber color in it.Test tube 3 did not change inside the 30 minute interval so it was placed into the 37? C water bath for other 30 minutes. Then one drop of solution and one drop of iodine was added into the well. There was unbosom no color change, so another 1cm of alpha-amylase was added to it and was incubated for another 30 minutes, the color became amber. Table 1. import of Temperature on Amylase Enzyme Conversion of Starch to Sugar Time/Minutes Test Tube 1 / 0? C Test Tube 2 / 37? C Test Tube 3 / 100?C 5 + - ++ 10 + ++ 15 - ++ 20 ++ 25 ++ 30 ++ 60 ++ 90 - The color of iodine that indicated that starch was still present in the test tube was the Black/ Blue-Black color. The color that indicated that the starch was gone was the Amber color. Amylase breaks up the starch which makes it disappear, it does not react with iodine anymore. The variable in this experiment can be an independent variable. The change in the experiment is the starch to sugar. The temperature that is optimal is 37? C.The temperature that facilitated first was the 37? C, then it was 0? C. No 37? C was chosen as the mid-range temperature because that degrees in Fahrenheit(postnominal) is 98. 6? F. It was important to immediately observe the color because after a while it changes colors again. If you would be possessed of just added the iodine in test tubes that would have ruined the entire experiment because there wouldnt be anything to test if it didnt work the first time. Once the test tube was incubated at 37? c null happen. The starch did not disappear, maybe because there wasnt enough amylase. After 1cm of alpha-amylase was added and incubated at 37? C the starch eventually disappeared.This probably didnt happen the first time because the more amylase there is the better chance of it shift up the starch. pic My initial hypothesis was that the more starch solution there is the longer it will wee for it to break down amylase. My hypothesis was actually supported, there was more starch then amylase, when there undeniable to be more amylase then starch. I learned that amylase breaks up starch, and that its an digestive enzyme. Exercise 2 First a water bath was set at 37? C. Then the microplate was label this time five test tubes were used. One drop of iodine was placed into each well (1,2,3,4,5) up to 30 minutes. Then 1/2cm was marked from the bottom of the test tube. Next for each tube an additional cm was added.For tes t tube one 2cm above the bottom, test tube two 3cm above the bottom, test tube three 4cm above the bottom, test tube four 5cm above the bottom, test tube five 6cm above the bottom. Then 1/2 alpha-amylase was added to the 1/2 cm mark on the test tube. Afterwards the test tubes were placed into the water bath for five minutes. Next 1% starch solution was added to the next cm mark on the test tubes. The test tubes were put back into the water bath for another five minutes. Then two drops of the solution was added to each of the corresponding wells. This was done for all five test tubes. presently record the colors. The steps were completed for an interval of 30 minutes.The tubes that had not changed color within the 30 minute interval was put back into the water bath for another 30 minutes. Two drops of solution and one drop of iodine was put into the well and the amber color was recorded. Table 2 The Effect of compactness on Amylase Enzyme Conversion of Starch to Sugar Time/Minutes Tube 1 Tube 2 Tube 3 Tube 4 Tube 5 Concentration of Amylase 0. 5cm/2cm 0. 5cm/3cm 0. 5cm/4cm 0. 5cm/5cm 0. 5cm/6cm Per test tube 25% 17% 12. % 10% 8% 5 ++ ++ ++ ++ ++ 10 ++ ++ ++ ++ ++ 15 ++ ++ ++ ++ ++ 20 ++ ++ ++ ++ ++ 25 ++ - ++ - ++ 30 ++ ++ ++ ++ ++ 60 ++ - - The variable in this exercise the substrate concentrations. Test tube 3 This experiment could be alter if each test tube had its own water bath.My hypothesis was that test tube five would convert to sugar first. The reason I hypothesized this was because test tube five had the most starch. In this exercise I learned that enzymes can be used over and over again to facilitate the conversion of substances beforehand they are denatured. Some practical applications can be Food and Beverages D. Another way this experiment could be done is by using different temperatures of water baths. Exercise 3 First a water bath was set at 37? C. The microplate was labeled, this time only four test tubes were used. Each t est tube was marked 1cm, 2cm, and 4cm from the bottom. Next one drop of iodine was added to the weel (1,2,3,4) and u to a 30 minute interval.Then a different pH pilot film was added to each test tube at the 1cm mark. For test tube one pH 3. 5 buffer was added, for test tube two pH 5 buffer was added, for test tube three pH 6. 8 was added, and for test tube four pH 11. 5 was added. Afterwards, 2cm of alpha-amylase was added. Then the test tubes was placed into the water bath to be incubated. After five minutes starch solution was added to the remaining 4cm mark, then placed back into the water bath. After five minutes two drops of solution was put into each corresponding well. Immediately record color. The steps were completed for an interval of 30 minutes. The fourth test tube showed color immediately, but the other three test tubes did not.Table 3 The Effect of pH on Amylase Enzyme Conversion of Starch to Sugar Time/ Minute Test Tube 1 Test Tube 2 Test Tube 3 Test Tube 4 pH 3. 5 5. 0 6. 8 11. 5 5 ++ ++ ++ - 10 ++ ++ ++ 15 ++ ++ ++ 20 ++ ++ ++ 25 ++ ++ ++ 30 ++ + + 60 + + + The variable in this exercise is the pH. Only the fourth test tube converted starch to sugar, I think this happend because it had an higher amount of buffer. Yes the first three test tube did not point a change in color.. There pH was much lower then the last one. I hypothesized that the test tube with the pH buffer would convert to sugar first. My hypothesis was refute. completion Temperature, and the times of incubation.You can get different substrates if you change the incubation times. In this laboratory I learned how to recognize enzyme-facilitate reactions, and how to submit when starch is converted into sugar. Some practical applications could be yeast, detergent, leather and bioethanol. Discussion/Error Analysis/Conclusion The first part of this laboratory was based upon testing the alpha-amylase enzyme activity on starch under three temperature environments , 0? C, 37? C, and 100? C. The next part was to demonstrate the effects of substrate concentration on enzyme reactivity. The last exercise was based upon testing how alpha-amylase functions at four different pH levels (3. ,5,6. 8, and 11. 5). A hardly a(prenominal) errors that occurred was one, the changing of the water bath temperatures. If the bath stayed at a constant temperature then it probably would have made a dissimilitude to some of the tubes. Another laboratory error could have been that there were only a select few of pipets. If there were pipets for each exercise that could have made a difference even though the pipets were cleaned after each experiment, it still would have made a different if it was a clean dry pipet. Another laboratory error could have been the incubating times. These errors could be minimized in the succeeding(a) if there were a few arrangements before hand.