How pH Affects the Break Down of Starch by the Enzyme Amylase
Hypothesis:
The optimum pH for the reaction of starch with amylase is pH 7. PH values lower or higher than this value will result in a slower rate of reaction. Amylase works in the range pH 3 to pH 11.
Biological Knowledge ====================
PH changes affect the structure of an enzyme molecule and therefore affect its ability to bind with its substrate molecules. Changes in pH affect the ionic bonds and hydrogen bonds that hold the enzyme together, which naturally affects the rate of reaction of the enzyme with the substrate. On top if this, the hydrogen ions neutralise the negative charges of the R groups in the
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7 Corvettes
These are used instead of test tubes as corvettes are required if a colorimeter is to be used. One corvette is needed for each pH and one for the control. These can then be reused for the repeats.
Syringe calibrated at 0.1 cm³ intervals
This is vital to measure out the volumes of the starch and amylase solutions accurately using these calibrations to achieve precision. This apparatus is far more accurate than using a measuring cylinder, as there are minimal transference losses.
2 Syringes
As the volume of buffer solution of iodine is not being measured, only the number of drops, apparatus with calibrations is not required for this.
Stopclock
This is important for measuring the time. It is not important whether a digital or traditional stopclock is used as there will be the same percentage error either way.
Method
* Measure out 1 cm³ amylase solution with calibrated syringe and put in corvette
* Add to this 5 drops of pH 4 buffer solution
* Measure out 2 cm³ starch solution
* Start stopclock and leave for 1 minute
* Measure out 1 cm³ amylase and place in second corvette
* Add to this 2 cm³ distilled water
* Add 3 drops of iodine solution
* Shake it well,
During these experimental procedures, the implication of multiple different temperatures on fungal and bacterial amylase was studied. In order to conduct this experiment, there were four different temperatures used. The four temperatures used were the following: 0 degrees Celsius, 25 degrees Celsius, 55 degrees Celsius, and 80 degrees Celsius - Each temperature for one fungal and one bacterial amylase. Drops of iodine were then placed in order to measure the effectiveness of the enzyme. This method is produced as the starch test. The enzyme was tested over the course of ten minutes to determine if starch hydrolysis stemmed. An effective enzyme would indicate a color variation between blue/black to a more yellowish color towards the end of the time intervals, whereas a not so effective enzyme would produce little to no change in color variation. According to the experiment, both the fungal amylase and bacterial amylase exhibited a optimal temperature. This was discovered by observing during which temperature and time period produced a yellow-like color the quickest. Amylase shared a similar optimal temperature of 55 degrees Celsius. Most of the amylases underwent changes at different points, but some enzymes displayed no effectiveness at all. Both amylases displayed this inactivity at 0 degrees Celsius. At 80 Celsius both the enzymes became denatured due to the high temperatures. In culmination, both fungal and bacterial amylase presented a array of change during it’s
enzymes that will be used during this lab to test the ability of amylase to break down starch ,a
These results shown from this experiment led us to conclude that enzymes work best at certain pH rates. For this particular enzyme, pH 7 worked best. When compared to high levels of pH, the lower levels worked better. The wrong level of pH can denature enzymes; therefore finding the right level is essential. The independent variable was the amount of pH, and the dependent being the rate of oxygen. The results are reliable as they are reinforced by the fact that enzymes typically work best at neutral pH
This experiment consisted of setting up a control group of starch in various temperature and then placing both fungal amylases and bacterial amylases in a mixture of starch and placing the solution of amylase and starch in various temperatures of water. After a certain amount of time- different amount of time needs to be used in order to have reliable results- iodine is added in a well on spot plates, then two drops of the mixture of amylase-starch is added from each temperature used, by adding iodine into the plates the mixture will show how much starch was hydrolyzed, this is used to calculate the amount of
For this experiment, we have to prepare our phosphorylase which extracted from a potato. We prepared by weighed about 250 grams of peeled potato and cut it into cubes. The extracts then blended with 100mL of 0.1M NaF. After filtered the contents into a clean 250mL centrifuge bottle, we centrifuged it for 3 minutes. Then, separated the supernatant into a centrifuge bottle, which is our phosphorylase preparation. The enzyme assay used in this experiment today is the iodine test. As the iodine reacts with starch, it will form a brown, blue or black precipitate due to the iodine ions forcing into a linear arrangement. The endpoint of the enzyme reaction indicates the presence of starch by using the iodine test to determine. The faster the endpoint is reached, the less active the phosphorylase is.
Add the same amount of Benedict’s solution in each tube and for the ones that need iodine use the same amount.
In this experiment we wanted to determine the optimal temperatures for fungal, Aspergillus oryzae, and bacterial, Bacillus licheniformis. In order to see if any of the starch was broken down, Iodine was mixed with the starch-amylase substance. In four spot plates, the groups labeled the different temperatures, once the iodine came in contact with the starch, the result would be a reaction that turns the fluid into this dark blue/black color. In a span of 10 minutes, with occasional check ups on the solution every 2 minutes, the amylase-starch solution was placed into five types of temperature, all being Celsius. The five temperatures were 0 degrees, 25 degrees, 55 degrees, and 85 degrees Celsius. The solution would change colors, so in order to measure the changes, a scale was used. Such scale was a 1-5 scale, with colors next to each number. One being the lightest color, or yellow, and 5 being the darkest color, or black. Based on the change of color, we could tell how fast it hydrolyzed the starch in a span of 10 minutes. To keep record of the results, the results were put in Data Tables used from the Lab Manual. The average optimal temperature for Bacteria Amylase was 85 degrees Celsius, while the Fungal was 55 degrees Celsius. You can see this by looking for the
Amylase experiment # 2 was done to see how the pH affected the efficacy of the enzyme. First we collected all of the materials that were necessary to make this experiment. We needed five clean test tubes, the following standard solutions, 1% Starch Solution pH 3,1% Starch Solution pH 5,1% Starch Solution pH 7,1% Starch Solution pH 9,1% Starch Solution pH 11
test the pH of the amylase a drop of the solution should be put on pH
Enzymes are high molecular weight molecules and are proteins in nature. Enzymes work as catalysts in biochemical reactions in living organisms. Enzyme Catecholase is found on in plants, animals as well as fungi and is responsible for the darkening of different fruits. In most cases enzymatic activities are influenced by a number of factors, among them is temperature, PH, enzyme concentration as well as substrate concentration (Silverthorn, 2004). In this experiment enzyme catecholase was used to investigate the effects of PH and enzyme concentration on it rate of reaction. A pH buffer was used to control the PH, potato juice was used as the substrate and water was used as a solvent.
The practical was carried out to investigate the effect of pH on the reaction of the enzyme acid phosphatase.
(6.2)Material and Methods in the process or exercise of measuring the starch we were used the following material and how we used them to conduct the experiment. Obtain seven tubes the material to be tested table 6.1 and then add seven to ten drops of iodine to each tube, and then record the color of the tubes contents in table 6.1
Measure and add 5cm3 of buffer solution using a measuring cylinder with the pH 3 into a test tube using a pipette and place the potato cylinders into the test tube.
Amylase is an enzyme that is in human’s saliva as well as the pancreas. Enzymes are biological catalysts that speed up a chemical reaction. They break down complex molecules into simple ones. In this case, amylase converts starches (complex molecule) into simple sugars. That is why foods like potatoes for example, may taste sweet to us, because they contain starch. The optimum pH for pancreatic amylase is the pH of 7. In the experiment I have used buffer solutions with the pHs of 2.8, 4 and 6.5. I have also used iodine and starch. Normally, iodine is orange-yellow, however when you add starch to it, the solution will turn blue-black.