In this experiment, in the first part, the best concentration of enzyme was determined by recording the absorption over time. In the second part, the best concentration was selected from the previous experiment which was C and the optimum pH was determined. Introduction In an article, ‘Lab Presentation’ on Prezi, the peroxidase enzyme was tested on factors such as enzyme concentration and pH. It was measured using guaiacol as it turns orange when oxidized; spectrophotometer was used to determine the rate of absorbance. The article states that as the peroxidase enzyme concentration increases, the rate of reaction increases as well. This happens because the more the enzyme concentration, the more substrates will be catalyzed by peroxidase. …show more content…
An assay determines the enzyme activity. Guaiacol and hydrogen peroxide are key factors for this experiment. As the enzyme breaks down hydrogen peroxide, it gives out hydrogen and oxygen. Guaiacol turns the solution brown in the presence of oxygen, so as the oxygen was given out from the breakdown of hydrogen peroxide the solution turned brownish which proved that the enzyme was reacting. In the first part of this experiment, the concentration of the enzyme was found. In four test-tubes, labelled A-D, each test-tubes were diluted starting from the the test-tube A which had 5ml enzyme stock solution. 1 ml of the enzyme was added to the test-tube B containing 4ml buffer, which was 1:5 dilution after the mixture. 1 ml of the solution from test-tube B was then added to test-tube C, which then had 1:25 dilution. The same procedure was repeated with test-tube D, which had 1:125 dilution. Then, nine test-tubes were taken, out of them one was blank. The rest eight test-tubes had different volumes of buffer, enzyme, hydrogen peroxide and guaiacol. Guaiacol is a dye used to determine the presence of oxygen, which turns the solution brown if oxygen is present. Mixtures of two test-tubes were added and labelled A-D accordingly. The absorption rate of the four test-tubes were then determined over a total time of 120 seconds, having a gap of 20 seconds within the 120 seconds. The more dilute the solution was, the less was the absorption, which …show more content…
Test-tube C had the best concentration according to the results. Three test-tubes were labelled A-C. Test-tube A had 1ml enzyme solution which was added to test-tube B which had 4ml buffer (pH 5 was used). 1ml of the solution from test-tube B was then added to the test-tube C which also had 4ml buffer (pH 5). Test-tube C was used as the enzyme in all the reactions. Nine test-tubes were taken out of them one was used as the the blank, labelled as test-tube 9. The blank had 5ml buffer (pH 5), 2ml hydrogen peroxide, 1ml guaiacol and no enzyme. Then, 3ml of buffer (pH 3) and 2ml of enzyme were added to test-tube 1. Test-tube 2 had 2ml hydrogen peroxide and 1ml guaiacol. Test-tube 1 and 2 were mixed. The same procedure was used for test-tube 3 as test-tube 1, but this time the buffer was pH 5. Test-tube 4 was prepared the same way as test-tube 2. Then, Test-tube 3 and 4 were mixed. Test-tube 5 was prepared as test-tube 1 but with buffer of pH 7 and test-tube 6 was prepared as test-tube 2. Next, test-tube 5 and 6 were mixed. Last but not the least, test-tube 7 was prepared as test-tube 1 but with buffer of pH 9 and test-tube 8 was prepared as test-tube 2. Then, test-tube 7 and 8 were mixed. The spectrophotometer was set to 470nm and using the blank it was set to zero. The four test-tubes with different pH’s (pH 3, pH 5, pH 7, pH 9) were read
This yellow species can then be measured using UV absorbance (max abs = 420 nm), and thus the concentration of the can species determined.1 Horseradish peroxidase in important in the glucose assay because it catalyzes a reaction that includes one of the products from the glucose oxidase reaction, H2O2. There will be one H2O2 produced for every oxidized B-D-glucose, which will then be used to oxidize one ferrocyanide into the one measurable ferricyanide. Therefore, using the enzymes glucose oxidase and horseradish peroxidase in a consecutive manner, users can determine the concentration of glucose present in solution by simply measuring the amount of ferricyanide produced because of it (this is a one to one ratio).
After conducting this experiment and collecting the data I would have to say that the optimal temperature for enzyme activity would have to be room temperature which in my experiment was thirty-four degrees Celsius. I came to this answer because the glucose test strip showed that at room temperature there was more glucose concentration that at either of the other temperatures. Due to temperature extremes in the boiling water the enzymes could no longer function because the breakdown of lactose stopped. The cold water also hindered the breakdown of the lactose but as the water warmed the enzymes were more active which can be seen in the results for the cold water at 20 minutes B. Describe the relationship between pH and the enzymatic activity of lactase.
However, at 3% substrate concentration, the hydrogen peroxide decomposition showed an immediate peak of up to 3.8 mm in height. As the substrate concentration slowly increased, enzyme
The purpose of this study is to analyze the activity of the enzyme, catalase, through our understanding
Catalase is a common enzyme that is produced in all living organisms. All living organisms are made up of cells and within the cells, enzymes function to increase the rate of chemical reactions. Enzymes function to create the same reactions using a lower amount of energy. The reactions of catalase play an important role to life, for example, it breaks down hydrogen peroxide into oxygen and water. Our group developed an experiment to test the rate of reaction of catalase in whole carrots and pinto beans with various concentrations of hydrogen peroxide. Almost all enzymes are proteins and proteins are made up of amino acids. The areas within an enzyme speed up the chemical reactions which are known as the active sites, and are also where the
For example, incubating the samples at different temperatures would create more data points to establish an optimal temperature. From the results in the experiment in this study, it is known as temperature increases, enzymatic activity increase, and vise versa. However, what can not be observed is at what point does the increase in temperature begin to denature the enzyme, above 60°C. Furthermore, assays can be preformed to determine optimal pH, as well. From Dutta’s, and his partners, experiment it shows that there is a range where the Heliodiaptomus viduus’s lactase shows the most activity, which is between 5.0 and 6.0
Finally, the last part of the experiment examined the enzyme activity at different pH levels. Four sets of 11 tubes were set up in this part. The procedure for this part is the same as before, but 4 other buffers were substituted for the standard pH 7.3 phosphate buffer. Set A used the 5.5 pH buffer while set B used the 6.5 pH buffer. The buffer of pH 8.5 was used for set B and for set D the pH was 9. The absorbance readings for 4 sets were taken and recorded in table 13. Using the linear equation that the best-fit line gave for each set, the Km and the Vmax of each set were determined. Then, table 15 was made by dividing the Vmax by the Km. of the four pHs. The Vmax and Km of the control set were also used to make
The purpose of this experiment was to discover the specificity of the enzyme lactase to a spec...
The independent variable for this experiment is the enzyme concentration, and the range chosen is from 1% to 5% with the measurements of 1, 2, 4, and 5%. The dependant variable to be measured is the absorbance of the absorbance of the solution within a colorimeter, Equipments: Iodine solution: used to test for present of starch - Amylase solution - 1% starch solution - 1 pipette - 3 syringes - 8 test tubes – Stop clock - Water bath at 37oc - Distilled water- colorimeter Method: = == ==
In this experiment as a whole, there were three individual experiments conducted, each with an individualized hypothesis. For the effect of temperature on enzyme activity, catalase activity will be decreased when catalase is exposed to temperatures greater than or less approximately 23 degrees Celsius. For the effect of enzyme concentration on enzyme activity, a concentration of greater or less than approximately 50% enzymes, the less active catalase will be. Lastly, the more the pH buffer deviates from a basic pH of 7, the less active catalase will be.
Investigating the Effect of Substrate Concentration on Catalase Reaction. Planning -Aim : The aim of the experiment is to examine how the concentration of the substrate (Hydrogen Peroxide, H2O2) affects the rate of reaction. the enzyme (catalase).
2) A small amount is required as the enzyme is used again and again as
From looking at the results I can conclude that when the pH was 3 and 5. No oxygen was produced, therefore no reactions were taking place. This was because the pH had a high hydrogen ion content, which caused the breaking of the ionic bonds that hold the tertiary structure of the enzyme in place of the syringe. The enzyme lost its functional shape.
Abstract: Enzymes are catalysts therefore we can state that they work to start a reaction or speed it up. The chemical transformed due to the enzyme (catalase) is known as the substrate. In this lab the chemical used was hydrogen peroxide because it can be broken down by catalase. The substrate in this lab would be hydrogen peroxide and the enzymes used will be catalase which is found in both potatoes and liver. This substrate will fill the active sites on the enzyme and the reaction will vary based on the concentration of both and the different factors in the experiment. Students placed either liver or potatoes in test tubes with the substrate and observed them at different temperatures as well as with different concentrations of the substrate. Upon reviewing observations, it can be concluded that liver contains the greater amount of catalase as its rates of reaction were greater than that of the potato.
Without enzymes, reactions wouldn’t occur and living organisms would die. For instance, the enzyme in the stomach breaks down large molecules to smaller molecules to absorb nutrition faster. Researchers experimented with enzyme activity with a potato extract. Researchers will test enzyme activity by increasing and decreasing pH levels, lowering and increasing temperature, and substrate concentration effects. In the first experiment, researchers hypothesized whether different pH levels would change how much Benzoquinone are created and how will the enzymes function in neutral pH levels than higher and lower levels. Researchers used potato extract and different levels of pH to test their hypothesis. In addition, researchers questioned at what temperature does the greatest amount of potato extract enzyme activity take place in. Researchers then hypothesized that the results would indicate the greatest amount of potato enzyme activity level will take place in room temperature. In this experiment, researchers used potato extract and different temperature levels to test the hypothesis. Moreover, researchers wanted to test the color intensity scale and how specific catechol oxidase is for catechol. In this experiment, researchers used dH2O, catechol solution, hydroquinone, and potato extract. Lastly, researchers tested the substrate concentration and how it has an effect on enzyme activity. In this experiment researchers used different measurements of catechol and 1cm of potato extract. Researchers hypothesized that the increase o substrate would level out the enzyme activity