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2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 1 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea926Exported for Hang Nguyen on Mon, 13 Feb 2023 18:10:14 GMTExperiment 3: E!ect of SubstrateConcentrationHypothesis: The rate of enzymatic activity will increase proportionally with the increase of substrateconcentration because the increased concentration of substrate allows for a greater number of catecholoxidation reactions to occur simultaneously, resulting in a greater rate of product accumulation.Procedure1. Evaluate the hypothesis presented above and formulate the predictions based on the hypothesis in theLab Report below.2. Label four tubes for serial dilution of the substrate, S1:10, S1:100, S1:1000, S1:10000), and fill the tubeswith 9 ml of distilled water (“S” for “substrate”).3. Following the serial dilution procedure outlined in Figure 3.2, transfer 1 ml of the stock solution ofcatechol into tube S1:10 and mix thoroughly by shaking the tube. This is a ten-fold dilution of theMaterialsChecklistCatecholstocksolutionCatecholasestocksolution(keeponice)DistilledwaterSharpiemarkerO10mLgraduatedcylinderTesttubesonarack400mLbeakersDisposablepipettesOWaterbathScissors
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 2 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea926catechol into tube S1:10 and mix thoroughly by shaking the tube. This is a ten-fold dilution of thesubstrate.4. Transfer 1 ml of the ten-fold dilution of catechol from tube S1:10 to tube S1:100 and mix thoroughly byshaking the tube. This is a hundred-fold dilution of the substrate.5. Transfer 1 ml of the hundred-fold dilution of catechol from tube S1:100 to tube S1:1000 and mixthoroughly by shaking the tube. This is a thousand-fold dilution of the enzyme.6. Transfer 1 ml of the thousand-fold dilution of catechol from tube S1:1000 to tube S1:10000 and mixthoroughly by shaking the tube. This is a ten thousand-fold dilution of the enzyme.7. Label test tubes C1, C2, C3, C4, C5, and C6 and place 3 ml of the enzyme (catecholase) in each tube.8. Add 3 ml of catechol stock solution to tube C1 (Table 3.5).9. Add the diluted substrate to the tubes C2C5 from the tubes S1:10, S1:100, S1:1000, and S1:10000,respectively, as shown in Table 3.5.10. Add 3ml of distilled water to tube C6 (see Table 3.5).Table 3.5. Test tube contents for e”ect of substrate concentration experiment.Tube #Chatechol (mL)Chatecholase(mL)Water (mL)Total volume(mL)C13 (from stock)306C23 (from TubeS1:10)306C33 (from TubeS1:100)306C43 (from TubeS1:1000)306C53 (from TubeS1:10000)306C6033611. Record the initial color intensities in the “0 min” column of Table 3.6.12. Place all six tubes in the 40 ¡C water bath for 15 minutes.
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 3 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea92612. Place all six tubes in the 40 ¡C water bath for 15 minutes.13. Agitate the tubes every 5 minutes, and record the resulting relative color intensities in Table 3.6; rankcolor intensity as 0, +, ++, +++, ++++.14. Graph the results.15. Clean up the work space:Discard test tube contents in the sink;Rinse the glassware with tap water and leave it to dry upside down on the rack;Discard used plastic pipettes in regular trash;Collect all the equipment and materials into containers, and return to the cart;Clean the bench top.Lab Report: Experiment 3 E!ect of Substrate Concentration1. What are your predictions regarding the color intensity of the resulting solutions based on thehypothesis?Predictions of the outcome of the reactions in termsof the color intensity of the product.HomeworkAnsweredAt which substrate concentrations the solutions will turn most intense brown (+++++), least intense brown (+), andthe intermediate shades of brown (++/+++/++++)?Drag and drop options on the right-hand side and submit. For keyboard navigation…SHOW MORE+++++++++++++++Stock1:101:1001:10001:10000
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 4 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea926+Answered1:10000Resubmit2. Report the data for the solutions tested in the Table 3.6.The color intensity data at di!erent substrateconcentrations.HomeworkAnsweredReport the data for the tested solutions in terms of brown color intensity (0/+/++/etc.) in the tablebelow.UndoStart OverColumnRowTable TitleColumn 1Column 2Column 3Column 4Column 5Column 6Table 3.6. The effect of varying substrate concentration on catecholase activity.Test TubeSubstrate Concentrati0 min5 min10 min15 minC1++++++++++++++++++++++C2+++++++++++++++++C3+++++++++++++C4++++++++C5+++00C600000Resubmit3. Which solution was the control?Control solutions.HomeworkAnswered
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 5 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea926Which of the solutions were the control? Consult the Scientific Skills chapter for the definition of the control.Select an answer and submit. For keyboard navigation, use the up/down arrow keys to select an answer.aC1bC2cC3dC4eC5fC6AnsweredResubmit4. Identify the data type that the rate of catechol oxidation as represented by the color intensity of thereaction product.Identify data type.HomeworkAnsweredWhat data type do the color intensity measurements represent?Select an answer and submit. For keyboard navigation, use the up/down arrow keys to select an answer.aQualitativebQuantitative discretecQuantitative continuousAnsweredResubmit5. Plot the results as a line graph with the y-axis indicating the relative product quantity (represented by
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 6 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea9265. Plot the results as a line graph with the y-axis indicating the relative product quantity (represented bycolor ranking from 0 to +++++) for all five solutions and the x-axis indicating time in minutes. Each trial ofspecific substrate concentration is represented by a curve. Provide the graph with axis labels, units, and acomprehensive legend. Please note: the intensity values represented by “+” symbols cannot be enteredinto the table’s cells, so convert your symbols into digits first, representing the number of pluses byintegers: “+” = 1; “++” = 2; “+++” = 3, etc.Construct a graph.HomeworkGenerate GraphPlot the color intensity vs. time. Note: due to the software limitations, the graph can display dataonly for up to five data sets. For this experiment, do not include the data for the control in yourgraph. Additional rows and columns can be added by clicking the “+ Column” or “+ Row” buttons on the top right side of the table editor.UndoStart OverColumnRowGraph TitleX Axis LabelY Axis LabelX ValuesEnter X valuesin columnbelowY Values1Y Values2 Y Values3²Y Values4¼Y Values5The effect of Substrate ConcentrationTime(Minutes)Color intensityC1C2C3C4C505432155432110432101532210
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 7 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea926
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 8 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea926oxidation? If not, what can be a potential explanation?Hypothesis evaluation.HomeworkUnansweredDo any of the curves reach a plateau or change the slope? If so, speculate why.UnansweredSubmit8. Summarize your findings to evaluate the hypothesis.Hypothesis evaluation.HomeworkUnansweredGiven your data and a likely disagreement between the predictions and the observations,provide potential explanations for the discrepancy between the expected and theexperimental results, and modify the hypothesis accordingly.
2/13/23, 1:10 PMExperiment 3: Effect of Substrate Concentration | Top HatPage 9 of 9https://app.tophat.com/e/263759/assigned/content/987130::f9c14ee7-5757-5d57-a42b-7a1f962ea926Exported for Hang Nguyen on Mon, 13 Feb 2023 18:10:14 GMTUnansweredSubmit
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