Copy of Lab 5 Photosynthesis Post-lab_Fall22

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Feb 20, 2024

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Post-lab Assignment LAB 5: PHOTOSYNTHESIS Bio 1AL Fall 2022 Last Name _____Kim_____ First Name ________Juliana_______ Lab Section #______223______ Lab 5 Post-Lab 1) (0.5 pt) Upload an image of your chromatogram with a ruler next to it, setting the 0 cm mark at the starting line. The position of the pigment bands, starting line, and solvent front should be marked. 2) (1 pt) Use your chromatogram to complete the table below. Identify which pigment is chlorophyll a vs. chlorophyll b. Table 1. Pigment chromatography Rf values. Band # & Color (from top (solvent front) to bottom (starting line)) Pigment name* Migration distance from starting line (cm) Rf value Solvent front n/a 10.6 n/a 5 (yellow) carotene 10.6 1 4 (yellow) carotenoid 1 8.5 0.802 3 (yellow) carotenoid 2 5.4 0.509 2 (green) chlorophyll a 4.9 0.462 1 (green) chlorophyll b 2.3 0.217 *For the pigment name, you should differentiate between chlorophyll a and b, but you do not need to differentiate between the two carotenoids (just list them as carotenoid 1 and carotenoid 2) 5-1
Post-lab Assignment LAB 5: PHOTOSYNTHESIS Bio 1AL Fall 2022 3) (0.5 pt) A key consideration in this lab is to make a standardized suspension of chloroplasts (Page 5D , Part IV). In the table below, follow the directions in Part IV to find C X and V X . Calculate the values based upon the absorbance that you measured in lab for tube “L”. Table 2. Standardization of chloroplast suspension Absorbance of Tube “L” (diluted chloroplast suspension) C X = concentration of chlorophyll in enriched chloroplast suspension (Tube “E”) (mg Chl/mL) V X = volume (mL) of Tube “E” suspension to add to 5 mL buffer for 0.1 mg Chl/mL 0.45 .0240 0.863 4) (1 pt) Using your own group’s data (not the data for all stations), create a graph of absorbance at 600 nm versus light exposure time for the four experimental conditions . See Excel Graphing Resources page in bCourses. Save the graph in Excel as an image (Ctrl Click on the graph > Save as Picture or take a screenshot) and paste the image below. 5) (0.5 pts) Based on your graph above, indicate what time interval (starting and ending time points) would be appropriate to use to calculate the rate of DCPIP reduction (absorbance change / min) for each treatment. Only the decreasing, steepest linear part of each curve should be used for the rate calculations. Table 3. Time intervals for calculating rate of DCPIP reduction. Time interval for calculating rate of DCPIP reduction (min) Dark 0 - 3 min Light + DCMU 0 - 3 min Light 0 - 1.5 min Light + methylamine 0 - 0.75 min 5-2
Post-lab Assignment LAB 5: PHOTOSYNTHESIS Bio 1AL Fall 2022 (2 pt) Download the Photosynthesis data template Excel file from the Lab 5 homepage and open in Excel (do not open in your web browser). Copy all data from your lab section’s class data sheet on page 5G into the Excel file. Create a graph of average absorbance at 600 nm ± SEM versus light exposure time for the four experimental conditions (see graphing tables on the second tab of the Excel file). See Excel Graphing Resources page in bCourses. Save the graph in Excel as an image (Ctrl Click on the graph > Save as Picture or take a screenshot) and paste the image below. 6) (1 pt) Use the class data to calculate the average corrected rate ( ± SEM) of DCPIP reduction for the four conditions. Notes: Only the decreasing linear part of the curve was used for the rate calculations in Excel (timepoints used in the Excel file are noted below). The Excel file also has a function to correct the light reaction rates by subtracting off any reduction rate of DCPIP that occurred in the dark control, so report the corrected values as indicated. Table 4. Effect of DCMU and methylamine on rate of DCPIP reduction Timepoints used (min) Mean Corrected Rate of DCPIP Reduction ± SEM (absorbance / min) Dark (no correction needed) 0 - 3 min 0.675 ± 0.476 Light + DCMU (corrected) 0 - 3 min 1.153 ± 0.027 Light (corrected) 0.25 - 0.75 min 0.857 ± 0.086 Light + methylamine (corrected) 0.25 - 0.75 min 0.597 ± 0.11 7) (1 pt) Compare the DCPIP reduction rates from the light reaction positive control and the methylamine experimental tube. Report the mean corrected DCPIP reduction rates ± SEM for each condition (same as reported in Table 4 above). Calculate the percentage change between the two means, and conduct an 5-3
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Post-lab Assignment LAB 5: PHOTOSYNTHESIS Bio 1AL Fall 2022 Independent Samples one-tailed t-test on these data in VassarStats . Record the relevant values below. (See the Comparing Two Means page in bCourses.) Table 5. Effect of methylamine on rates of electron transport Light Light + methylamine Mean rate ± SEM (absorbance / min) 0.857 ± 0.086 0.597 ± 0.11 Percentage change (%) (0.5 pt) 18.99% t-test results (0.5 pt) t = 1.09 , df = 24 , p (one-tailed) = 0.143 9) (a) (0.5 pt) Write one sentence explaining to a fellow student what your results in Question 8 mean. Our results from the t-test show us that there is similarity between the two treatment absorbance results, and since our p-value is above 0.05, we cannot conclude that a significant difference exists between the two data sets from light and light+methylamine. 9b) 0.5 pt) Write a sentence that explains, in your own words, how methylamine works to affect the electron transport rate. Methylamine buffers and binds H+ because it’s a weak base, and that will speed up photosynthesis which in turn indicates that the speed of electron transport has been increased. 10) Imagine you repeat the DCPIP experiment, but add both DCMU and methylamine in the tube along with the chloroplast suspension. You put the reaction in the light and measure the absorbance every 15 seconds. a) (0.5 pt) How would the rate of DCPIP reduction in this treatment compare to DCMU alone and methylamine alone? (Circle/highlight one) i) Similar to DCMU alone ii) In between rates for DCMU alone and methylamine alone iii) Similar to methylamine alone iv) Faster than methylamine alone b) (0.5 pt) Briefly explain (1-2 sentences). Because methylamine causes the rate of DCPIP reduction to decrease while DCMU causes the rate to stay the same, the mixture of the two should cause a rate that is above methylamine but below DCMU. 11) (0.5 pt) You probably noticed that methylamine increased the rate of electron transfer in your isolated chloroplasts. You wonder if this would work to increase rates of photosynthesis in living plants. Would the addition of methylamine applied to leaves on a living plant increase plant growth? Assume it is absorbed and enters mesophyll cells and enters chloroplasts. 5-4
Post-lab Assignment LAB 5: PHOTOSYNTHESIS Bio 1AL Fall 2022 Briefly explain (1-2 sentences). Circle Yes / No. Explanation: It will increase the rate of plant growth / photosynthesis because electron transfer provides the concentration gradient that is used by ATP synthase to create ATP for photosynthesis; Therefore, with increased rates of electron transfer, comes increase rates of photosynthesis or plant growth. 12) (1 pt) Complete the following table using the absorption spectrum and pigment fluorescence data you recorded on your data collection sheet. Table 6. Absorption spectrum and pigment fluorescence data Absorption Spectrum λ max (nm) (0.5 pt) Peak 1: 428.7 nm Peak 2: 663.5 nm Pigment Fluorescence (0.5 pt) Maximal emission wavelength (nm): 680.9 nm Color of emitted light: red 5-5

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