Download LIGHT REACTION AP LAB Due Friday, 11/8 To prepare for this lab

LIGHT REACTION AP LAB
Due Friday, 11/8
To prepare for this lab, please …
 Review principles of the Light Reaction
o Use the link provided on my web page to listen to the AP Lab 4 tutorial provided by Lab Bench.
o Watch the corresponding Bozeman videos.
 Title new page in Lab Notebook, Light Reaction AP Lab
 Sub-title the first page, “Part A – Leaf Pigment Chromatography” and complete the information below.
 Sub-title the second page, “Part B – Light Reaction of Photosynthesis” and complete the information below.
 Update your Table of Contents
Please Note: A formal lab write-up is not required. The abbreviated lab write-up will consist of
 Title
 Data
o Part A
 Hypothesis
 Data Table
o Part B
 Hypotheses
 Data Table
 Data Analysis – Part B
o Graph, including key and absorbance rate for each set-up
o Statistical Analysis
 Conclusion
…………………………………………………………………………………………………………….
Part A – Leaf Pigment Chromatography
The photosystems of the light reaction contain various types of pigment molecules in order to enhance light absorption. The different pigments can be
separated in a chromatography procedure. Plant pigment is deposited on paper, and then the paper is placed in a non-polar solvent. The distance each
pigment type travels is determined by the solubility of the pigment in the solvent and the tendency of the pigment to hydrogen bond with the paper.
There are four different pigments found in spinach leaves:
 Carotene – Forms faint yellow to yellow-orange bands; Highly soluble in chromatography solvent; Non-polar
 Xanthophyll – Produces a yellow band; Slightly less soluble than carotene; Forms some hydrogen bonds with paper
 Chlorophyll a – Produces bright-green to blue-green band; Polar
 Chlorophyll b – Produces yellow-green to dark, olive-green band; Identical in structure to chlorophyll a but methyl group of chlorophyll a is
replaced with an aldehyde group
Purpose: To observe the separation and migration of photosynthetic pigments used to capture light energy.
Pre-Lab – On the first page …
 Formulate a hypothesis predicting the relative distance each pigment will move; in other words, rank the pigments according to how far they
will travel on the paper.
 Justify your hypothesis.
 Construct a data table to record the distance traveled for each pigment and the Rf value. List the pigments in the order you predict them to
move, beginning with the pigment that should travel the shortest distance. Note the expected color of each band in the data table.
Part B – Light Reaction of Photosynthesis
In order to provide the energy needed for photosynthesis, plants contain pigment molecules capable of absorbing photons of light energy from the visible
spectrum. As light energy is absorbed, electrons within the pigment molecules of the photosystems are raised to a higher energy level. The energy from
the “excited electrons” is passed from one pigment molecule to another in the photosystem until it is received by a special complex known as the
reaction center. At this point, the pair of chlorophyll a molecules that make up the reaction center give up a pair of electrons that are ultimately used to
generate ATP (in Photosystem II) and reduce NADP+ (in Photosystem I). In the lab, a dye-reduction technique will be used to visualize this process.
DPIP (2,6-dichlorophenol-indophenol) will be used as an electron acceptor. It is normally a deep blue color, but as DPIP is reduced, it changes from
blue to colorless. This color change will be quantified with a colorimeter by measuring light Absorbance. As the color changes from blue to clear,
absorbance decreases.
Purpose: To determine the effect of different wavelengths of light on the rate of the light reaction of photosynthesis.
Pre-Lab – On the second page …
 Formulate hypotheses predicting relative reduction rates of DPIP based on the color of the cup placed over each cuvette during the light
incubation as compared to the set-up with the clear cup.
o Cuvette 1 → Red
o Cuvette 2 → Blue
o Cuvette 3 → Green
 Justify your hypotheses
o A few things to consider . . .
 The transparent colored cups act as filters, meaning the “color” represents the portion of the visible spectrum that is able
to pass through
 What portion(s) of the visible spectrum do plants normally utilize for the light reaction?
 Construct a Data Table for your results.
o You do not need to record any results for the blank.
o Absorbance readings will be taken at the following time increments:
 0 min
 5 min
 10 min
 15 min
 20 min
Part B Data Analysis
 Construct a graph showing your table results for each of the four cuvettes.
o Calculate the Absorbance Rate for each cuvette → Total Change in Absorbance / Total Amount of Time (AU/min)
o Include this information with the key for your graph.
 To determine if the effect of light color on the Absorbance Rate is significant,
o Record a null hypothesis for each of the three test cuvettes as compared to the control.
o Use class data to complete a T-test to determine if the Class Average Change in Absorbance over Time for each set-up is
significantly different than the Mean Change in Absorbance over Time for the Control. Please show your work.
o For the control and each color:
 Calculate the class mean using class data posted.
 Determine the variance (s2)
 Calculate the deviation from the mean for each value.
 Square each deviation from the mean.
 Determine the variance or s2 by finding the sum of the squared deviations from the mean and dividing the sum
by n – 1 or the (number of values minus 1).
 Use the following equation to set up the t-test for each color set-up as compared to the control
 t-value = (Difference between the two means)
√ [ (s12 + s22) / (n1 + n2) ]
Determine the Degrees of Freedom
 (n1 – 1) + (n2 – 1)
 Use the chart provided/posted to determine if the difference between the means is statistically significant.
 Find the row representing the correct number of Degrees of Freedom
 Look under the column for 0.05
 If the calculated t-value is greater than the value listed, the null hypothesis is rejected
 This means there is less than a 5% chance that the difference between the means is statistically
insignificant; in other words, there is a significant difference between the means.
 If the calculated t-value is less than or equal to the value listed, the null hypothesis is accepted
 This means the difference between the means is most likely due to random chance and the means can
be considered “the same”.
 You will compare the control with each color set-up so you will need to complete these steps a total of three times.
Calculate the standard deviation or spread for each set-up by taking the square root of the variance (s2).

o
Conclusion
Write a well-written paragraph in which you …
 Write a brief overview describing the light reaction of photosynthesis
 Discuss the importance of plant pigments, light energy to the reaction.
 Evaluate your data from Part A. Was your hypothesis correct? Explain.
 Discuss your data, statistical analysis in Part B.
o Were your original hypotheses correct?
o Did the statistical analysis support or reject the null hypothesis for each comparison?
o Evaluate the standard deviation found in the data