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Photosynthesis involves multiple oxidation-reduction reactions, where electrons are transferred from one complex to another through an electron transport chain. A proton gradient is generated due to the pumping of protons across the lipid bilayer. This gradient is used to drive ATP synthesis. High energy electrons are transferred onto carrier molecules (NAD+/NADH or NADP+/NADPH) which allow these electrons to be used in other chemical reactions (Johnson, 2009). Photosynthesis is actually a three stage process. The first two stages are called the light-dependent reactions; they use absorbed protons of two difference wavelengths to generate NADPH. This is also the step that creates the proton gradient necessary to drive ATP synthesis. In the third stage, the light-independent reactions, the enzyme RUBISCO uses ATP and NADPH to combine CO2 with 1, 5-ribulose biphosphate (Karp, 2008) to form a 6-carbon glucose molecule (Johnson, 2009). Photosystem II is the first stage of the light-dependent reactions. Energy absorbed from light causes the electrons to leave chlorophyll and move to plastiquinone. These electrons can be observed using 2, 6-dichlorophenol indophenol (DCPIP), an electron acceptor that is able to extract the electrons that would have passed to the plastiquinone. DCPIP is a dark blue aromatic compound that becomes colorless when it is reduced. This color change can be measured using a spectrophotometer at 600nm wavelength. The absorbance reading is an indication of the reduction of DCPIP. The lower the absorbance the more reduced the DCPIP, therefore the more photosynthetic activity was present (Johnson, 2009).
The general purpose of this experiment was to determine the effect of light color on photosynthetic activity. If photosynthetic activity of leafy greens is affected by the color of light, then exposing kale (Brassica oleracea var. acephala) to yellow light should reduce the photosynthetic activity as measured by absorbance when compared to that of white light. In a study of lettuce growth it was found that yellow light appeared to suppress the growth of lettuce (Dougher, 2001). The absorption spectrum for photosynthetic activity shows little activity around 580-600nm which is the region of yellow light visibility (Dougher, 2001).
This experiment was designed to determine if yellow light influenced photosynthetic activity in kale. Two kale leaves were used to obtain an enriched chloroplast pellet using grinding buffer and differential centrifugation. Once the pellet was obtained grinding buffer was once again added to form a liquid that could be added to each of the experimental and control tubes. Photosynthetic activity was tested under three conditions. The blank contained 8ml water, no DCPIP, 2ml reaction buffer and 100ÎÂ¼l chloroplasts. The other tubes contained 6ml water, 2ml DCPIP, and 2ml reaction buffer. Tubes tested under yellow light also contained 100ÎÂ¼l chloroplasts as that was the experimental condition where decreased activity was expected. Tubes under yellow light with no chloroplasts acted as the negative control where no activity was expected, and white light with 100ÎÂ¼l chloroplasts acted as the positive control where normal activity was expected. Five replicates were tested in each condition. The tubes to be tested under yellow light were placed in a glass beaker covered with yellow cellophane while the tubes under white light were covered with clear cellophane. Absorbance readings were taken on each replicate at 600nm using a spectrophotometer at time 0, 10, 20 and 30 minutes. The percent change in absorbance was calculated to determine the amount of photosynthetic activity. A P-one tail t-test assuming unequal variance was used to compare the photosynthetic activity of yellow light versus white light, with p<0.05 determined to be significant.
The mean percent change in photosynthetic activity in yellow light was 6.74%. The mean percent change in photosynthetic activity in white light was 8.79%. The negative control in yellow light without chloroplast showed a mean percent change of 0.54%. The mean percent change in absorbance of the chloroplast-containing solutions in yellow light was not significantly different than that of those in the white light (p=0.258193). Figure 1 illustrates that there was little change in absorbance between white light and yellow light.
The hypothesis was not supported by this research. Apparently plant chloroplasts have learned to adapt to different light conditions. The PS II and PS I complexes allow a plant to adapt to various light conditions. PS II complexes are favored in red light, while PS I complexes are favored in yellow light. This allows the plant to able to overcome the prevailing light quality by synthesizing new compounds (Glick, 1986). This idea of photosynthetic redox control of chloroplast transcription was also addressed by Pfannschmidt. Different light absorption properties of the reaction center chlorophylls PS I and PS II work electrochemically in series to ensure that photosynthesis is not interrupted (Pfannschmidt, 2005). This could be an explanation for why there was no significant change between white light and yellow light. The plant is able to overcome the difference in light and continue photosynthetic activity. In our particular research it is possible that the presence of the cellophane itself could have caused the light to be reflected rather than passing through and being absorbed by the chloroplasts. Very little research has been done on the effect of yellow light (580-600nm) and photosynthetic activity because most researchers classify 500-600nm as green light (Dougher, 2001). Research found in testing lettuce growth that chlorophyll or chloroplast formation was suppressed by yellow light (Dougher, 2001). Some of the research in this article would lead one to begin to examine further the affect that yellow and or green light has on photosynthetic activity. If it could be determined that growth is in fact suppressed under these conditions the scientific ramifications of such a discovery could be phenomenal in the way of new pesticides.
Dougher T, Bugbee B. 2001. Evidence for yellow light suppression of lettuce growth. Photochemistry and Photobiology. 73(2): 208-212.
Glick R, McCauley S, Gruissem W, Melis A. 1986. Light quality regulates expression of chloroplast genes and assembly of photosynthetic membrane complexes. Proc. Natl Acad. Sci. USA. 83: 4287-4291.
Johnson A. Spring 2009. Energetics & Photosynthesis. Laboratory Manual Cell Biology. Dept. Biology, Wake Forest University, Winston-Salem, NC. pp. 65-79.
Karp Gerald. 2008. Cell and Molecular Biology. Danvers: Wiley. 230 pp.
Pfannschmidt T, Liere K. 2005. Redox regulation and modification of proteins controlling chloroplast gene expression. Antioxidants & Redox Signaling. 7: 607-618.
Figure 1. Points represent the mean absorbance (600 nm) at 0, 10, 20, and 30 minutes for kale chloroplasts with DCPIP exposed to yellow light (blue diamond) and white light (yellow triangle). The pink squares indicate the mean percent change in absorbance of the negative controls, which contained no chloroplasts and was exposed to yellow light,. There was no significant difference in the absorbance readings between white light and yellow light.