Purpose: The purpose of this lab was to separate plant pigments using chromatography, calculate Rf values using the collected data, and study photosynthesis with isolated chloroplasts. Light energy
Background Information (Activity A): In photosynthesis, plant cells convert light energy into chemical energy that is stored in sugars and other organic compounds. It is an endergonic and anaerobic reaction. Critical to the process is chlorophyll, the primary photosynthetic pigment in chloroplasts. The chemical equation for photosynthesis is: 36 ATP + 6CO2 + 6H2O C6H12O6 + O2 (From: "LabBench for Lab 4." LabBench. PHS School, n.d. Web. 22 Dec. 2012. <http://www.phschool.com/science/biology_place/labbench/lab4/intro.html>.) Pigments are chemical compounds which reflect only certain wavelengths of visible light. This makes them appear "colorful". Flowers, corals, and even animal skin contain pigments which give them their colors. More important than their reflection of light is the ability of pigments to absorb certain wavelengths. Because they interact with light to absorb only certain wavelengths, pigments are useful to plants and other autotrophs --organisms which make their own food using photosynthesis. In plants, algae, and cyanobacteria, pigments are the means by which the energy of sunlight is captured for photosynthesis. However, since each pigment reacts with only a narrow range of the spectrum, there is usually a need to produce several kinds of pigments, each of a different color, to capture more of the sun's energy. Four pigments are usually found in many leaves: carotene, xanthophyll, chlorophyll a and chlorophyll b. Carotene is very soluble in the solvent used in the lab. Its molecules don’t form hydrogen bonds with cellulose, an important polysaccharide in cell walls used for support. Carotene makes a faint yellow to yellow-orange band. Xanthophyll is less soluble than carotene in the solvent. It forms some hydrogen bonds with cellulose. Xanthophyll produces a yellow band. Both chlorophyll a and chlorophyll b easily make hydrogen bonds with cellulose. Chlorophyll a makes a bright green to blue-green band, while chlorophyll b produces a yellow-green to dark olive green band. (From: "Photosynthetic Pigments." Photosynthetic Pigments. N.p., n.d. Web. 22 Dec. 2012. <http://www.ucmp.berkeley.edu/glossary/gloss3/pigments.html>. & Carolina Student Guide for AP Biology Laboratory 4: Plant Pigments and Photosynthesis) Background Information (Activity B): In the light reactions of photosynthesis, light energy is taken in by chlorophyll, the pigment that makes plants green, and is used to excite electrons, the negatively charged subatomic particle. The excited electrons then enter one of two electron transport chains. One chain turns ADP + P to ATP. The other chain changes NADP + H to NADPH. In this part of the lab, we will add a solution of DPIP, which is a blue dye to a suspension of chloroplasts, the plant cell organelle that conducts photosynthesis. The DPIP will replace NADP in the light reactions: DPIP + H DPIPH. DPIPH is colorless, so as the light reactions occur, the blue color of the solution will decrease. We will use this color change as an indication that the light reactions are occurring and we will use the rate at which the color change is happening as a measure of the rate of the light reactions. Independent Variable: The amount of light and the boiling/unboiling/no chloroplasts in the suspension Dependent Variable: % of light transmittance
Hypothesis: If a cuvette contains boiled chloroplasts or has unboiled chloroplast in the dark, then they will have a lower percentage of light transmittance than the cuvette containing unboiled chloroplasts exposed to light.
Materials (Activity A)
* Chromatography jar tightly capped with solvent
* Chromatography paper
* Green Leaf
* Small staples or paper...
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