PhotosynthesisFranclene Milla, Chanel Mirafuentes, Gianica Monteagudo, Bradley Ong

BIO 21, LAB1A MHAB1

Submitted to: Professor Elena RagragioSubmitted: September 20, 2012

Photosynthesis

ABSTRACTPhotosynthesis is a complex process that autotrophs or food-producing organisms undergo by combining carbon dioxide with water to reduce it to form glucose, a molecule that stores energy[2]. The various factors that are needed for such a process and the different roles that they play have been witnessed in this experiment. Three parts of the procedure were concentrated on chlorophyll as a driver, light as an energy source and carbon dioxide as raw material. Several pigments present in the chloroplasts of leaves were identified as absorbers and channels of energy. It was also affirmed that oxygen was a by-product of photosynthesis.

1. IntroductionAs with any living organism, plants need energy to perform the different processes that keep it healthy and functional. One source is glucose, a molecule containing considerable amounts of energy. Plants can obtain glucose easily because they are capable of manufacturing in a process called photosynthesis. Photosynthesis is a very complicated procedure with numerous sub-processes yet it cannot occur without several factors, namely chlorophyll, light, water, and carbon and dioxide. These factors will serve as the foci of this activity.

This experiment aims to:(1) Determine the role of chlorophyll, light and carbon dioxide in photosynthesis.(2) Observe oxygen liberation during photosynthesis.(3) Demonstrate how chloroplast pigments are separated and to identify these pigments.

If a plant is sufficiently exposed to light, chlorophyll is present in the leaves and there is ample supply of carbon dioxide, the plant is able to undergo photosynthesis.

2. Methodology2.1. The Role of Chlorophyll in Photosynthesis We boiled the leaf for 10 minutes. The boiled leaf was immersed in a test tube in with 95% ethyl alcohol. We placed the test tube in a hot water bath until the pigments were extracted. The bleached leaf was washed with water and tested for presence of starch using IKI solution.

2.2. The Role of Light inPhotosynthesisWe place a potted plant in the dark for 48 hours. After this, we selected several leaves and wrapped portions of them with black or carbon paper fixed by paper clips. Then the whole plant was exposed to light for about 5 hours. The leaf was plucked and tested for starch using IKI solution. 2.3.The Role of CO2 in PhotosynthesisWe labeled three test tubes A, B and C and filled these with previously boiled then cooled distilled or tap water. We put Hydrilla sprigs into test tubes A and B in an inverted position. A pinch of NAHCO3 was added in test tubes B and C. We exposed the 3 test tubes to bright light. We determined if bubble evolution occured in test tubes And B. The Hydrilla sprigs in test tubes A and B were removed. Five drops of phenol red were added to each test tube (A, B & C) and shake gently. (Phenol red changes toyellow at low pH). We compared the color intensities of the solutions in the test tubes.

2.4. Oxygen Liberation in PhotosynthesisWe filled a one-liter beaker with tap water. To enrich CO2 content of water, we blew air gently into the beaker for 3-5 minutes using a plastic straw. We cut the ends of Hydrilla under water and insertde a thistle funnel over the Hydrilla sprigs. The cut ends of the plant should face the tube of the funnel. We placed a test tube filled with water upside down over the tube of the funnel partially dipped in thewater of the beaker. We observed for bubbles emerging out from the cut ends.

2.5. Separation of Chloroplast PigmentsTo prepare the leaf extract, we got 15-20 mature Hibiscus leaves and cut into small pieces. We put them in a mortar with a pinch or two of sand. The extract was expressed. We put in a vial and set aside.

To prepare the filter paper strip, we fit a piece of paper into a clean, dry test tube. With this paper as a pattern or mold, we cut the filter paper strip. We held the filter paper by the edges. Refrain from touching the filter paper. (Fingerprints on the surface of the paper may affect the chromatogram.)

The filter paper should easily slide in and out of the tube without touching the sides. With a pencil, not a pen, we marked about 2 cm from the top and 1 cm from the base of the strip. We cut a point on the base of the strip. We hung the strip on a cork by means of a hook made from a paper clip. We marked the outside of the test tube to show proper solvent level. We removed the paper strip from the dry test tube and lay it on the table. In the meantime, we poured the solvent into the dry tube up to the marked line. We replaced the cork (without the paper strip) and kept inside.

To prepare the chromatogram, we dipped a capillary tube or a fine-tip dropper into the vial containing the extract. We placed the drop that comes out of the capillary tube onto the center of the 1 cm mark at the base of the strip. Let it dry completely by waving the paper in the air. We added another drop directly on the first. The green spot should be as small as possible. We dried and repeated the process about 10-15 times until the spot became saturated with the green extract. We took note that the spot must be completely dry before each drop was added. When we were finished, the paper had a small, dark green spot in the center of the 1 cm mark. The spot did not reach the sides of the strip.

When the spot wass completely dry, we removed the cork from the tube and placed the paper strip on the hook and inserted into the test tube containing the solvent mixture 95 parts petroleum ether and 5 parts acetone. The solvent level was below the 1 cm mark. We replaced the cork with the strip and observed the flow of the solvent until the 2 cm mark. Immediately, we removed the paper from the tube and let it dry.

When the strip was completely dry, the chromatogram showed the different pigments present in the specimen.

3. Results3.1. The Role of Chlorophyll in Photosynthesis

The green pigment extracted was chlorophyll. Starch was detected in the green areas while there were only minimal amounts in the white areas.

3.2. The Role of Light in PhotosynthesisThe leaf that had been covered was almost or completely devoid of starch. The test showed that the leaf only contained minimal amounts of starch.

3.3. The Role of Carbon Dioxide in PhotosynthesisBubble evolution occurred in test tubes A and B. However, the rate at which said bubbles were released was slow and had long intervals. The table below shows the color of the solutions after phenol red has been added. Table 1. Color Intensities of Solutions in A, B, and CTest Tube LabelColor

Test Tube ALight orange

Test Tube BMagenta

Test Tube CMagenta

3.4. Oxygen Liberation in PhotosynthesisBubble evolution occurred at an extremely slow rate with an average of 1-2 bubbles released per minute. 3.5. Separation of Chloroplast PigmentsTable 2. Pigments Present in HibiscusColorPigment

Green-Blue GreenChlorophyll A

Yellow Green-Light GreenChlorophyll B

OrangeCarotene

YellowXantophyll

4. Discussion4.1. The Role of Chlorophyll in PhotosynthesisThe leaf was boiled prior to extraction to cause the chloroplasts to burst thus exposing the pigments. The green areas mentioned earlier were the photosynthetic areas because they contain chlorophyll, an important component of photosynthesis. Starch test was employed because the presence of starch indicates the occurrence of photosynthesis. Starch was evident in the green areas because they contain abundant amounts of chlorophyll and therefore are able to undergo photosynthesis. Meanwhile, the white areas have only limited reserves of chlorophyll. In conclusion, only areas with chlorophyll can undergo photosynthesis.

4.2. The Role of Light in PhotosynthesisThe plant was placed in the dark to consume its starch reserves. Some leaves were also covered to prevent light penetration. The uncovered portions of the leaves responded positive to the starch test because these are the only parts that were able to absorb enough light to trigger photosynthesis. The covered portions, having used up their starch reserves for energy consumption and having been unable to capture light energy were ultimately unable to begin food production. Therefore, light is needed in photosynthesis.

4.3. The Role of Carbon Dioxide in PhotosynthesisThe bubble formation that occurred in test tubes A and B was actually the release of oxygen gas as the Hydrilla sprigs underwent photosynthesis. The addition of phenol red was to identify the pH of the solutions while photosynthesis was occurring. Solution A (from test tube A) turned light orange which signifies a low pH or it was acidic. Carbon dioxide which forms carbonic acid with water wasnt being consumed at the same rate as in the other set-up. On the other hand, solution B was magenta which means that it was considerably more basic. The addition of sodium bicarbonate to the solution increased the rate of photosynthesis hence carbon dioxide was used up more rapidly by the sprig in set-up B than in set-up A. This led to the solution being slightly more basic. Solution C served as the control set-up. The less carbon dioxide present, the more basic a solution is.

4.4. Oxygen Liberation in PhotosynthesisInverting the test tube over the funnel enables the Hydrilla plant to obtain carbon dioxide from its environment, enabling it to undergo photosynthesis. The bubbles were oxygen gas being released as by-product.

4.E. Separation of Chloroplast PigmentsThe pigments were identified to be chlorophyll a and b, carotene and xanthophyll[3]. Chlorophyll a and b are involved in photosystems I and II. Carotenoids absorb different wavelengths and transfer energy to chlorophyll a. Chlorophyll is the most soluble, followed by xanthophyll with carotene being the least soluble[4].

REFERENCES1. Committee on Biology 21 Laboratory Manual FS AY 2008-2009. (2008). Biology 21: General Botany Laboratory Manual.1. Mauseth, J. (2009). Botany: An Introduction to Plant Biology, 4th edition. Sudbury, MA: Jones and Bartlett Publishers.

1. Chloroplast Pigments. (n.d.) Photosynthesisinfo. Retrieved September 19, 2012 from http://www.photosynthesisinfo.com/chloroplast-pigments/1. B. Lowe. (1937). Experimental Cookery from the Chemical and Physical Standpoint. IO: John Wiley and Sons