Bio Lab Report Final - Grade: A+ PDF

Preview

Summary

bio lab 1 final lab report...

Description

Mance 1 Lauren Mance Dustin Partridge Intro to Bio Lab 9 December 2016 Photosynthesis: Affects of Wavelength, Separation and Identification of Pigments, and Determining the Absorption Spectrum for Leaf Pigments Abstract: This study investigates the photosynthetic activity of different wavelengths of light. The purpose of this experiment is to determine if products of photosynthesis are present in a leaf tissue that has been exposed to different wavelengths of light for several days. Various pieces of different colored filter papers (black, green, blue, and red) are cut and placed onto small portions of a leaf from a germanium plant and after a few days the paper is taken off and the leaves are dropped into the boiling alcohol solution in order to extract the pigments. When the leaves are mostly devoid of color, they are removed from the boiling alcohol bath and drops of I2KI are then added in order to show which areas of the leaf test positive for starch. The portion of the leaf that allows most photosynthetic activity is the portion of the leaf that does not have filter paper on it because all wavelengths of light are able to pass through the leaf. The portion of the leaf with red and blue filter paper also have high levels of photosynthetic activity since only blue and red light are transmitted and the rest are absorbed, still allowing photosynthesis to happen at a high rate. Starch is not present in the area of the leaf where green and black filter papers are placed. The reason as to why photosynthesis does not take place under the green filter papered area is because even though green light is able to go through the filter and get reflected, that color doesn't make photosynthesis. Under the area where the black filter paper is placed, there is no photosynthesis, because black absorbs all wavelengths of light and no light goes to the plant. The continuation of this experiment includes plotting the absorption spectrum of leaf pigments separated by paper chromatography. The purpose of this is to determine the absorption spectrum for leaf pigments. The pigments separated by paper chromatography are cut out and distributed into six groups. Groups one through four dilute the pigments and groups five and six determine the absorption spectrum of the total pigment solution. In terms of optimal wavelength, 720 is the optimum for chlorophyll a and b, 680 is the optimum wavelength for xanthophyll, 640 or 700 is the optimum wavelength for carotene, 700 is the optimal wavelength for total pigment 1, and 720 is the optimal wavelength for total pigment 2. Chlorophyll a is the most important pigment for photosynthesis. Chlorophyll b is an accessory pigment and simply aid in harvesting the energy from different wavelengths of light. Carotenoids absorb wavelengths of light not picked up by chlorophyll a and help aid absorption of light and efficiency of photosynthesis. Introduction: Photosynthesis is the process by which plants convert solar energy from the sun into useful chemical energy for food. They release molecular oxygen and remove CO2 from the air. Plants are photoautotrophs, which means they are able to synthesize food directly from inorganic compounds using light energy, instead of eating other organisms or relying on material derived from them. The energy for photosynthesis comes from absorbed photons found in light and involves a reducing agent, in this case water. Oxygen is released as a product. All the organisms that undergo

Mance 2 photosynthesis convert CO2 to organic material by reducing this gas to carbohydrates. This is done through a rather complex set of reactions. The pigments that absorb the light are primarily chlorophylls and carotenoids. Chlorophylls absorb blue and red light while carotenoids absorb blue-green light, but green and yellow light are not effectively absorbed by photosynthetic pigments in plants; therefore, light of these colors is either reflected by leaves or passes through the leaves. This explains why plants are green. Chlorophyll is also unique in that it is capable of converting the active energy of light into a latent form that can be stored and used when needed (Campbell). This study investigates the photosynthetic activity of different wavelengths of light. The purpose of this experiment is to determine if products of photosynthesis are present in a leaf tissue that has been exposed to different wavelengths of light for several days, and then plot the absorption spectrum of leaf pigments separated by paper chromatography. Leaves of a geranium plant that have had small pieces of black, green, red, and blue filter paper placed on small portions of the leaf are analyzed and levels of photosynthetic activity are determined by testing for the presence for starch under the filter papers. Starch can be easily tested as an indirect product of photosynthesis and will turn dark in the presence of iodine. Filter papers used in this experiment are designed to reflect and transmit the appropriate wavelengths of light to correspond to the visible light spectrum. Leaf portions covered in black or green filter paper will have no reaction with starch due to a lack of photosynthesis. Leaf portions covered in red or blue filter paper will produce carbohydrates and will react with starch since photosynthesis will have still taken place. As for pigment wavelengths, the absorption spectrum is the absorption pattern for a particular pigment, showing relative absorbance at different wavelengths of light. The absorption spectrum can be determined by utilizing a spectrophotometer/calorimeter. In terms of optimal wavelength, chlorophyll and total pigment will share the same optimal wavelength, since chlorophyll a is the most important pigment in photosynthesis. Chlorophyll b will have a lower but closely related wavelength, followed by xanthophyll and carotene since they are all accessory pigments in photosynthesis. Xanthophyll absorbs light in the green-blue-indigo-violet region and reflects mostly yellow light, chlorophyll a absorbs light in the blue-violet region and reflects green light, chlorophyll b absorbs red-blue light and reflects green light, and carotene and carotenoids absorb light in the blue-green-violet region and reflect the yellow, red, and orange light (Govardovskii). Methods: Four to five days prior to the experiment, various pieces of different colored filter papers (black, green, blue, and red) are cut and placed onto small portions of a leaf from a germanium, then are placed under bright light. On the day of the experiment, before boiling the germanium leaf, I distinguish which leaves have which colored filter papers on them by introducing distinguishing features; on the leaf with the black and red filter paper, a small hole is poked through the right side of the leaf and the stem is cut off, and on the leaf with the green and blue filter paper, the leaf remains unchanged. A boiling alcohol bath containing 300mL of water and 200mL of 80% ethyl alcohol is placed on a hot plate in a 1,000mL beaker and the beaker is brought to a boil. The filter papers are to be removed from each leaf and, using forceps, the leaves are dropped into the boiling alcohol solution in order to extract the pigments (Harborne). When the leaves are mostly devoid of color, they are removed from the boiling alcohol bath using forceps, placed in separate petri dishes, and are rinsed with distilled water in order to cover the leaf. Drops of I2KI are then added to the water in the petri dishes until a pale amber color is obtained. Results are to be recorded, showing which areas of the leaf test positive for starch. For chromatography,

Mance 3 using a capillary tube, the leaf pigment extract is streaked on a pencil line previously drawn 1cm from the edge of the paper cylinder. The paper is put in cylindrical form so that the paper has a small gap between the left and right sides of the chromatogram, which prevents solvent or compound from crossing over (Irreverre, Filadelfo, and Martin). After initially allowing the chlorophyll to dry, this step is to be repeated three to four times, allowing the extract to air-dry each time. A band of green pigments become present along the pencil line, and their intensity determines how good the results of the experiment will be (the darker the bands, the better the results). A jar containing petroleum ether and acetone solvent is obtained and, using forceps, the loaded paper cylinder is loaded into the solvent and quickly covered tightly with the lid. The chromatography proceeded until the solvent reaches within 3cm of the top of the cylinder. The cylinder is removed from the jar, dried, and the staples are removed. The paper with separated pigments are cut out and distributed as follows: team 1 is carotene, team 2 is xanthophyll, team 3 is chlorophyll a, team 4 is chlorophyll b, and teams 5 and 6 determined the absorption spectrum of the total pigment solution. I am a part of team 6. I add drops of the original chlorophyll extract solution (acetone pigment mixture) to 10mL of acetone until it looked pale green. This becomes pigment solution for cuvette B. The acetone with no pigment becomes the solution for cuvette A. The purpose of the cuvette with reference material is because they are always required for setting the spectrophotometer to read zero absorbance at each wavelength used. Both cuvettes are filled two-thirds full and are wiped with Kimwipes to remove fingerprints. The absorption spectrum is measured and results are recorded. Absorption spectrum is measured by using a spectrophotometer; the beginning wavelength is selected (400nm) and the instrument is zeroed by adjusting the 0 control knob so that the meter reads 0% transmittance. The instrument is then calibrated by inserting cuvette A into the sample holder and closing the lid. The light control is adjusted until it read 100% transmittance and the mode is changed to absorbance. The readings can then begin by removing cuvette A and inserting cuvette B. The spectrophotometer is recalibrated every time the wavelength is changed and observations continue until 720nm is reached. Results: The area where filter paper was not placed was the area of highest photosynthesis activity. On the portion of the leaf where red paper was placed, red light was transmitted and blue was absorbed, so photosynthesis still took place at a relatively high rate. On the portion of the leaf where blue paper was placed, blue light was transmitted and red was absorbed, so photosynthesis still took place at a relatively high rate. On the portion of the leaf where a green filter was placed, green light was transmitted and red and blue were absorbed, preventing any photosynthetic activity. On the portion of the leaf where black filter paper was placed, no light was able to get through to the leaf, thus preventing any photosynthetic activity. Refer to Figure 1.

Before:

After:

Black=no starch Red=starch

Green=no starch Blue= starch

Mance 4 Using the readings recorded, the absorption spectrum for each pigment is able to be determined and plotted as shown in Figure 2:

Absorption Spectrum of Leaf Pigments 1.2 1 Chlorophyll a Chlorophyll b Xanthophyll Carotene Total Pigment 1 Total Pigment 2

Absorption

0.8 0.6 0.4 0.2 0 400 420 440 460 480 500 520 540 560 580 600 620 640 660 680 700 720 Wavelength (nm)

Discussion: The portion of the leaf that allows most photosynthetic activity is the portion of the leaf that does not have filter paper on it because all wavelengths of light are able to pass through the leaf. The portion of the leaf with red and blue filter paper also have high levels of photosynthetic activity since only blue and red light are transmitted and the rest are absorbed, still allowing photosynthesis to happen at a high rate. Starch is not present in the area of the leaf where green and black filter papers are placed. The reason as to why photosynthesis did not take place under the green filter papered area is because even though green light is able to go through the filter and get reflected, that color does not make photosynthesis. Rather, the red and blue wavelengths aid photosynthetic activity, and since those colors are absorbed in the green filter paper, photosynthesis does not occur.Under the area where the black filter paper is placed, there is no photosynthesis, because black absorbs all wavelengths of light and no light reaches the plant. The results therefore support the hypothesis and predictions. Potential error sources include improperly labeling the leaf with regard to where what color filter paper was placed, thus leading to skewed results, light and temperature exposure, and overall environment for the plant/leaves. In terms of optimal wavelength, 720 is the optimum for chlorophyll a and b, 680 is the optimum wavelength for xanthophyll, 640 or 700 is the optimum wavelength for carotene, 700 is the optimal wavelength for total pigment 1, and 720 is the optimal wavelength for total pigment 2. These results also support the hypothesis and predictions. Chlorophyll a is the most important pigment because it is common to all types of plants. Chlorophyll a is known as the primary pigment in plants. It traps light and is also the source of electrons used in the process of photosynthesis. The other pigments such as chlorophyll b are accessory pigments and simply aid in harvesting the

Mance 5 energy from different wavelengths of light (Thornber). In terms of chlorophyll b and carotenoids, they absorb those wavelengths of light not picked up by chlorophyll a and so increase the absorption of light and efficiency of photosynthesis. They are part of the antenna complex of pigments, which pass the energy to chlorophyll a in the reaction centers of the light stage.

Works Cited

Mance 6

Campbell, Beth. "Photosynthesis." http://wmich.edu/engineer. Western Michigan University, Apr. 2006. Web. Govardovskii, Victor I., et al. "In search of the visual pigment template." Visual neuroscience 17.04 (2000): 509-528. Harborne, A. J. Phytochemical methods a guide to modern techniques of plant analysis. Springer Science & Business Media, 1998. Irreverre, Filadelfo, and William Martin. "Versatile technique of paper chromatography." Analytical Chemistry 26.2 (1954): 257-260. Thornber, J. Ph. "Chlorophyll-proteins: light-harvesting and reaction center components of plants." Annual Review of Plant Physiology 26.1 (1975): 127-158....