Purpose: to divide the assorted photosynthetic pigments of Spinacia oleracea leaves. Materials:
– methyl alcohol
– H2O
– filter paper
– sand
– trial tubing
– 10 S. oleracea foliages
– 2 wooden splints
– howitzer
– stamp
Method: First. 10 S. oleracea foliages were gathered. A piece of filter paper. a wooden splint. a howitzer and stamp were besides gathered. The foliages were placed inside the howitzer. make fulling the howitzer about half full. The foliages were so crushed with the stamp. Sand. H2O and methyl alcohol were added and so the foliages were crushed once more. This was repeated until the foliages had reached the consistence of a soupy green paste. The leaf infusion ( soupy green paste ) was strained. taking the hempen Spinacia oleracea paste. which was placed into the refuse. and go forthing the liquid foliage infusion. The wooden splint was dipped into the leaf infusion and so traced in a thin consecutive line onto the filter paper. The filter paper was allowed to dry for about 2 proceedingss. The following coat could non be applied until the paper was wholly dry. The procedure was repeated 10 times. from dip. hint and drying. A trial tubing was gathered every bit good as methyl alcohol and a 2nd wooden splint. The wooden splint was pierced through the top of the filter paper and so the filter paper was placed into the empty trial tubing. It was measured how much methyl alcohol would be needed so that the tip of the filter
paper was submerged but the extract line was non submerged. After measuring. the filter paper was removed from the trial tubing. The methyl alcohol was poured into the trial tubing up to the measured fill point. The filter paper was so carefully inserted so that it ne’er touched the borders of the trial tubing. The trial tubing was carefully placed vertically into a trial tubing holder and allowed to absorb methyl alcohol for 45 proceedingss. After the 45 proceedingss were complete. the filter paper was so removed. measured as to how far pigments had travelled and left to dry.
Observations:
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Calculations: Rf xanthophyll= distance pigment travelled
distance to solvent forepart
= 7
9. 7
= 0. 72
Rf chlorophyll a= distance pigment travelled
distance to solvent forepart
= 8. 4
9. 7
= 0. 87
Rf chlorophyll b= distance pigment travelled
distance to solvent forepart
= 8. 5
9. 7
= 0. 88
Rf carotene= distance pigment travelled
distance to solvent forepart
= 8. 7
9. 7
= 0. 90
Discussion: 1. The Mortar and Pestle is used to interrupt down the intramolecular bonds within the S. oleracea leaves. This helped interrupt the molecule down into smaller molecules and single atoms so that chloroplasts and their pigments were non buried inside the foliage and were easy accessible.
2. Methanol is used to fade out the pigments. as methyl alcohol is polar and so are the pigments. As we saw. the pigments stayed with the methyl alcohol in the liquid portion of the mixture but everything else ( cell walls. etc ) floated to the top of the mixture. so we could easy take it.
3. It is of import that the filter paper did non touch the walls of the trial tubing because if touching the side walls. H2O would absorb from the sides every bit good as the underside of the filter paper. If this were to go on. the flow of methyl alcohol up the filter paper would be disturbed. therefore upseting the flow and separation of the pigments.
4. It is really of import that the foliage infusion non be dissolved in the dissolver because it would non go upward the same. If the extract line were to be dissolved in the methyl alcohol. it would merely fade out into the methyl alcohol. without really holding the opportunity to go up the filter paper. With the extract line above the semilunar cartilage. the pigments’ merely pick was to follow the way of the methyl alcohol.
5. Water would non be able to transport the pigments up the filter paper because it does non hold the same physical belongingss as methyl alcohol. One of methanol’s alone characteristics is that it dissolves compounds of really high mutual opposition. While H2O will fade out polar molecules. it would be unable to fade out these pigments every bit good as methyl alcohol can.
6. Harmonizing to my findings. chloroplast a is in the highest copiousness in the foliage infusion. This is known because of the really bright green coloring material of the pigment. every bit good as the sweep of chloroplast a on the filter paper.
7. Pigments separate instead than go up the paper at the same rate due to their solubility. Pigments that are the most soluble in methyl alcohol will go to the highest degrees. about every bit high as methyl alcohol itself. but pigments that are least soluble in methyl alcohol will remain lower down on the filter paper. The pigments will merely go far plenty so that they are wholly dissolved from the methyl alcohol to the filter paper. Therefore. those pigments that are least soluble in methyl alcohol. take less clip and infinite to fade out from methyl alcohols so they stop going at an earlier phase that those that are most soluble with methyl alcohol.
8. The significance of the Rf value is to assist place pigments. Each pigment holds an Rf value. which is used like a fingerprint. Although coloring material can be used to place pigments. their Rf value is the true indicant of which pigment is present. Decision: the assorted photosynthetic pigments in S. oleracea were separated utilizing the chromatography technique.
