Question

please do two pargraph about this pre lab and lab include Title,purpose ,chemical eqution if their any ،Procedure ,Hazards(hazard Identification,physical and chemical properties),Experimental Phase, Data &Observations,Data&Observations cont (melting point and meling point). and Calcuations

Spinach Laboratory Procedure PROCEDURES Isolation of pigment from leaves: 1. Wash the spinach leaves. Weigh 1.0 g-2.0 g of dry spinach leaves. Remove any stems and veins from the leaves before you weigh. 2. Combine 1.0 g of anhydrous magnesium sulfate with 0.5 g of sand with the 2.0 g of spinach. Using mortar and pestle, grind the mixture of spinach leaves to a fine paste (green power, 5-10 minutes) 3. Transfer the paste using a Pasteur pipette to a centrifuge tube without cap. Wash the mortar with 2 mL of acetone. Transfer the acetone to the centrifuge tube. Wash again with 2mL of acetone. Mix the acetone with the mixture well. 4. Centrifuge the test for 5 minutes a medium speed. 5. Transfer the liquid portion (after centrifuging) using a Pasteur pipette to another centrifuge tube with stopper. 6. Add 3 mL of acetone to the solid paste.. Mix well and centrifuge again for 5 minutes. 7. Using air decrease the volume of acetone to around 1 mL TLC Separation: beaker walch glass cover JLC plase Gher paper solvent Figure 1. Development Chamber 1. Prepare a TLC development chamber with 70% hexane and 30%acetone. Make sure that the solvent is around 5mm in height. Add half of a filter paper "wick" to the development chamber. The purpose of the wick is to ensure that the chamber is saturated in the vapors of the mobile phase. 2. Prepare a TLC plate. Draw a line at 1 cm from the edge using a graphite pencil. ... 3. Using a TLC capillary, Seed the extract in the TLC line. 4. Place the TLC plate in the development chamber (opposite to the wick) and making sure than the samples in the plates is not in contact with the solvent. 5. Remove the plate when the solvent front is 1-2 cm from the top of the plate. 6. Using a lead or graphite pencil mark the position of the solvent front. 7. Leave the solvent evaporate, and the plate dried out. Outline the spots with a pencil and indicate the colors. Analysis of the results: In the crude extract, you should be able to see the following components (in order of decreasing R fvalues): Carotenes (1 spot) (yellow-orange) -5 /S .1 Pheophytin a (gray, may be nearly as intense as chlorophyll b) ls Pheophytin b (gray, may not be visible) 3.7 1s Chlorophyll a (blue-green, more intense than chlórophyll b) Chlorophyll b (green) 1 /s Xanthophylls (possibly three spots: yellow) re 2 /s = o.3 Pe Depending was spotted on the TLC plate, you may observe other pigments. These additional components can result from air oxidation, hydrolysis, or other chemical reactions involving the pigments. It is also common to observe components in the green band that were not present in the extract. on the spinach sample, the conditions of the experiment, and how much sample The distance traveled by each component is measured and this value is called the retardation factor or retention factor, designated as the R/value. The Rfvalue for a component is calculated using the following expression. distance traveled by the component R distance traveled by the solvent There is a Rfvalue associated with each developed spot on the TLC plate. ieasL To L B 3 cm Rfor Spot A: -0.3 distance between baseline and solvent frout 10 cm 10 cm 7 cm Refor Spot B -0.7 10 cm Po s Figure 2. Calculation of Rfvalues for spots on a developed TLC plate Tcm

Answer 1

Separation of vegetable photosynthetic pigments by thin layer chromatography (TLC).

Aim: Extract the photosynthetic pigments and separate them by means of TLC.

Procedure: 1) - Isolation of pigments from spinach leaves. Mechanical breakdown of plant tissue (anhydrous magnesium sulphate + sand) for the release of the pigments and their extraction with acetone (polar solvent). Centrifuge to separate the liquid phase (pigments + acetone) from the cell debris. Caution: acetone is flammable and irritating, it is recommended to work with personal protection and under hood. 2) - Separation of pigments by TLC. To prepare the chamber where the TLC plate will be placed with the mobile phase - 70% hexane-acetone 30% - (Caution with the mixture, flammable and irritant). When the chamber is stabilized, the TLC plate is introduced to separate the pigments on it depending on the distance that each pigment travels from the place where it was initially placed.

Analysis of the results: to quantify the distance traveled by each pigment, the retention factor or retardation factor (Rf value) is calculated, which is the ratio of the distance traveled by the pigment and the distance traveled by the liquid phase (hexane-acetone in this case) from its baseline on the TLC plate. Expected results: the pigments will be separated in the stationary phase of the TLC plate, depending on their degree of solubility in the hexane-acetone mixture. The Rf values ​​in decreasing order for the pigments studied will be: carotenes > pheophytin a > pheophytin b > cholophyll a > cholophyll b > xanthophylls.