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Lab Menual Spectrophoto

This document outlines an experiment to perform paper chromatography to separate and identify components in leaf samples. Paper chromatography works by using a mobile phase like acetone or ethanol to carry compounds from a sample up a stationary chromatography paper. The distance each compound travels allows calculation of its Rf value, which can identify the compound. The experiment involves grinding leaf samples with solvent, applying the extract to paper, and developing the chromatogram by letting the solvent carry compounds up the paper. Marking the solvent front will allow calculation of Rf values to identify the leaf sample's components.

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312 views2 pages

Lab Menual Spectrophoto

This document outlines an experiment to perform paper chromatography to separate and identify components in leaf samples. Paper chromatography works by using a mobile phase like acetone or ethanol to carry compounds from a sample up a stationary chromatography paper. The distance each compound travels allows calculation of its Rf value, which can identify the compound. The experiment involves grinding leaf samples with solvent, applying the extract to paper, and developing the chromatogram by letting the solvent carry compounds up the paper. Marking the solvent front will allow calculation of Rf values to identify the leaf sample's components.

Uploaded by

Nityantini
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Download as DOCX, PDF, TXT or read online on Scribd
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Experiment: Paper Chromatography.

Aim : To obtain a paper chromatogram of various sample


: To identify components in sample by calculating Rf values

INTRODUCTION.

Chromatography is a method of physically separating mixtures into its individual components. It is a


common laboratory technique used to identify unknown components in mixtures.

There are several types of chromatography; all types employ a mobile phase or eluent (it can be
liquid or gas), which is forced through a stationary phase (a solid or semi-solid). Mixtures are
separated because some components will be more attracted to the stationary phase (and stick to it)
while some components will be more attracted to the mobile phase (and travel with it).

Rf values

Some compounds in a mixture travel almost as far as the solvent does; some stay much closer to the
base line. The distance travelled relative to the solvent is a constant for a particular compound as
long as you keep everything else constant - the type of paper and the exact composition of the
solvent, for example.

The distance travelled relative to the solvent is called the Rf value. For each compound it can be
worked out using the formula:

Apparatus: 600-mL beaker, pencil, ruler, plastic wrap, tape and chromatography paper

Chemicals: acetone, ethanol.

Procedures:

Part A: Preparation of Chromatography Paper

1. Wash your hands thoroughly to remove excess oils from your skin. Obtain a ruler and a piece
of chromatography paper from your instructor. Handle the paper only on the edges to avoid
leaving fingerprints, as these may hinder the elution process.

2. Place the chromatography paper on a sheet of clean notebook paper or paper towel to avoid
picking up dirt or contaminants from the bench top. Using a pencil and ruler to measure
accurately, draw a straight line across the paper, about 1.5 cm above the bottom edge. This is
the starting line. Draw another line about 5 cm above .This is the finish line.
3. On the starting line, measure in from one side about 2.5 cm and lightly draw a small “X”
centered on the starting line.

Part B: Chromatography of leaves

1. Finely cut up some leaves and fill a mortar to about 2 cm depth.


2. Add a pinch of sand and six drops of solvent from the teat pipette.
3. Grind the mixture for at least three minutes.
4. On a strip of chromatography paper, use a fine glass tube to put liquid from the leaf extract
onto the centre of the line. Keep the spot as small as possible.
5. Allow the spot to dry, then add another spot on top. Add five more drops of solution, letting
each one dry before putting on the next. The idea is to build up a very concentrated small spot
on the paper.
6. Put a small amount of solvent in a beaker and hang the paper so it dips in the solvent. Ensure
the solvent level is below the spot.
7. Leave until the solvent has soaked near to the top.
8. Mark how high the solvent gets on the paper with a pencil and let the chromatogram dry.

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