The Arizona STEM Acceleration Project
Exploring Plant Pigments through Paper Chromatography
Exploring Plant Pigments through Paper Chromatography
Grades 6-9
Biology, Botany, Environmental Science
Reny Mathew
11/28/23
Notes for teachers
The Rf (retention factor) values of pigments can vary depending on the specific chromatographic conditions used, including the type of chromatography (e.g., paper chromatography, thin-layer chromatography), the type of stationary and mobile phases, and the solvent system. Additionally, the Rf values can be influenced by factors like temperature and humidity.
However, here are approximate Rf values for some common plant pigments when using a standard solvent system for chromatography (e.g., acetone or ethanol as the mobile phase):
Chlorophyll a: Rf ≈ 0.3 - 0.4
Chlorophyll b: Rf ≈ 0.2 - 0.3
Carotenes : Rf ≈ 0.8 - 0.9
Xanthophylls : Rf ≈ 0.4 - 0.6
These values are approximate and can vary based on the specific conditions of the chromatography.
List of Materials
Standards
MS-PS1-2. Analyze and interpret data on the properties of substances before and after the substances interact to determine if a chemical reaction has occurred. (Grades 6 - 8)
HS.L2U1.21 Obtain, evaluate, and communicate data showing the relationship of photosynthesis and cellular respiration; flow of energy and cycling of matter.
Science and Engineering Practices
Objectives:
Agenda (90 minutes)
Introduction
Students will observe the phenomenon and teacher poses the question: What do you notice in the picture?
Ask students to write their observation on a T-chart
What do you notice? | What do you wonder? |
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Students will work in groups and discuss
Teacher will elicit response from each group by asking: Why do you think that happened?
Student answers varies
Teacher explains the phenomenon
What is chromatography?
How does paper chromatography work?
This technique uses paper as the stationary phase. The mixture is applied to a paper strip, and the solvent moves through the paper, separating the components.
The principle behind the paper chromatography is that the most soluble substances move further on the filter paper than the least soluble substances. Different plant pigments can be separated by using the technique of paper chromatography.
What is Retention factor (Rf)?
The retention factor (Rf) is a quantity used in chromatography to quantify the separation of components in a mixture.
It is a ratio that describes the distance a particular component travels compared to the distance traveled by the solvent front (the furthest point reached by the solvent) during the chromatographic separation.
The formula for calculating the retention factor (Rf) is: Distance traveled by the solute / Distance traveled by the solvent front
Chromatography is a laboratory technique used to separate and analyze mixtures of chemicals. It is based on the principle that different components in a mixture have different affinities for a stationary phase and a mobile phase. The stationary phase is typically a solid or liquid material, and the mobile phase is a liquid or gas.
Activity:
Students will work in groups of two and practice lab techniques.
Preparation of Spinach Extract: (a.)Collect fresh spinach leaves and wash them thoroughly with water to remove any surface contaminants. (b.)Dry the leaves using a paper towel.(c.) Cut the spinach leaves into small pieces.(d.) Use a mortar and pestle to grind the spinach leaves, adding a small amount of extraction solvent (acetone or ethanol) to create a pigmented extract. (e.)Filter the extract to remove solid particles. Collect the liquid in a small beaker or glass jar.
Preparing the Chromatography Paper: (a.) Cut a strip of filter paper or chromatography paper to the desired length (usually a few centimeters wide and long enough to hang into the developing chamber).(b.) Use a pencil to draw a baseline near the bottom of the paper strip, about 1-2 cm above the edge.
Application of Spinach Extract: Place a small spot of the spinach extract on the baseline, slightly above the bottom edge of the paper. Be careful not to overload the paper.
Developing the Chromatogram:(a.)Pour a small amount of the extraction solvent into the developing chamber, ensuring that the solvent level is below the baseline.(b). Hang the chromatography paper in the chamber, making sure that the spot with the extract is above the solvent level.(c.)Seal the chamber with a lid to create a closed environment.
Observation:
(a.) Allow the solvent to move up the paper until it nearly reaches the top.
(b.) Remove the paper from the chamber and mark the solvent front.
(c.) Observe and mark the positions of any separated pigments on the paper.
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Example to calculate Rf value
Students will work in groups of two and their final assessment is based on their analysis of the lab.
Students will measure the distance from the baseline to each pigment spot and the distance from the baseline to the solvent front.
They will calculate the Rf values for each pigment using the formula:
Distance traveled by the pigment / Distance traveled by the solvent front
Students will discuss reflection questions in their groups:
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ASSESSMENT
Students understand the information and can elaborate, explain, and answer questions about the pigments in plants.
Students will be able to complete their lab report.
Students will work in groups and discuss the reflection questions.
Differentiation
For struggling students, instead of having students mark their own observations, provide a template with pre-determined observation points for where pigments might appear.
Encourage students to focus on changes in color without requiring them to interpret complex chromatograms.
Struggling learners can be paired with more confident peers to gain support. They can be allowed extended time if needed. Some struggling learners may need more time to process information and complete the task.
Remediation
Extension/Enrichment
For students who grasp the material quickly, create extension activities or projects. This could involve additional research, experimentation, or real-world applications.
Students can be offered choice assignments that allow students to explore topics related to chromatography in real life situations.
For example, how might the ability to separate solutions into their components be important to environmental engineers looking into water quality issues like water contamination?