Leaf Chromatography Instructor guide and answer key Materials: Note: these are the items needed for each group (2-3 students). Multiply by the number of groups you will need. • • • • • • • •
1 jar or test tube (50mL beakers work great!) 1 piece of chromatography paper approximately 12cm x 3cm Coin or some other object for transferring pigment onto the leaf. Leaf material Pencil Rubbing alcohol (just enough to cover the bottom of the jar or test tube) Ruler Student lab
Instructor Prep The nice thing about this lab is that it involves very little prep for the instructor. You will need to gather the alcohol, jars, leaves, and chromatography paper. To save time, you may choose to cut the chromatography paper down to size in advance. Depending on the age and trustworthiness of your students, you may decide to pour the rubbing alcohol into the jars before class.
Safety concerns The only dangerous material in this lab is the rubbing alcohol. The MSDS from Flinn Scientific has been provided to alert you to possible safety issues.
Suggestions Chromatography paper is a cotton based paper that is highly absorbent. It can be purchased from Flinn Scientific by the roll or in individual sheets. If cost is an issue, coffee filters can also be used. Purchase white, round coffee filters, flatten them out and cut them into strips. Common rubbing alcohol (70% alcohol, 30% water) can be used. Any type of leaf can provide pigment, however some have greater amounts of pigment than others. If this lab is done in the winter when leaves are not available, spinach leaves can be used.
Leaf Chromatography © John R. Sowash | March 2009 | Permission to redistribute granted
Common mistakes 1. Drawing on chromatography paper with pen. • Remind students that ink is a pigment just like chlorophyll. If they use a pen, their line will slowly move up the paper! 2. Not transferring enough pigment onto the chromatography paper. • A generous amount of pigment is required in order to achieve clear separation. Instruct students that they can’t have too much! 3. Touching the leaf pigment to the alcohol at the bottom of the jar. • If the pigment touches the alcohol, it will immediately dissolve into the liquid at the bottom of the jar and will not move up the chromatography paper. 4. Not leaving the chromatogram in the alcohol long enough to allow the pigments to separate. • The chromatogram should be left in the jar until the solvent front (alcohol) reaches the top of the chromatography paper. This will take approximately 30 min.
Further Investigation 1. Experiment with different types of leaves. Which leaf has the greatest amount of pigment? 2. Practice chromatography using other pigments. Visa-vie markers work great!
Leaf Chromatography © John R. Sowash | March 2009 | Permission to redistribute granted
Answer Key
Chlorophyll a
Chlorophyll b
Xanthophyll
Carotene
Pigment Origin
2cm
2cm
2cm
2cm
Pigment front
Will vary
Will vary
Will vary
Will vary
Solvent front
Near 12cm
Near 12cm
Near 12cm
Near 12cm
Rf Value
Will vary
Will vary
Will vary
Will vary
Rf = distance moved by pigment from original spot distance moved by solvent from original spot
Pigment Identification: Chlorophyll a= blue-green Chlorophyll b = Olive green Xanthophyll = yellow Carotene = orange yellow Post-Lab Questions: 1. Why is chromatography useful? Chromatography is a process that helps separate pigments that have been mixed together. It is commonly used in forensic science.
Staple Filter Paper Here
2. Why are chlorophyll a & b green? Chlorophyll a & b are green because they reflect green light.
Leaf Chromatography © John R. Sowash | March 2009 | Permission to redistribute granted
3. Why do leaves change colors in the fall? Leaves turn different colors in the fall because chlorophyll a & b are not being made. As these pigments dissipate out of the leaves, the accessory pigments become visible.
4. How could you predict the color a tree’s leaves will turn in the fall? You could predict the fall color of a leaf by separating out the pigments found in the leaf. The fall color will be a combination of the accessory pigment colors.
5. Use the chromatogram below to complete the data table.
Solvent Front (12cm from solvent front) Xanthophyll (9cm) Carotene (7.7cm)
Chlorophyll a (5.2cm)
Chlorophyll b (4.7cm)
Pigment/Solvent Origin (1cm)
Chlorophyll a
Chlorophyll b
Xanthophyll
Carotene
Pigment origin
1cm
1cm
1cm
1cm
Pigment front
5.2cm
4.7cm
9cm
7.7cm
Solvent front
12cm
12cm
12cm
12cm
Rf Value
0.43cm
0.39cm
0.75cm
0.64cm
Leaf Chromatography © John R. Sowash | March 2009 | Permission to redistribute granted
FLINN SCIENTIFIC INC. "Your Safer Source for Science Supplies"
Material Safety Data Sheet (MSDS)
MSDS #:
420.50
Revision Date:
February 24, 2003
Section 1 — Chemical Product and Company Identification
Isopropyl Alcohol Flinn Scientific, Inc. P.O. Box 219 Batavia, IL 60510 (800) 452-1261 CHEMTREC Emergency Phone Number: (800) 424-9300 Section 2 — Composition, Information on Ingredients Isopropyl Alcohol (67-63-0) 70%, and Water (7732-18-5) 30% Synonym: 2-propanol, rubbing alcohol CAS#: None Established Section 3 — Hazards Identification Clear colorless liquid; distinctive odor; like rubbing alcohol. Irritant to body tissues. Slightly toxic by ingestion and inhalation. The single lethal dose for a human adult is about 400 mL, although as little as 150 mL can be fatal.
FLINN AT-A-GLANCE Health-1 Flammability-2 Reactivity-1 Exposure-0 Storage-3 0 is low hazard, 3 is high hazard
Section 4 — First Aid Measures Call a physician, seek medical attention for further treatment, observation and support after first aid. Inhalation: Remove to fresh air at once. If breathing has stopped give artificial respiration immediately. Eye: Immediately flush with fresh water for 15 minutes. External: Wash continuously with fresh water for 15 minutes. Internal: Induce vomiting. Call a physician or poison control at once.
Section 5 — Fire Fighting Measures Combustible liquid. Flash Point: 75 F Upper: 12% Lower: 2.5% Autoignition Temperature: 860 F When heated to decomposition, emits acrid smoke and fumes. Fire Fighting Instructions: Use triclass, dry chemical fire extinguisher. Firefighters should wear PPE and SCBA with full facepiece operated in positive pressure mode.
NFPA CODE H-1 F-0 R-0
Section 6 — Accidental Release Measures Restrict unprotected personnel from area. Remove all ignition sources and ventilate area. Contain spill with sand and absorbent material; deposit in sealed bag or container. See Sections 8 and 13 for further information.
Section 7 — Handling and Storage Flinn Suggested Chemical Storage Pattern: Organic #2. Store with alcohols, glycols, amines and amides. Store in a dedicated flammables cabinet. If a flammables cabinet is not available, store in Flinn Saf-Stor can. Store in a cool dry place. Use and dispense in a hood. Section 8 — Exposure Controls, Personal Protection Avoid contact with eyes, skin and clothing. Wear chemical splash goggles, chemical-resistant gloves and chemical-resistant apron. Use ventilation to keep airborne concentrations below exposure limits. Always wear a NIOSH-approved respirator with proper cartridges or a positive pressure, air-supplied respirator when handling this material in emergency situations (spill or fire).
© 2003 Flinn Scientific, Inc. All Rights Reserved.
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FLINN SCIENTIFIC INC. "Your Safer Source for Science Supplies"
Material Safety Data Sheet (MSDS)
Isopropyl Alcohol MSDS #:
420.50
Revision Date: February 24, 2003
Section 9 — Physical and Chemical Properties Clear colorless liquid. Solubility: Water soluble, soluble in alcohol and ether. Formula: C3H8O Formula Weight: 60.11
Specific Gravity: .8745 (70%) Boiling Point: 81.65 C (70%)
Section 10 — Stability and Reactivity Avoid contact with strong oxidizers, acetaldehyde, chlorine, ethylene oxide, acids, isocyantes. Shelf life: Poor.
Section 11 — Toxicological Information Acute effects: Severe eye irritant, nausea, headache, vomiting Chronic effects: N.A. Target organs: Nerves, kidneys
ORL-RAT LD50: N.A. IHL-RAT LC50: N.A. SKN-RBT LD50: N.A.
N.A. = Not available, not all health aspects of this substance have been fully investigated. Section 12 — Ecological Information Data not yet available. Section 13 — Disposal Considerations Please consult with state and local regulations. Flinn Suggested Disposal Method #18a is one option.
Section 14 — Transport Information Shipping Name: Not regulated Hazard Class: N/A UN Number: N/A N/A = Not applicable
Section 15 — Regulatory Information TSCA-listed, EINECS-listed (200-661-7), RCRA code D001. Section 16 — Other Information Consult your copy of the Flinn Scientific Catalog/Reference Manual for additional information about laboratory chemicals. This Material Safety Data Sheet (MSDS) is for guidance and is based upon information and tests believed to be reliable. Flinn Scientific Inc. makes no guarantee of the accuracy or completeness of the data and shall not be liable for any damages relating thereto. The data is offered solely for your consideration, investigation, and verification. Flinn Scientific Inc. assumes no legal responsibility for use or reliance upon this data.
FLINN SCIENTIFIC INC. "Your Safer Source for Science Supplies"
Flinn Is No. 1 in Safety
[email protected] www.flinnsci.com P.O. Box 219 Batavia IL 60510 (800) 452-1261 Fax (866) 452-1436
© 2003 Flinn Scientific, Inc. All Rights Reserved.
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