Chromatography Lab

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Environmental Health Sciences Center

Community Outreach & Education Program

(Adapted from: Forensic Science Activities. University of Colorado Boulder Hughes Initiative. .) DESCRIPTION: Students will use paper chromatography to separate ink molecules and identify the pen used on an unknown sample of handwriting. Students will graph and analyze data they collect using paper chromatography. PURPOSE/GOAL: Students will be able to: • Gain understanding of the purpose of chromatography. • Measure and graph pigment separation. • Use evidence gathered from the chromatograms to support their conclusions about which suspects can be excluded from suspicion based on evidence. • Understand that the separation of dye molecules in chromatography is a physical property of the dye and movement of the dye is unique for each different dye molecule. • Defend their conclusions about the dyes based on the evidence that they gathered. TIME ESTIMATE: Prep: 60 minutes (cutting paper stripes and marking the crime scene strips takes a lot of time.) Activity: 90 minutes + answering the questions as homework MATERIALS: • Hydroville Science Journal • Pencil • Ruler Per group of four students: • Test tubes or culture tubes (20 X 150 mm) in test tube rack • Strips of chromatography paper (1 x 16 cm) • Strip of chromatography paper with crime scene ink (1 x 16 cm) Class Material on Tool Table: • set of black ink pens labeled “A”, “B”, “C”, and/or • set of black ink pens labeled “D”, “E”, “F” • Paper towels • Colored pencils • Wash bottle with rubbing alcohol MATERIALS TO PHOTOCOPY: • Using Paper Chromatography - Instructions • Team Worksheet • Student Worksheet MYSTERIOUS ILLNESS OUTBREAK – Investigation of an Outbreak HYDROVILLE CURRICULUM PROJECT ©2004, Oregon State University Funded by NIEHS grant #66 R25 ES10721


Environmental Health Sciences Center

Community Outreach & Education Program

BACKGROUND INFORMATION: What is Chromatography? Chromatography is a laboratory method that is widely used for the separation, identification, and determination of chemical components of a complex mixture. More specifically, chromatography separates compounds based on differences in their structure, size, and/or composition. Analytical chemistry uses chromatography to conduct qualitative analysis (identify the components) and quantitative analysis (determine the concentration) of unknown substances. No other separation method is as powerful and generally applicable as chromatography. Qualitative Analysis – identifying chemicals (what kind?) • Species identification , e.g., "killer" bees can be distinguished from native bees by comparing gas chromatograms of cuticle extracts • Tracing contraband sources and detecting drugs in urine Quantitative Analysis – finding concentrations (how much?) • Each peak corresponds to a separate component in the mixture • Area of each peak is proportional to concentration How Does Chromatography Work? There are numerous different chromatography techniques; however, they all use a stationary phase and mobile phase. The part that stays in one place is called the stationary phase, and the part that moves is called the mobile phase. Substances have different attractions for the stationary and the mobile phases and can be separated by these different attractions. Components of a mixture are carried through the stationary phase by a flow of a gaseous or liquid mobile phase. Each sample will migrate through the stationary phase at a different rate. Using Paper Chromatography In paper chromatography, a small amount of the substance to be analyzed (analyte) is placed on a strip of paper (the stationary phase) above the level of the solvent (mobile phase). In this activity, you will be using ink as the analyte and alcohol as the solvent. As the alcohol moves up the paper, the dye molecules from the ink mixture will move with it. If they are more strongly attracted...
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