Chromatographic Separation Alina Guna Ryan Young Edward Sun Jonathan Lam Mr. Vincent SCH3U3

Contents Introduction to Chromatography Applications Conclusion Adsorption Paper Chromatography Experiment Materials Procedure Observations and Calculations Video Applications Conclusion Sources of Experimental Error

Ex: Chromatography in Action Introduction Chromatography – Separating Mixtures Includes a mobile/stationary phase Preparative or Analytical Preparative: Separate to use components Analytical: Measure relative proportion of a substance 5 different types of Chromatography: Adsorption Partition Ion Exchange Molecular Exclusion Affinity Ex: Chromatography in Action

Adsorption Chromatography When a mobile gas or liquid collects onto a stationary solid Forms a thin layer of molecules Solvent travels up adsorbent to meet sample Continues upwards as a solvent/solute solution (adsorbate) Different compounds in sample Different solubilities Travels different distances Analyze Identify Purify Quantify Separate Mixture Components

The other types … Partition Chromatography Ion Exchange Separation of similar substances by repeated extraction by two immiscible liquids. Ion Exchange Separation of ions and polar molecules Molecular Exclusion Particles are separated based on size. Usually applied to large molecules such as proteins and industrial polymers Affinity Method of separating biochemical mixtures We focus on Adsorption in our project.

Paper Chromatography Stationary Phase Mobile Phase Chromatography Paper Mobile Phase Unreactive solvent Primarily used to separate and identify coloured mixtures Two-way paper chromatography Complicated substances Amino Acids Solvent’s ability to move upwards via capillary action

Process of Paper Chromatography

Experiment Explain adsorption What it is How it is used to separate mixtures Demonstrate adsorption through paper chromatography Calculate retention factors of the substances

Safety Concerns Safety Goggles Rubbing Alcohol Ventilation of Area Spillage/Leakage Consumption Ventilation of Area Permanent Markers Condition Wastage

Materials Glass Jars Chromatography Paper Pencils Rubbing alcohol (70% Isopropyl Alcohol) Sharpie Pens of various colours Measuring cup Ruler Scissors Tape Safety Goggles

Procedure Use the scissors and ruler to cut equally long strips of Chromatography Paper. With a pencil, draw a horizontal line 1 cm above the bottom edge of each strip. Using each Sharpie pen, place a spot along the line of the Chromatography Paper. Tape each strip onto a different pencil, and place the pencil onto the jar. Make sure that the strip of Chromatography Paper is touching the Isopropyl Alcohol inside. Measure approximately 15 mL of Isopropyl Alcohol and pour it in each jar. Let the strips develop until the ascending solution is approximately 2 cm from the top of the paper. Then remove the strips and let them air dry. Record your observations and determine the retention factor.

Data Spot Colour Adsorbate Colour Distances Travelled (Respective) Distance Travelled by Solvent Retention Factor (Respective) Black Blue, Orange, Red 4.3cm, 1.6cm, 3cm 4.3cm 1, 0.3271, 0.6977 Gray Purple 1 Blue 3.9cm Green Green, Yellow 4.1cm, 4.4cm 4.4cm 0.9318, 1 Light Green 4cm, 4.4cm 0.9090, 1 Orange Orange, Red 3.1cm, 4cm 4cm 0.7750, 1 Burgundy Green (2) 3.8cm, 4.2cm 4.2cm 0.9048, 1 Orange (2) 3.2cm, 4.2cm 0.7619, 1 Burgundy (2)

Calculations Black Ink Blue Dye Red Dye Orange Dye

Applications Anything involving separating a solution Medicine Sports Medicine – Testing for drug usage (Gas chromatography) Pharmaceutical industries – Check if the right reactants were used. Criminology & Forensic Science DNA sequencing (Capillary electrophoresis) Analyzing evidence (Thin-layer chromatography) Petrochemical Industries Checking the purity of fuel (Gas chromatography)

Sources of Experimental Error Chromatography Paper Condition Measurements of Distance Marker Dot Size

Experiment Modifications More Solutes Different Colours Different Rf Values E.g. Food Colouring, Homemade mixture Different Solvents Different Concentrations of Isopropyl Alcohol

Experiment Modifications Testing known Solvents E.g. Known chemical formula Easier to find relationships Record speed of Solution Relationships with respect to time

Conclusion Intermolecular Forces Retention Factor Stronger forces = Slower speed traveling upwards Longer time in stationary phase Retention Factor Can help identify an unknown compound Surface tension of Markers All solutes tested were polar