Chromatography
DEFINITION CHROMATOGRAPHY The separation of a mixture by distribution of its components between a mobile and stationary phase over time mobile phase = solvent stationary phase = column packing material
HISTORY Chromatography (from Greek :chromatos -- color, "graphein" -- to write) 1903 Tswett - plant pigments separated on chalk columns 1931 Lederer & Kuhn - LC of carotenoids 1938 TLC and ion exchange 1950 Reverse phase LC 1954 Martin & Synge (Nobel Prize) 1959 Gel permeation 1965 instrumental LC (Waters)
Purpose of Chromatography Analytical - determine chemical composition of a sample Preparative - purify and collect one or more components of a sample
Uses for Chromatography Real-life examples of uses for chromatography: Pharmaceutical Company determine amount of each chemical found in new product Hospital detect blood or alcohol levels in a patient s blood stream Law Enforcement to compare a sample found at a crime scene to samples from suspects Environmental Agency determine the level of pollutants in the water supply Manufacturing Plant to purify a chemical needed to make a product
Classification of Methods There are two classification schemes: mobile phase attractive forces
Mobile Phase Gas (GC) Water (LC) Organic solvent (LC) Supercritical fluid (SCFC)
Classification based on Mobile Phase Gas Chromatography Gas - solid Gas - liquid Stationary Phase
Classification based on Mobile Phase Liquid chromatography (LC) Column (gravity flow) High performance (pressure flow) Thin layer (adsorption)
Classification based on Attractive Forces 1. Adsorption 2. Ion Exchange 3. Partition 4. Size Exclusion
Adsorption Chromatography Separation based on their adsorption onto the surface of solid (stationary phase). Normal phase-like separation Nonpolar mobile phase for polar non-ionic compounds Ex; Column chromatography
Partition Chromatography solute are separated based on their partition between a liquid mobile phase and a liquid stationary phase coated on a solid support. Normal analyte is nonpolar organic; stationary phase MORE polar than the mobile phase Reverse analyte is polar organic; stationary phase LESS polar than the mobile phase Ex : TLC, Paper Chromatography Phase 2 Phase 1 Phase 2 Phase 1
Ion Exchange Chromatography Use ionic stationary phase ions separated on the basis of their tendency to displace counter ions adsorbed on stationary phase (Depends on charge, hydration, solubility ) Anionic stationary phases: used for cation separation Cationic stationary phases : for anion separation for ionic compounds - Ex : CC (K.turus), HPLC
Size Exclusion Chromatography Separation is a result of trapping of molecules in the pores of the packing material Very large molecules can t get into the pores unretained Very small molecules get hung up in to pores for a long time - most retained longest retention time stationary phase is a porous matrix Ex: CC, HPLC
Types of Chromatography Liquid Chromatography separates liquid samples with a liquid solvent (mobile phase) and a column composed of solid beads (stationary phase) Gas Chromatography separates vaporized samples with a carrier gas (mobile phase) and a column composed of a liquid or of solid beads (stationary phase) Paper Chromatography separates dried liquid samples with a liquid solvent (mobile phase) and a paper strip (stationary phase) Thin-Layer Chromatography separates dried liquid samples with a liquid solvent (mobile phase) and a glass plate covered with a thin layer of alumina or silica gel (stationary phase)
STATIONARY PHASE Type of chromatography Material Paper chromatography (KK = kertas kromatografi) Thin Layer Chromatography (KLN = Kromatografi lapisan nipis) Gas chromatography (GC) High Performance Liquid Chromatography (KCPT = kromatografi cecair prestasi tinggi) Filter paper, cellulose Silica gel, alumina, polyamide Squalene, apezion, carbowax M C-8, C-18, Licosorb, Silicone
MOBILE PHASE Type of chromatography Solvent Paper chromatography (KK = kertas kromatografi) Thin Layer Chromatography (KLN = Kromatografi lapisan nipis) Gas chromatography (GC) High Performance Liquid Chromatography (KCPT = kromatografi cecair prestasi tinggi) Air, alcohol Hexane, ether petroleum, alcohol. He, Ar, N 2 Cyclohexane, n-hexane, carbon tetrachloride, ethanol, methanol, air