only touch plastic side * Organic Chemistry 205 Laboratory Experiment - A. Ahmed CHROMATOGRAPHY "TLC" Chromatography is involved in the most modern and sophisticated methods of separating mixtures available to the organic chemist. Chromatography is defined as the separation of mixture of two or more different compounds or ions by distribution between two phases, one of which is stationary and the other of which is moving (mobile). Various types of chromatography are possible, depending on the nature of the two phases involved: A. Solid-liquid: a. Column chromatography (sizing and ion-exchange, affinity columns). b. Thin layer chromatography [TLC] c. Paper chromatography [PC] B. Liquid-liquid: High performance liquid chromatography [HPLC, FPLC] C. Gas-liquid (vapor-phase): Gas chromatography [GC] All chromatography works on much the same principle. Basically, the method depends on the differential solubilities or absorptivity of the substances to be separated relative to the two phases between which they are to be partitioned. In solid-liquid chromatograph the solids used most commonly are silica gel SiO2.xH2O, alumina A1203.xH2O and sepharose. The possible interactions of organic compounds with alumina involve: coordination interaction, dipole-dipole interaction, or hydrogen bonding interaction. Solvent Selection is one of the parameters affecting the separation. Developing the TLC plates: UV light and lodine vapor in order to measure the Rf values. Identification of Drugs: TLC analysis of Analgesic Drugs Factors affecting the separation in TLC: solvent system - adsorbent-thickness of the adsorbent layer-relative amount of material spotted. The TLC plates will be used are silica gel coated with a fluorescent indicator. cetami * Different drugs will be used; Acetaminophen, Aspirin, Caffeine, Ibuprofen, Acetanilide, Salicylamide, and a mixture. Procedure: * Dissolve the drugs in a mixture (1:1) of Ethanol: Methylene chloride. (One tablet per 10 mL. * The developing solvent is 0.5% glacial acetic acid in ethyl acetate. Spot diameter about 1-2 mm. Run the solvent up to 0.5 cm from the top. Baseline *The plates dimension about 10 x 6.6 cm. Rf value (retardation value) = distance traveled by substance / distance traveled by solvent front. * Recording: the Rf values of the used drugs and the unknown sample. * draw w/ pencil Compare reactant + produc The Lab Report: briefly outline the procedure - calculations-R+values of each drug and the name of the unknown sample. 6 Samples 1) a, aspirin 3 See spors under uv JUST a dash 6-salicylic acid 28 2/3 drops of 1:1 2) a. acetanilide 4 b. aniline 4.4 (3) a. dibromostilbine 4 6. stilbene 4.2 0.5cm 8 Rf = 12 b 12cm or 6 36 R₁ = 12 Rf Baseline 115 #20, 338 Organic Chemistry 205 Laboratory Experiments >6 spots w/ flouricine -> See under UV (4x6cm) w/ lodine stains for next week touch only 120 Solvent APPLICATION OF THIN-LAYER CHROMATOGRAPH IN ORGANIC CHEMISTRY t A. Ahmed Thin-layer chromatography has several important uses in organic chemistry and environmental chemistry. It is reflect the distribution of a compound between a fixed phase (adsorbent layer on a plate) and a mobile phase (solvent system) allow to determine the R-values. The TLC can be used in the following applications: a. To examine that two compounds are identical b. Determination the number of components in a mixture C. Determination the good solvent (or solvent system) for a mixture separation d. Monitoring a column chromatograph separation of a mixture components e. Check the purification efficiency of different methods. Monitoring organic reaction progress. f. g. Detect pollutants in по N collected samples (water, soil) after extraction. 5cm After After 5 mins 10mins C D R: = distance travelled by the component distance travelled by the solvent P Steps of Thin Layer Chromatography H Fraciun Original mixture - SOR 916 abi The advantage of using TLC is using small amount of materials about (107g) can be observed. The disadvantage of TLC is dealing with the volatile materials. The Rf (retardation factor) value under same conditions is fixed for a certain compound, and it is the ratio between the distance that compound ravel to the distance that solvent front travel. The condition that affect the Rf value include: a. Type of the Adsorbent b. Solvent system c. Thickness of the adsorbent layer d. Relative amount of the applied (spotted) materials. TLC jar TLC plate Wet filter paper Mobile phase lova mo HOMEWORK: 1. In TLC experiment Calculate the Rf value of a spot that travels 5.0 cm, with a solvent front that travels 15.0 cm. What are the factors affecting the Rf value? 2. Types of chromatograph used in the purification and identification of organic compound? 3. What is the difference between one and two dimension TLC? 37