Improvements in the Separation of p-Carotene and Lycopene by Column Chromatography An excellent experiment for demonstrating column chromatography is the separation of &carotene and lycopene in carrot and tomato paste, respectively. The following is a modification of the experiment as written in Fieser and Williams.' Three grams each of carrot and tomato paste were placed in a beaker with 15 mL of 95% e t h a n ~ lThe . ~ mixture was well stirred with a glass rod for several minutes and filtered using vacuum filtration. and the residue was messed with another oiece of filter oaner . . to eet out as much liouid as oossihle. T h r nlrohul extract was discarded and the dehydrated residue u,as plilced'h n 50-ml. flask aiong u,i;h 10 ml. of dicllloromethnnt~.3Thrmixture was retluxed tor 6 min and then filtered usinr vacuum liltnition. Thr residue was returned to the flask and the procedure was repeated twice more. The &chloromethane extracts were combined, washed once with saturated sodium chloride solution, dried over sodium sulfate, and evaporated to dryness. The oily residue was taken up in 2 mL of low-boiling petroleum ether.' A 150 X ll-mm column of activity I alumina was prepared using low-boiling petroleum ether. The carotanoid mixture was applied to the alumina column in a narrow band using petroleum ether to wash down the sides of the column. After several milliliters of petroleum ether were passed through the column (the carotanoids should remain in a narrow hand a t the top of the column), the solvent was changed to 10%diethyl ether-petroleum ether. The &carotene separated as a distinct yellow hand from the orange lycopene. Once all the 8-carotene was eluted off, the solvent was switched to 50%diethyl ether-petroleum ether in order to elute off the lycopene. The solvent was evaporated from the two fractions and approximately 0.2 mL of dichloromethane was added to each residue. Thin-layer chromatography was done using silica gel plates and 5% chloroform-cyclohexane as the eluting solvent. The @-carotenegave a distinct yellow spot with an Rr of 0.4, and lycopene gave an orange spot with a n Rr of 0.2. This experiment requires about 3.5 h.
' Fieser, L. F. and Williams. K. L., "Organic Experiments." 5th ed.. D.C. Heath and Company. Lexington. MA. 1983, p. 253. -~~
Neither tomato nw carrot pastes contain sufficient amounts of both B-cardene and lycopene to demonstrate their separation on a column. If the alcohol and dichloromethane extracts are combined, then problems with emulsion are encountered, Low-boiling petroleum ether was sufficiently polar to dissolve most of the carotanoid residue. Mher solvents such as toluene or dichloromethane caused band widening on the column so that an incomplete separation of the &carotene and lycopene resulted. Peter Ronman Brevard Community College Melbourne Campus 3865 N. Wickham Road Melbourne. FL 32935
540
Journal of Chemical Education
