Abstract
A method of thin-layer chromatography was developed for the separation of the 2,4-dinitrophenylhydrazones of eight keto-acids. When petroleum ether-ethyl formate/monocarboxylic acid was used as the solvent system higher RB values of the components—measured with reference to 2,4-dinitrophenylhydrazine—were obtained with increasing acid concentration or with decreasing molecular weight of the acid in the solvent system. If the mobile phase contained even small amounts of formic acid, some of the compounds gave two successive spots due to isomers. With higher homologues of formic acid no separation of isomers could be observed when the acid concentration was low. When the concentration was raised, the number of components that gave two spots also increased. Owing to the extraction procedure to which the precipitates were subjected some of the compounds were not stable in solution. A thicker layer of Kieselgel G and also addition of acid to the adsorbent-water mixture before coating caused the RB values to decrease. The separation of the individual keto-acids from the mixture could be improved by acidifying the spreading mixture and using a thicker layer.
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