A simple, rapid, and reliable technique for identifying small amounts of phenylthiohydantoin (PTH) derivatives has been developed for use in the analysis by Edman degradation of small amounts of peptides isolated from fingerprints (10). Solvent systems used in paper and column chromatography of PTH amino acids have been adapted for use in thinlfayer chromatography (TLC) on precoated flexible TL sheets with an inborporated fluorescent indicator (7). Twenty PTH amino acids can be idgntified on one TL sheet by ascending chromatography in one direction by the repeated utilization of two different solvent systems.
Methods
The PTH amino acids were synthesized as described elsewhere (2-4) or purchased from Mann Research Laboratory, Inc., New York. n-But&nol, ethylene chloride, formic acid, heptane, and propionic acid were reagent-grade products and were used without further purification.
Eastnian Chromagram sheets type K301R with fluorescent indicator were used. The sheets were activated at loo', 30 min before use. A line was scored 5 mm from each edge of a sheet in the direction of chromatography in order to prevent front line convection. Samples were applied 1.5 cm from one end and l-l.5 cm apart.
The compositions of the four standard solutions, each of which contained 10 IJ;moles of each PTH amino acid/ml 90% acetic acid, are shown in Figures 1 and2; 0.5-l ,pl of each standard solution was applied under a str.eam of nitrogen.
Glass jars made for TLC on 20 X 20 cm glass plates were used. The jars were lined with Whatman 3MM paper on all four sides. The lining papers were wet with the appropriate solvent before each run. The Chromagram sheet was equilibrated in the jar for 0.5 hr before the solvent system was introduced. Ascending chromatography was performed for l-2 hr. In our hands the "sandwich" technique recommended by Przybylowicz et al. ( 7) failed to give satisfactory results.
Several solvent systems were tried, including those used earlier for