Determination of radioactive proline and hydroxyproline by a simple paper-chromatographic procedure

A simple method to separate and determine radioactive proline and hydroxyproline by paper chromatography is described. The localization of the imino acids after separation is achieved by direct nondestructive staining with 1-fluoro-2,4-dinitrobenzene dye. The imino acids are quantitated directly by liquid scintillation counting in the presence of paper strips. The method was applied to bone cultures with good reproducibility, sensitivity and linearity over a wide range of radioactivity. The procedure was also tested in fibroblast cultures. The results for hydroxyproline were in good correlation with the widely used method of Juva and Prockop (Juva, K. and Prockop, D.J. (1966) Anal. Biochem. 15, 77-83), in which hydroxyproline is oxidized to pyrrole, and then extracted and purified by column chromatography before counting radioactivity.