Abstract
Thin-layer chromatography allows rapid separation of proline and hydroxyproline over a wide quantitative range. The present method utilizes the nitrous acid reaction to destroy amino acids in protein hydrolyzates. After ether extraction of amino acid derivatives, proline and hydroxyproline are present unchanged or as unstable n-nitroso derivatives and can be rapidly separated by thin-layer chromatography on cellulose. Mixtures including up to 500 μg of each imino acid are separable on a 250-μ thick layer. The imino acids can be eluted and determined colorimetrically. Studies using isotopically labeled proline and hydroxyproline are possible even when low specific activities are involved. This simple and rapid technique has proved useful in determination of the specific activity of radioactive protein bound hydroxyproline in crude collagen extracts.
Published Version
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