An Improved Method for the Determination of Hydroxyproline

An improved method for the determination of hydroxyproline (Hyp) in animal tissues has been established by modifying Kivirikko's and KISO methods. This method consists of the oxidation of Hyp by the addition of chloramine T at 0°C for 2 h in 0.5 M borate buffer (pH 8.2) containing 1 M KCl. It was confirmed that Hyp was satisfactorily oxidized to an oxidation intermediate, which was stable throughout the process of the oxidation at 0°C for 2 h. The degree of color development with the colorimetric reagent in this method was 2-3 times higher than that in conventional methods, and the reproducibility was invariably good. The present method is very useful for the microanalytical determination (0.05-4 μg/tube) of Hyp.