Related Experiment Video
Updated: Jul 26, 2026

Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification
Published on: September 21, 2011
A new procedure for the specific high-performance liquid chromatographic determination of hydroxyproline
P A Biondi1, L M Chiesa, M R Storelli
1Istituto di Fisiologia Veterinaria e Biochimica, Milano, Italy.
Abstract:
A procedure suitable for a selective high-performance liquid chromatographic (HPLC) analysis of the imino acid hydroxyproline in the presence of a large excess of amino acids is proposed. To deaminate the amino acids, the well-known reaction with nitrous acid is exploited. The N-nitroso derivatives of imino acids obtained are extracted in ethyl acetate, denitrosated by hydrobromic acid, and treated with dabsyl-chloride. The final HPLC separations are carried out on a reversed-phase column in a rapid isocratic run. The use of the cis isomer of hydroxyproline as an internal standard allows good reproducibility. As an application of the described method, the hydroxyproline content in samples containing collagen and an excess of bovine serum albumine (up to 20:1) is determined.
Related Concept Videos
High-Performance Liquid Chromatography: Introduction
In HPLC, two phases play a critical role in the separation process:
High-Performance Liquid Chromatography: Elution Process
High-Performance Liquid Chromatography: Instrumentation
High-Performance Liquid Chromatography: Types of Detectors

