RAPID-DETERMINATION OF DESMOSINE AND ISODESMOSINE IN TISSUE HYDROLYSATES BY ISOCRATIC HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY AND PRECOLUMN DERIVATIZATION

A rapid and sensitive isocratic high performance liquid chromatographi c method has been developed for the single and specific determination of low concentrations of desmosine (Des) and isodesmosine (Ide), the m ajor specific crosslink aminoacids in elastin. Samples of isolated ela stin or whole tissue were hydrolysed in 6N HCl, and the hydrolysates w ere prefractionated on cellulose CF1. Des, Ide, gamma-glutamyl-glutami c acid as internal standard were dansylated and derivatives were extra cted from reaction mixture by ethylacetate. Their separation on a Lich rosphere 100-NH2 column, using methanol-water as mobile phase containi ng acetic acid and 0.25 M sodium acetate, final pH 6.5, was followed b y fluorescence detection (340-510 nm). The overall reproducibility was 5.9% for Des and 5.0% for Ide. The limits of detection were 2.2 pmol and 2.5 pmol, respectively. The method was successfully applied for th e determination of Des and Ide in normal pig aortas.