Hydroxylysine glycosides: preparation and analysis by reverse phase high performance liquid chromatography

Hydroxylysine diglycoside was prepared from hydrolyzed sponge by a combination of ion-exchange chromatography and gel filtration. The product obtained was chemically pure on amino acid analyzer and was active as substrate of the enzyme alpha-(1----2)glucosidase. Hyl monoglycoside was prepared by mild acid hydrolysis from diglycoside. A reverse phase HPLC system was devised for Hyl glycosides, hydroxylysine and other basic amino acids. The separation was achieved using an octadecyl bonded silica column on the compounds derivatized with dabsyl chloride to produce di-dabsyl derivatives. Elution was followed in the visible region at 436 nm. In the conditions used no or very low amount of mono-dabsyl derivatives was observed. Hyl di- and monoglycoside resulted a mixture of two diastereoisomers, which form during the alkaline hydrolysis. The separation of the diastereoisomers of each compound depended on pH and ionic strength of the eluent in the HPLC column, whereas they were not separated by our short column on amino acid analyzer. The HPLC system was also used for the analysis of Hyl glycosides on two collagen preparations.