Comparative Study of Carbohydrate-Protein Complexes II. Determination of Hydroxylysine and Its Glycosides in Human Skin and Scar Collagens by an Improved Method

Abstract

A modification of the existing methods for measuring hydroxylysine, galactosyl-hydroxylysine, and glucosylgalactosylhydroxylysine is described. The method is based on analysis with an automated amino acid analyzer using a conventional separation system for basic amino acids. The prior removal of acidic and neutral amino acids was necessary. This was achieved by passing an alkaline hydrolysate of collagen through a column of Amberlite CG-120. Type II (H⁺) and washing the column with 8% aqueous pyridine. A basic fraction containing the hydroxylysine compounds was then recovered from the column by elution with 3 M NH⁴OH. Model experiments showed that hydroxylysine and its glycosides could be analyzed within an hour and that recoveries exceeded 90%. This method was applied to human tissues to investigate whether the dermal scar is different in collagen Composition from normal skin. With the limited number of samples analyzed, the data suggested that long-standing scar tissues reverted to a composition similar to that of normal skin. The composition of hydroxylysine-linked carbohydrate units is also discussed on the basis of the age-related change.