Determination of hydroxyproline by high pressure liquid chromatography.
A rapid, precise, and simple HPLC method provides an assay of hydroxyproline from tissue extracts or solutions of collagen. Samples are hydrolyzed with 6 N HCl, derivatized with phenyl isothiocyanate, and chromatographed on a small, C18 reverse-phase HPLC column. Hydroxyproline (Hyp) is separated from other amino acids and detected by absorption at 254 nm. The method detects 0.40 to 36 micrograms of Hyp with a linear response. Separation requires a total of 6 min, including column cleanup and reequilibration. All components are commercially available, making this a convenient method for routine measurement of collagen concentration.
Green
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Abbott Biotech, Inc., Needham Heights, Massachusetts 02194.
Reagan
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D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003124:Colorimetry; D006909:Hydroxyproline / Q000032:analysis*; D013997:Time Factors
D006909:Hydroxyproline