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Determination of elastase activity by reversed-phase high-performance liquid chromatography

Authors
Journal
Journal of Chromatography A
0021-9673
Publisher
Elsevier
Publication Date
Volume
461
Identifiers
DOI: 10.1016/s0021-9673(00)94292-4
Disciplines
  • Medicine

Abstract

Abstract The action of elastase on elastin was measured by high-performance liquid chromatography with on-line post-column derivatization. After alkaline hydrolysis, a 20-ml sample was injected into a ODS-2 gel column. An elastin-specific dipeptide, Val-Pro, and the internal standard, Gly-Leu, were eluted by a linear gradient of 0 to 10% of 1-propanol in 50 m M heptafluorobutyrate (pH 3) at a flow-rate of 1 ml/min. The eluent was reacted with fluorescamine, and the fluorescent products were measured. Retention times for Val-Pro and Gly-Leu were 17.33 and 23.54 min. The peak areas of 0.2–16 μg of Val-Pro gave a straight-line plot. Elastase activity was constant from 6 to 24 h and was 0.95 ± 0.02 (S.D.) μg/h. The method may be useful for the measurement of the elastolytic activity in some diseased tissues.

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