DETERMINATION OF THE MOLECULAR-WEIGHT DISTRIBUTION OF LOW-MOLECULAR-WEIGHT HEPARINS USING HIGH-PERFORMANCE GEL-PERMEATION CHROMATOGRAPHY

The molecular-weight distribution of commercially available low-molecular-weight heparins (LMWHs) was estimated by high-performance gel permeation chromatography (HPGPC) using a LMWH molecular-weight calibrant, supplied by the National Institute for Biological Standards and Control for the calibration of columns. This calibrant is a mixture of many polysaccharides with a disaccharide unit prepared by a random depolymerization of unfractionated heparin, and the ends of the fragments have a UV absorption. The ratio of the refractive index to the UV absorption is a relative measure of molecular weight. Various molecular-weight parameters such as weight-, number- or z-average molecular-weights, polydispersity and relative distributions of high, middle and low molecular-weight components were estimated, indicating that LMWHs have a wide variation in molecular-weight distribution. This variation was ascribed to the difference in preparation methods from unfractionated heparins.