MOLECULAR-MASS DETERMINATION OF LOW-MOLECULAR-MASS HEPARINS - APPLICATION OF WIDE COLLECTION ANGLE MEASUREMENTS OF LIGHT-SCATTERING USING AHIGH-PERFORMANCE GEL-PERMEATION CHROMATOGRAPHIC SYSTEM EQUIPPED WITH A LOW-ANGLE LASER LIGHT-SCATTERING PHOTOMETER

A high-performance gel permeation chromatographic system with on-line low-angle laser light-scattering detection (HPGPC-LALLS) was used to d etermine molecular masses of low-molecular-mass heparins (LMMHs). Meas urements at wide and narrow collection angles were compared and the ap plication of the HPGPC-LALLS method to small molecules, with molecular masses in the range 1000-10 000, was assessed. The molecular mass ave rages of fractionated heparins and commercially available LMMHs were a lso determined by ordinary HPGPC analysis using an LMMH molecular-mass calibrant, supplied by the National Institute for Biological Standard s and Control for the calibration of columns. The LALLS intensity at t he routinely used narrow collection angle (theta(col) = 1-degree) abou t doubled at the wide collection angle (theta(col) = 2-degrees) and th e signal-to-noise ratio was improved. The present study thus indicates that wide collection angle measurement of light scattering allows the application of the HPGPC-LALLS method to very small biopolymers of mo lecular mass <10000.