Structural Analysis of Heparin by Raman Spectroscopy

The Raman spectra of commercially available heparin disaccharide standards exhibit bands associated with the N-sulfate and the 6-O-sulfate groups of the glucosamine and the 2-O-sulfate of the iduronic acid. The N-sulfate has a strong band at 1039 cm-1. The 6-O-sulfate and the 2-O-sulfate exhibit bands at 1055 and 1065 cm-1, respectively. The pattern of these modes, which are assigned to the symmetric SO3 vibrations, was supported by semiempirical quantum mechanical calculations. The above bands were identified in the Raman spectrum of a commercial preparation of porcine mucosal heparin and were used to determine the relative proportion of the N-sulfate, 6-O-sulfate, and 2-O-sulfate groups in the heparin molecule. This information, which is complementary to that obtained by NMR spectroscopy, is of particular importance in relation to biological activity. This study also extends the usefulness of Raman spectroscopy to include structural details required for the quality assurance of pharmaceutical preparations of heparin.