ABSTRACT: Sulfated epitopes of alpha-glucosamine (GlcN sulfoforms) were prepared by solid-phase synthesis as models of internal glucosamines within heparan sulfate. An orthogonally protected 2'-hydroxyethyl GlcN derivative was immobilized on a trityl resin support and subjected to regioselective deprotection and sulfonation conditions, which were optimized with the aid of on-resin infrared or Raman analysis. The sulfoforms were cleaved from the resin under mild Lewis acid conditions without affecting the O- or N-sulfate groups and purified by reversed-phase high-performance liquid chromatography (HPLC). The alpha-GlcN sulfoforms and their 4- O-benzyl ethers were examined by electrospray ionization tandem mass spectrometry (ESI-MS/MS), with product ion spectra produced by collision-induced dissociation (CID). ESI-MS/MS revealed significant differences in parent ion stabilities and fragmentation rates as a function of sulfate position. Ion fragmentation by CID resulted in characteristic mass losses with strong correlation to the positions of both free hydroxyl groups and sulfate ions. Most of these fragmentation patterns are consonant with elimination pathways, and suggest possible strategies for elucidating the structures of glucosamine-derived sulfoforms with identical m/ z ratios. In particular, fragmentation analysis can easily distinguish GlcN sulfoforms bearing the relatively rare 3- O-sulfate from isomers with the more common 6- O-sulfate.
The Journal of Organic Chemistry 08/2008; 73(16):6059-72. · 4.45 Impact Factor