Microscale preparation of even- and odd-numbered N-acetylheparosan oligosaccharides

In order to prepare a series of N-acetylheparosan (NAH)-related oligosaccharides, bacterial NAH produced in Escherichia coli strain K5 was partially depolymerized with heparitinase I into a mixture of even-numbered NAH oligosaccharides, having an unsaturated uronic acid (ΔUA) at the non-reducing end. A mixture of odd-numbered oligosaccharides was derived by removing this ΔUA in the aforementioned mixture by a 'trimming' reaction using mercury(II) acetate. Each oligosaccharide mixture was subjected to gel-filtration chromatography to generate a series of size-uniform NAH oligosaccharides of satisfactory purity (assessed by analytical anion-exchange HPLC), and their structures were identified by MALDITOF-MS, ESIMS, and 1H NMR analysis. As a result, a microscale preparation of a series of both even- and odd-numbered NAH oligosaccharides was achieved for the first time. The developed procedure is simple and systematic, and thus, should be valuable for providing not only research tools for heparin/heparan sulfate-specific enzymes and their binding proteins, but also precursor substrates with medical applications.