A New Method of Carbohydrate Synthesis in Both Solution and Solid Phases Using a Special Hydroxy Protecting Group
Authors: Komba Shiro , Kitaoka Motomitsu , Kasumi Takafumi 
Published: November 2005

A new carbohydrate synthesis method using a special hydroxy protecting group (uni‐chemo protection = UCP) in both solution and solid phases was developed. The UCP group was comprised of polymerized amino acid derivatives. Each hydroxyl group was protected by a UCP group with a different degree of polymerization, which allowed them to be uniquely identified. To deprotect the UCP group, one cycle of Edman degradation was performed as follows: 1) removal of the amino protecting group; 2) phenyl isothiocyanate coupling; 3) removal of the N‐terminal phenyl thiocarbamoyl mono‐amino acid derivative by treatment with trifluoroacetic acid; and 4) re‐protection of the newly exposed amino group with a Boc group. The hydroxyl groups were deprotected successively from the UCP group with the lowest to the highest degree of polymerization by repeating this Edman degradation cycle. First, commercially available N‐α‐t‐Boc‐sarcosine, N‐α‐t‐Boc‐N‐α‐methyl‐L‐alanine, and N‐α‐t‐Boc‐L‐phenylglycine were examined as UCP groups. Despite successfully protecting and selectively deprotecting the hydroxyl groups, there were problems with stability or reactivity. To address these problems, N‐α‐1‐ethylpropylglycine was chosen as the UCP group, and we successfully synthesized two sialyl‐T antigen analogues. Tri‐UCP and mono‐UCP were attached to the 6‐ and 3‐positions of the D‐galactosamine (GalN) derivative, respectively, using cyanuric chloride. To selectively deprotect the mono‐UCP group on the 3‐position of the GalN derivative, one cycle of Edman degradation was performed. As a result of this cycle, the tri‐UCP on the 6‐position of the GalN derivative was degraded to di‐UCP, and the mono‐UCP on the 3‐position was selectively deprotected to yield a GalN derivative with a free 3‐position. Glycosylation of this selectively deprotected free 3‐position GalN derivative with a suitably protected D‐galactose (Gal) derivative in which the 3‐position was protected by a mono‐UCP group using N‐iodosuccinimide (NIS) and trifluoromethanesulfonic acid (TfOH) in dichloromethane yielded the desired disaccharide in high yield in both a position‐ and stereoselective manner. By repeating one cycle of Edman degradation, the 3′‐position‐free disaccharide in which the 6‐position was protected by a mono‐UCP group was prepared selectively. Subsequent acetylation and a final cycle of Edman degradation (except the third and fourth steps) yielded the 6‐position‐free disaccharide selectively. Two disaccharides, one with a free 3′‐position and another with a free 6‐position, were coupled with a sialic acid donor using NIS and TfOH in acetonitrile to obtain sialyl (2→3) T and sialyl (2→6) T antigen derivatives, respectively. Solid‐phase synthesis was demonstrated using polystyrene‐type beads and a new linker, 2‐{4‐(hydroxymethyl)benzamido}acetic acid (HMBA‐Gly), to synthesize Galβ (1→3) Gal from a suitably protected Gal derivative in which the 3‐position was protected by mono‐UCP. Solid‐phase glycosylation was successfully monitored by measuring the removal of the Fmoc group, which protected the amino group on UCP, similar to the method for monitoring solid‐phase Fmoc peptide synthesis. This UCP hydroxy protecting group was suitable for solid‐phase synthesis, and will be the key technique in both automated oligosaccharide synthesis and oligosaccharide library synthesis. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)