Elsevier

Carbohydrate Research

Volume 345, Issue 9, 16 June 2010, Pages 1123-1134
Carbohydrate Research

Synthesis of α- and β-d-glucopyranosyl triazoles by CuAAC ‘click chemistry’: reactant tolerance, reaction rate, product structure and glucosidase inhibitory properties

https://doi.org/10.1016/j.carres.2010.03.041Get rights and content

Abstract

CuI-catalysed azide alkyne 1,3-dipolar cycloaddition (CuAAC) ‘click chemistry’ was used to assemble a library of 21 α-d- and β-d-glucopyranosyl triazoles, which were assessed as potential glycosidase inhibitors. In the course of this work, different reactivities of isomeric α- and β-glucopyranosyl azides under CuAAC conditions were noted. This difference was further investigated using competition reactions and rationalised on the basis of X-ray crystallographic data, which revealed significant differences in bond lengths within the azido groups of the α- and β-anomers. Structural studies also revealed a preference for perpendicular orientation of the sugar and triazole rings in both the α- and β-glucosyl triazoles in the solid state. The triazole library was assayed for inhibition of sweet almond β-glucosidase (GH1) and yeast α-glucosidase (GH13), which led to the identification of a set of glucosidase inhibitors effective in the 100 μM range. The preference for inhibition of one enzyme over the other proved to be dependent on the anomeric configuration of the inhibitor, as expected.

Introduction

Glycoside hydrolases are involved in a large number of natural processes related to carbohydrate metabolism. Detailed knowledge of the mechanisms1 that these enzyme use to hydrolyse glycosidic bonds of complex carbohydrates will facilitate the search for new inhibitors,2, 3 which is directly relevant to the discovery of new therapeutics.4 During the last few years there has been increasing interest in the use of the CuI-catalysed azide-alkyne cycloaddition reaction (CuAAC), an example of so-called ‘click chemistry’,5, 6 in the field of carbohydrate research.7, 8 This reaction has been used to assemble mimics of oligosaccharides and glycopeptides,7, 8, 9, 10, 11, 12 as well as glycoclusters and dendrimers;13, 14 it has also found applications in in vivo imaging of glycoconjugates.15, 16 Several attempts have been made to utilise click chemistry for the synthesis of potential glycosidase inhibitors.17, 18, 19 For instance, β-linked 1-glycosyl-4-phenyltriazoles 1 and 2 (Fig. 1), prepared from β-glycosyl azides and phenylacetylene, were assessed for inhibitory activity against three different glycosidases.18 Compound 1 displayed 48% inhibition of the bovine liver galactosidase activity at 0.24 mM concentration, but in general the inhibitory activity of compounds 1 and 2 was weak. A series of acarbose-like pseudo-oligosaccharides, for example triazole derivative 3, was assayed against α- and β-glycosidases from various sources and showed only weak inhibitory activity.17 Compounds 4a4c were synthesised19 using ‘click chemistry’ in order to combine two distinct classes of therapeutic agents—iminosugars and aryl-triazoles—which can interfere with both glycosidases and methionine aminopeptidase involved in angiogenesis.20, 21

The approach adopted in the present work was based on a simple assumption that new glucosidase inhibitors can be identified by screening libraries of glucosides having variable aglycones.22, 23, 24, 25, 26, 27 Utilisation of ‘click chemistry’ for building libraries of this type should allow one to overcome common synthetic challenges associated with the preparation of O- or C-glycosides and make it possible to incorporate a wide variety of functional groups into the aglycone portion of the glucopyranoside analogues. In this study we have used the CuAAC reaction of glucopyranosyl azide building blocks, both acetylated and deprotected, with assorted commercially available alkynes possessing lipophilic, hydrophilic, acidic or basic functionalities. The presence of these functional groups provides an opportunity to probe interactions of putative inhibitors with the active site while the relevance of the anomeric configuration can be explored using compounds synthesised by CuAAC of the same set of alkynes and either α- or β-glucopyranosyl azide. A small library of prospective glucopyranosyl triazole-based glycosidase inhibitors assembled using the CuAAC reaction (Fig. 2) was screened against the cognate and non-cognate GH1 sweet almond β-glucosidase and GH13 yeast α-glucosidase, which are readily available and well-characterised model glucosidases. The different reactivities of α- and β-glucopyranosyl azides towards the CuAAC reaction, which was noted in recent work from this group,28 are also discussed in more detail.

Section snippets

Synthesis of glucopyranosyl triazoles

Since the discovery of the CuAAC reaction29, 30 a large number of different reaction conditions and various forms of Cu(I) catalyst have been reported.6 For our purpose, we adopted a method based on the procedure first described by Sharpless30 that involves 0.25–0.5 M reactants, 0.01 mol equiv of CuSO4 plus 0.1 mol equiv of sodium ascorbate (NaAsc) as a catalyst, in 1:1 t-BuOH/H2O solvent mixture at room temperature. Testing these conditions for coupling between peracetylated β-d-glucopyranosyl

General methods

Microwave-assisted syntheses were conducted in Biotage Initiator system. Thin-layer chromatography (TLC) was performed on aluminium-backed, pre-coated silica gel plates (Silica Gel 60 F254, Merck). Spots were detected by immersion in a 5% ethanolic H2SO4, followed by heating to 200 °C. Column chromatography was performed on a Biotage Horizon purification system using pre-packed silica gel cartridges and gradient elution. Evaporation of solvents was performed under reduced pressure at 25–40 °C.

Acknowledgements

This work was supported by the UK BBSRC and the University of East Anglia.

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