Glucopyranosyl Fluoride Research Articles

The interaction of 1-fluoro-d-glucopyranosyl fluoride with glucosidases

1. 1-Fluoro-D-glucopyranosyl fluoride undergoes pH-independent loss of F- ion at a rate of 1 x 10(-8) s-1 at 50.0 degrees C, some 10(3)-fold slower than alpha-D-glucopyranosyl fluoride and 4 x 10(4)-fold slower than beta-D-glucopyranosyl fluoride. 2. The (inverting) amyloglucosidase II of Aspergillus niger hydrolyses the difluoride according to Michaelis-Menten kinetics (Km 34 mM and kcat. 0.27 s-1), by apparently the same (simple) mechanism by which it hydrolyses alpha-D-glucopyranosyl fluoride (Km 38 mM and kcat. 730 s-1), rather than by the Hehre resynthesis-hydrolysis mechanism used to transform beta-D-glucopyranosyl fluoride. 3. The difluoride is also a substrate for the (inverting) trehalase of pig kidney [Km 17.3 mM and Vmax. 6.2 x 10(-4) relative to alpha-D-glucopyranosyl fluoride (Km 38 mM]). 4. The quantitatively similar effect of fluorine substitution on the one-step enzymic reactions and on the non-enzymic reactions suggests that they go through similar (oxocarbonium-ion-like) transition states. 5. The difluoride is a substrate for the (retaining) beta-glucosidases from Aspergillus wentii (A3 enzyme) and sweet-almond meal (B isoenzyme) and for the retaining alpha-glucosidase from rice: comparison with the appropriate monofluoride reveals a variable rate-retarding effect of the second fluorine atom on kcat./Km that correlates with other measures of oxocarbonium ion character in the transition state. 6. The difluoride is a substrate for the (retaining) alpha-glucosidase from yeast, but also gives an insidious mimicry of active-site-directed irreversible inhibition, which we tentatively attribute either to formation of the non-covalent complex or to the fluoroglucosyl-enzyme increasing the well-known tendency of this enzyme to come out of solution by adsorption on the walls of the vessel.

Hydrogen fluoride saccharification of wood: Lignin fluoride content, isolation of α‐D‐glucopyranosyl fluoride and posthydrolysis of reversion products

Wood chips from bigtooth aspen (Populus grandidentata Michx.) were saccharified by reaction with liquid hydrogen fluoride either anhydrous or containing up to 10% v/v water. The reaction products were separated into a solid lignin fraction and a water-soluble saccharide fraction. The fluoride content of the lignin (determined after alkaline fusion) was initially about 1 mg/g wood, but was lowered to 0.1 mg/g wood by grinding and washing. Thus little or no chemical binding of fluoride to lignin occurred during hydrogen fluoride (HF) solvolysis. Analysis of the water-soluble fraction by gel filtration on Biogel P2 columns showed a range of low-molecular-weight oligosaccharides and only 10-20% sugar monomers. Thus considerable reversion occurred during HF evacuation. Posthydrolysis conditions were optimized for these reversion products by varying temperature and acid concentration. Optimal conditions at 1 h were 140 degrees C with 100mN sulfuric acid or 225mN Hydrofluoric acid resulting in monomer yields of > 90% for 0.5% sugar solutions and > 80% for 10% sugar solutions. After reaction of pure cellulose (Filter paper) with hydrogen fluoride in the absence of water, and terminating the reaction with calcium carbonate, the reaction intermediate alpha-D-glucopyranosylfluoride was isolated with a maximal yield of 0.2 g/g paper. Upon purification via paper chromatography glucosylfluoride was identified by its specific rotation and also by gas chromatography-mass spectrometry of its tetra-O-trimethylsilyl derivative.