Anomeric Equilibrium Research Articles

Phosphate buffer-catalyzed kinetics of mutarotation of glucosamine investigated by NMR spectroscopy

Glucosamine (2-amino-2-deoxy-d-glucose, GlcN) is a naturally occurring amino monosaccharide that is essential for a variety of biological functions, it is mainly involved in the formation of polysaccharide structures. It was recently reported to enable the imaging of cancerous tumors as an exogenous contrast agent using the MRI technique of chemical exchange saturation transfer (CEST). In preparation for the clinical use of GlcN, its anomeric equilibrium and mutarotation rate constants were directly investigated in this study utilizing high resolution 1H and 13C NMR spectroscopy. The effects of GlcN concentration, temperature, pH and buffer on the mutarotation rate constant and mutarotation equilibrium were measured. The mutarotation rate constant increased markedly with increasing GlcN concentrations. The rate constant of mutarotation of GlcN at room temperature was 2.2 × 10−4 - 5.0 × 10−4 s−1 at concentrations of 0.02–0.5 M, corresponding to a time of 3.8–1.7 h to reach 95% equilibrium. The anomeric ratio was strongly pH-dependent. The influence of phosphate buffer on the apparent rate constant of GlcN mutarotation was investigated. For phosphate buffer saline values between 0 and 50 mM, there was a six-fold increase in rate at pH 7.0. The mutarotation rate constant rose rapidly with pH at a phosphate concentration of 50 mM: from 0.4 × 10−3 s−1 at pH 5.0 to 7.8 × 10−3 s−1 at pH 9.4, suggesting that the catalysis is due to the HPO42− and PO43− ions. These findings might help researchers design the experimental setting for employing GlcN for cancer detection using GlcN-CEST MRI.

Experimental and Computational Studies on Structure and Energetic Properties of Halogen Derivatives of 2-Deoxy-D-Glucose.

The results of structural studies on a series of halogen-substituted derivatives of 2-deoxy-D-glucose (2-DG) are reported. 2-DG is an inhibitor of glycolysis, a metabolic pathway crucial for cancer cell proliferation and viral replication in host cells, and interferes with D-glucose and D-mannose metabolism. Thus, 2-DG and its derivatives are considered as potential anticancer and antiviral drugs. X-ray crystallography shows that a halogen atom present at the C2 position in the pyranose ring does not significantly affect its conformation. However, it has a noticeable effect on the crystal structure. Fluorine derivatives exist as a dense 3D framework isostructural with the parent compound, while Cl- and I-derivatives form layered structures. Analysis of the Hirshfeld surface shows formation of hydrogen bonds involving the halogen, yet no indication for the existence of halogen bonds. Density functional theory (DFT) periodic calculations of cohesive and interaction energies (at the B3LYP level of theory) have supported these findings. NMR studies in the solution show that most of the compounds do not display significant differences in their anomeric equilibria, and that pyranose ring puckering is similar to the crystalline state. For 2-deoxy-2-fluoro-D-glucose (2-FG), electrostatic interaction energies between the ligand and protein for several existing structures of pyranose 2-oxidase were also computed. These interactions mostly involve acidic residues of the protein; single amino-acid substitutions have only a minor impact on binding. These studies provide a better understanding of the structural chemistry of halogen-substituted carbohydrates as well as their intermolecular interactions with proteins determining their distinct biological activity.

Synergistic effect between CaCl2 and γ-Al2O3 for furfural production by dehydration of hemicellulosic carbohydrates

γ-Al2O3 has successfully been used as solid acid catalyst for xylose dehydration in a water:toluene biphasic system. The addition of alkaline earth metal chlorides improved the partition coefficient, thus facilitating the extraction of furfural from the aqueous phase. In this sense, the best catalytic performance was attained by using CaCl2 and γ-Al2O3 with almost full xylose conversion and a furfural yield of 55% after 50 min, at 150 °C. A 1H NMR study has demonstrated that CaCl2 shifts the α-xylopyranose/β-xylopyranose anomeric equilibrium toward the α-form, more favorable for initiating the dehydration process, and γ-Al2O3 promotes the furfural production. γ-Al2O3 can be reused during ten catalytic cycles without any pretreatment. Finally, the synergistic effect between CaCl2 and γ-Al2O3 could also be inferred from the increase in furfural yield, until a value of 83% after 50 min at 175 °C, by using a hemicellulosic liquor obtained from olive stones.

3-O-Methyl-D-glucose mutarotation and proton exchange rates assessed by 13C, 1H NMR and by chemical exchange saturation transfer and spin lock measurements.

3-O-Methyl-D-glucose (3OMG) was recently suggested as an agent to image tumors using chemical exchange saturation transfer (CEST) MRI. To characterize the properties of 3OMG in solution, the anomeric equilibrium and the mutarotation rates of 3OMG were studied by 1H and 13C NMR. This information is essential in designing the in vivo CEST experiments. At room temperature, the ratio of α and β 3OMG anomers at equilibrium was 1:1.4, and the time to reach 95% equilibrium was 6h. The chemical exchange rates between the hydroxyl protons of 3OMG and water, measured by CEST and spin lock at pH 6.14 and a temperature of 4°C, were in the range of 360-670s-1.

N-(1-Deoxy-α-d-tagatopyranos-1-yl)-N-methylaniline (“d-Tagatose-N-methylaniline”)

Tagatosamines form in thermally-processed dairy products and contribute to the foods’ organoleptic and nutritional value. d-Tagatose-N-methylaniline (N-(1-deoxy-d-tagatos-1-yl)-N-methylaniline, 1-deoxy-1-(N-methylphenylamino)-d-tagatose) was synthesized from d-galactose via the Amadori rearrangement. In aqueous solution, it established an anomeric equilibrium consisting of 62.8% α-pyranose, 21.3% β-pyranose, 1.5% α-furanose, 8.1% β-furanose, and 6.2% acyclic keto tautomer. The crystalline α-pyranose anomer of d-tagatose-N-methylaniline adopted the 5C2 chair conformation. All hydroxyl and ring oxygen atoms and the amino nitrogen are involved in an extensive H-bonding network dominated by infinite homodromic chains. The Hirshfeld surface analysis suggests a significant contribution of non-polar intermolecular contacts to the crystal structure.

Anomeric proportions of d-glucopyranose at the equilibrium determined from 1H NMR spectra II. Effects of alkali metal chlorides, CaCl2 and BaCl2 on the anomeric equilibrium at 25.0 °C

The isomeric proportions of saccharides are important information to discuss the properties of the saccharides such as the solvation states, structures, functions and so on in aqueous solutions. However, the isomeric composition, which is one of the fundamental properties of saccharides in the aqueous solution, has not been fully explored experimentally. Successively to our previous study on the anomeric proportion of d-glucopyranose in the aqueous solution [J. Mol. Liq., 232 (2017) 408–415.], the salt effect on these anomeric proportions at 25.0°C were investigated by 1H NMR measurement for the cases of all alkali metal chlorides, CaCl2 and BaCl2. The experimental conditions were; the glucose concentration was from 0.1wt% to 1.0wt% and the salt molality was from 0.1molkg−1 to 0.5molkg−1. At first, the optimal experimental conditions were confirmed in terms of the irradiation power and the target frequency of the presaturation pulse in 1H NMR measurement. The 1H NMR spectra measured under the optimal conditions with adequate accumulation number allowed us to determine the anomeric proportions within the error of 0.0004. The anomeric proportions of α-glucopyranose in the presence of each salts were determined from peak areas corresponding to the anomeric hydrogens of α- and β- d-glucopyranoses. For all kinds of the salts, the anomeric proportion increased consistently and the slope of the proportion decreased gradually with increasing glucose concentration at a given molality of the salts. The anomeric proportions at the infinite dilution of glucose were determined from these concentration dependences of glucose, and then the Gibbs energies of reaction for the anomerization ΔrG° were estimated from these results. The effects of the salts on ΔrG° were divided into two groups: (1) ΔrG° increased when LiCl, NaCl and CaCl2 were added, and (2) ΔrG° decreased when KCl, RbCl, CsCl and BaCl2 were added. The plots of ΔrG° against the ionic radius of the cations clearly illustrated this result, that is, ΔrG° for these two groups fell on the quite different cation radius dependences. ΔrG° of the first group of the salts were greater than that of the salt free system. ΔrG° for each molality of the salts linearly decreased with the cation radius, and became ΔrG° of the salt free glucose aqueous solution around 0.17nm irrespectively to the salt molality. On the other hand, ΔrG° of the second group of the salts were less than that of the salt free system. ΔrG° for each molality of the salts linearly decreased with the cation radius, and became ΔrG° of the salt free solution around 0.08nm irrespectively to the salt molality.

Behaviour of lactose with the presence of lactic acid and Ca as affected by pH.

Contradictory statements about the effects of pH change on crystallisation behaviour of lactose exist in the literature. Considering the importance of addressing the processability issue of acid whey, a systematic study is required to establish lactose crystallisation behaviour in the presence of LA and Ca at concentrations present in real acid whey waste streams emphasising impact of pH. Structural modifications of lactose were evident at elevated, more neutral pH in the presence of 1% w/w LA and 0·12% w/w Ca. These structural changes led to changes in the anomeric equilibrium of lactose, which manipulated the water-lactose behaviour and increased the crystallinity. Therefore, altering pH to 6·5 may be the solution to proper industrial processing of acid whey, enhancing the ability of lactose to crystallise properly.

Thermal Behavior of d-Ribose Adsorbed on Silica: Effect of Inorganic Salt Coadsorption and Significance for Prebiotic Chemistry.

Understanding ribose reactivity is a crucial step in the "RNA world" scenario because this molecule is a component of all extant nucleotides that make up RNA. In solution, ribose is unstable and susceptible to thermal destruction. We examined how ribose behaves upon thermal activation when adsorbed on silica, either alone or with the coadsorption of inorganic salts (MgCl2 , CaCl2 , SrCl2 , CuCl2 , FeCl2 , FeCl3 , ZnCl2 ). A combination of 13 C NMR, in situ IR, and TGA analyses revealed a variety of phenomena. When adsorbed alone, ribose remains stable up to 150 °C, at which point ring opening is observed, together with minor oxidation to a lactone. All the metal salts studied showed specific interactions with ribose after dehydration, resulting in the formation of polydentate metal ion complexes. Anomeric equilibria were affected, generally favoring ribofuranoses. Zn2+ stabilized ribose up to higher temperatures than bare silica (180 to 200 °C). Most other cations had an adverse effect on ribose stability, with ring opening already upon drying at 70 °C. In addition, alkaline earth cations catalyzed the dehydration of ribose to furfural and, to variable degrees, its further decarbonylation to furan. Transition-metal ions with open d-shells took part in redox reactions with ribose, either as reagents or as catalysts. These results allow the likelihood of prebiotic chemistry scenarios to be evaluated, and may also be of interest for the valorization of biomass-derived carbohydrates by heterogeneous catalysis.

Hyperconjugative and Electrostatic Interactions as Anomeric Triggers in Archetypical 1,4-Dioxane Derivatives.

The anomeric effect accounts for the greater thermodynamic stability of axially arranged six-membered heterocycles holding an electronegative substituent at the C1 position. Within a frame of no general consensus, two different theories are typically claimed to justify this effect mostly based on either hyperconjugative or electrostatic factors. Here we report a theoretical-experimental study of the role of both as anomeric triggers in two archetypical 1,4-dioxane derivatives, using a suitable combination of spectroscopic (IR and vibrational circular dichroism) and computational techniques for the analysis of the solvation environment effect in their anomeric choices. VCD and IR spectroscopies are used as conformer-discriminating tools: a detailed analysis of the evolution of the spectral profiles allows assessing the theoretically predicted changes in the experimental α/β ratios when changing the polar solvent, which are fully explained on the basis of an extensive NBO energy partition scheme that provides a detailed view of the role of hyperconjugative and electrostatic interactions as anomeric regulators. Our results suggest that the anomeric equilibrium cannot be described by a single stereoelectronic effect but by the combined contribution of hyperconjugation and electrostatic repulsions, so that the β-anomeric choice in polar solvents is markedly driven by the strong attenuation of electrostatic repulsive interactions that occurs in solution.

Cobalt(III) Complexes of D‐Galactosylamine

AbstractThe β‐pyranose isomer of D‐galactosylamine (1) formed complexes with three different cobalt(III) fragments. Crystals containing the dication [Co(tren)(β‐D‐Galp1N2H–1‐κ2N1,O2)]2+ (3) showed coordination through the anomeric amino group (N1) and the deprotonated hydroxy group (O2) of the 4C1 β‐pyranose form, which is also the major isomer of free galactosylamine. The cationic complexes [Co(fac‐dien)(β‐D‐Galp1N2H–1‐κ2N1,O2)]2+ (4) and [Co(phen)2(β‐D‐Galp1N2H–1‐κ2N1,O2)]2+ (5) were analysed by NMR spectroscopy and showed the same coordination mode as 3. In terms of available ligand isomers it was shown that 1 exhibits an anomeric equilibrium in solution of both pyranose and both furanose forms as is typical for the parent glycose, galactose.

Probing the influence of protecting groups on the anomeric equilibrium in sialic acid glycosides with the persistent radical effect.

A method for the investigation of the influence of protecting groups on the anomeric equilibrium in the sialic acid glycosides has been developed on the basis of the equilibration of O-sialyl hydroxylamines by reversible homolytic scission of the glycosidic bond following the dictates of the Fischer–Ingold persistent radical effect. It is found that a trans-fused 4O,5N-oxazolidinone group stabilizes the equatorial glycoside, i.e., reduces the anomeric effect, when compared to the 4O,5N-diacetyl protected systems. This effect is discussed in terms of the powerful electron-withdrawing nature of the oxazolidinone system, which in turn is a function of its strong dipole moment in the mean plane of the pyranose ring system. The new equilibration method displays a small solvent effect and is most pronounced in less polar media consistent with the anomeric effect in general. The unusual (for anomeric radicals) poor kinetic selectivity of anomeric sialyl radicals is discussed in terms of the planar π-type structure of these radicals and of competing 1,3-diaxial interactions in the diastereomeric transition states for trapping on the α- and β-faces of the radical.

Hydrolysis and Isomerization of Sugar Phosphates and Carbohydrate Phosphodiesters

Phosphoesters are abundant in carbohydrate structures, yet their chemical reactivity is less well known than that of nucleoside phosphoesters. Both classes of compounds contain sugar bound phosphoesters, but structural versatility of carbohydrates means that the reactivity range is wider, and reaction mechanisms not feasible in nucleic acid chemistry, are possible. Sugar phosphates, as well as their phosphodiester and phosphoanhydride derivatives with a phosphate group in glycosylic position, react like acetals under acidic conditions. Substrates with a phosphate group attached to an alcoholic OH react by intramolecular transesterification similar to that of RNA provided that there is a suitably positioned HO-group and a suitable leaving group. If there is a free carbonyl group allowing anomeric equilibria, base-catalyzed phosphate elimination through enediolate intermediates may compete with the cleavage, particularly under alkaline conditions. The few reports on phosphate migration show that the reaction is conceivable, but the competition between cleavage and phosphate migration possibly is different from reactions of nucleic acids and nucleotides.

Influence of protecting groups on the anomeric equilibrium; case of the 4,6-O-benzylidene acetal in the mannopyranose series

It is reported that the replacement of the 4- and 6-O-benzyl ethers in 2,3,4,6-tetra-O-benzyl-α,β-mannopyranose by a 4,6-O-benzylidene acetal results in an increased population of the β-anomer at equilibrium in CDCl3 solution. The phenomenon is considered to arise from the lower steric bulk of the benzylidene acetal that, through diminished buttressing interactions, reduces steric interactions normally present in the β-anomer.

Modelling the Effect of Solvents on Carbohydrates

Carbohydrates are polar molecules and their conformational and anomeric equilibrium can be strongly influenced by solvents. This review provides examples of studies addressing different issues of glycochemistry, such as anomeric equilibrium, conformational changes in rings, modelling of inter-residue linkages or complex carbohydrates and formation of carbohydrate-protein complexes. All these phenomena provide benchmark systems for testing of different solvent models.

PH-Dependent Mutarotation of 1-Thioaldoses in Water. Unexpected Behavior of (2S)-d-Aldopyranoses

The pH-dependent mutarotation of 1-thioaldopyranoses in aqueous media has been investigated. Anomerization readily occurred at lower and neutral pH for all aldopyranoses studied, whereas mainly for (2S)-D-aldopyranoses at higher pH. 1-Thio-D-mannopyranose and 1-thio-D-altropyranose showed very strong pH dependence where the anomeric equilibrium ratios changed dramatically from a preference for the β-anomer at lower pH to the α-anomer at higher pH.

Reference interaction site model study on the anomeric equilibrium of D-glucose in aqueous solution

The anomeric equilibrium of D-glucose in aqueous solution was studied by the extended reference interaction site model (XRISM) theory. In this study, all of the rotational degrees of freedom were considered upon the exocyclic hydroxyl and hydroxymethyl groups, namely 729 stereoisomers for each anomer. The free energy

Experimental and Predicted Results of Anomeric Equilibrium of Glucose in Alcohols

Total glucose solubility as well as equilibrium concentrations of α- and β-glucose anomers in 1-butanol, tert-butyl alcohol, 2-propanol, 1-propanol, ethanol, methanol, and an ethanol (1) + water (2) mixture (w1 = 90 %) at 40 °C and 60 °C were measured. Also, the recently developed mS-UNIFAC model is applied in the prediction of the total glucose solubility in these solvents and of the anomeric distribution in them yielding very satisfactory results.

Theoretical study of the relative stability of rotational conformers of alpha and beta-D-glucopyranose in gas phase and aqueous solution.

The alpha-beta anomer energy difference and the stability of 10 rotamers of counterclockwise D-glucopyranose were studied in vacuo and in aqueous solution at the B3LYP/6-31+G(d,p) level. To obtain the solute charge distribution and the solvent structure around it, we used the averaged solvent electrostatic potential from molecular dynamics method, ASEP/MD, which alternates molecular dynamics and quantum mechanics calculations in an iterative procedure. The main characteristics of the anomeric equilibrium, both in vacuo and in solution, are well reproduced. The relative stability of the different anomers is related to the availability of the free pairs of electrons in the anomeric oxygen to interact with the water molecules. The influence of solvation in the conformer equilibrium is also analyzed.

Metabolism of D-glucose anomers in rat pancreatic islets exposed to equilibrated D-glucose.

This study aims at establishing the contribution of alpha- and beta-D-glucose to the total generation of (3)HOH by rat pancreatic islets exposed to D-[2 - (3)H]glucose or D-[5 - (3)H] glucose at anomeric equilibrium. The islets were incubated for 60 min at 4 degrees C in the presence of equilibrated D-glucose (2.8 and 8.3 mM) mixed with tracer amounts of either alpha- or beta-D-glucose labelled with tritium on either the C (2) or C (5) of the hexose. Relative to their respective concentrations, (3)HOH generation from the anomers labelled with tritium on the C (2) or C (5) of the hexose provided beta/alpha ratios comparable to those previously found at both 2.8 and 8.3 mM, when the islets were exposed to each anomer separately. The relative contributions of each anomer to the total generation of (3)HOH was also close to the theoretical values derived from mathematical models for the catabolism of D-glucose at anomeric equilibrium in rat islets at both 2.8 and 8.3 mM and in the case of both D-[2 - (3)H]glucose and D-[5 - (3)H]glucose. Thus, even in islets exposed to D-glucose at anomeric equilibrium, the metabolic fate of alpha-D-glucose differs vastly from that of beta-D-glucose, the enzyme-to-enzyme channelling between hexokinase isoenzymes, especially glucokinase, and phosphoglucoisomerase being restricted to alpha-D-glucose 6-phosphate.

Metabolism of tritiated D-glucose anomers in rat erythrocytes.

It was recently proposed that alpha-D-glucose 6-phosphate may undergo enzyme-to-enzyme channelling between glucokinase and phosphoglucoisomerase in rat pancreatic islets. The present study aims at exploring whether a different situation prevails in cells deprived of glucokinase, namely in erythrocytes. At anomeric equilibrium, the ratio between D-[2-3H]glucose and D-[5-3H]glucose conversion to 3HOH was lower in rat erythrocytes incubated for 60 min at 4 degrees C in the presence of 2.8 mM, rather than 8.3 mM, D-glucose. This coincided with both a greater relative increase in beta-D-[5-3H]glucose, as compared to alpha-D-[5-3H]glucose, conversion to 3HOH and an increase in the beta/alpha ratio for 3HOH generation from D-[5-3H]glucose in response to an increase in the anomeric concentration from 2.8 to 8.3 mM, the suppression of the difference between the beta/alpha ratios for 3HOH generation from D-[2-3H]glucose and D-[5-3H]glucose in the erythrocytes incubated at 8.3 mM, as distinct from 2.8 mM, alpha- and beta-D-glucose, and a [2-3H]/[5-3H] ratio for 3HOH generation lower than unity in erythrocytes exposed to alpha-D-glucose but not significantly different from unity in the presence of beta-D-glucose. These findings emphasize the relevance of alpha-D-glucose 6-phosphate channelling between hexokinase and phosphoglucoisomerase as a determinant of the difference between D-[2-3H]glucose and D-[5-3H]glucose conversion to 3HOH, and reveal that the regulation of such a tunnelling process by the concentration of the D-glucose represents, in rat erythrocytes, a mirror image of that observed in rat pancreatic islets. The regulation of this process thus tightly depends on the identity of the hexokinase enzyme mainly responsible for the phosphorylation of D-glucose in distinct cell types.