Effect of molecular structure on thermodynamic properties of carbohydrates. A calorimetric study of aqueous di- and oligosaccharides at subzero temperatures

For aqueous solutions of di- and oligosaccharides thermodynamic properties have been investigated at subzero temperatures using differential scanning calorimetry. The amount of unfrozen water observed is found to increase linearly with the glass transition temperatures of anhydrous carbohydrates. Furthermore, the amount of unfrozen water shows a linear relationship with known solution properties of aqueous carbohydrates, such as partial molar compressibility and heat of solution. The different effectiveness among various di- and oligosaccharides to avoid ice formation is associated with the combination of constitutive monosaccharides and attendant molecular structure features including the position and type of the glycosidic linkage between the constituent units. More unfrozen water is induced in the presence of a carbohydrate having a poorer compatibility with the three-dimensional hydrogen-bond network of water. A series of these results obtained imply that there is a common key of carbohydrate stereochemistry governing several different thermodynamic amounts of a given system involving carbohydrates. In this context, a modified stereospecific-hydration model can be used to interpret the present results in terms of stereochemical effects of carbohydrates.     

Fluorometric determination of carbohydrate with 2-aminothiophenol

The 2-aminothiophenol-based fluorometric assay of Nakano et al. (1973, J. Pharm. Soc. Jpn. 93, 350-353) for monosaccharides has been modified to improve the speed, applicability, and sensitivity of the method. The improved assay is applicable to complex carbohydrates as well as to monosaccharides. Less than 50 ng of carbohydrate in a final volume of 2 ml can be quantitatively measured within 30 min. The assay is reasonably compatible with the presence of a variety of reagents commonly used in aqueous buffer solutions. The assay is especially useful for monitoring column eluents during the purification of small quantities of carbohydrates or their conjugates.     

Hydration of oligosaccharides: anomalous hydration ability of trehalose.

The disaccharide trehalose extensively exists in anhydrobiotic organism and is considered to play an important role in preserving the integrity of biomembrane. However, the preserving mechanism remains unclear. In this report, we examine the hydration abilities of trehalose and several oligosaccharides composed of alpha-D-glucopyranosyl residues. The unfrozen water fraction per molecule was determined from differential scanning calorimetry measurements of their aqueous solutions. Further, the NMR relaxation time of the natural abundance 17O of water is measured for several saccharide solutions. These results indicate that trehalose has the highest hydration ability among the saccharides studied. In other words, trehalose can effectively lower the mobility of water molecules hydrogen-bonded with saccharides. It is thus reasonable that, among the disaccharides studied, trehalose exhibits the maximum stabilizing effect on the bilayer structure of lipid whose acyl chains are bonded with each other by the apolar interaction, because the apolar interaction is strengthened with the stabilization of the surrounding water structure.     

Calorimetric and molecular simulation study on unfrozen water characteristics in aqueous sugar solutions: implications for biopreservation

Intensive studies have been conducted to determine the protective mechanisms of sugars that have proven beneficial to the biopreservation application. However, little has been known about the unfrozen water content that aqueous sugar solutions can possess when frozen at cryogenic temperatures. This study conducted calorimetric measurements to determine the unfrozen water content in frozen aqueous solutions of glucose, fructose, sucrose and trehalose of multiple concentrations. The hydrogen-bonding network in these solutions was characterised by molecular simulations. The experimental results showed that more water could be prevented from ice crystallisation in a more concentrated solution. Disaccharides, especially trehalose, are more effective than other protectants (e.g., glucose, glycerol and dimethyl sulfoxide) for detaining water in the unfrozen state. Moreover, it was found that, at molecular levels, there were more hydrogen bonds between sugar and water molecules in a more concentrated solution. From both macro- and microscopic perspectives, trehalose was demonstrated to be a much more effective cryoprotectant than others. This comparative study proved that the unfrozen water should be mainly attributed to hydrogen bonds between sugar and water in the mixture. Our findings will provide valuable information for determining the physical state of cryopreserved biomatrix and guiding the preparation of protective formulations.     

Thermodynamics of the interaction of RbCl with some monosaccharides (D-glucose, D-galactose, D-xylose, and D-arabinose) in aqueous solutions at 298.15K

The Gibbs energy interaction parameters of RbCl with some monosaccharides (D-glucose, D-galactose, D-xylose, and D-arabinose) in water, g(ES), were obtained from electromotive force (emf) measurements of the electrochemical cell without liquid junction and containing two ion-selective electrodes (ISE): K-ISEmid R:RbCl(m(E))mid R:ISE-Cl and K-ISEmid R:RbCl(m(E)),saccharide (m(S))mid R:ISE-Cl, at 298.15K. The enthalpy interaction parameters of RbCl with these monosaccharides in water, h(ES), are determined according to the McMillan-Mayer theory from the measurements of the enthalpies of mixing of aqueous RbCl solutions with aqueous monosaccharide solutions, as well as the enthalpies of dilution of RbCl and monosaccharide solutions in pure water at 298.15K by a calorimetric method. Furthermore, the entropy interaction parameters, s(ES), can be evaluated through g(ES) and h(ES). The results suggest that the electrostatic interactions of these monosaccharides with RbCl in water are predominant compared with structural interactions, and these parameters are controlled primarily by the stereochemical structure of the monosaccharides in water.     

Vibrational circular dichroism of carbohydrate films formed from aqueous solutions

Vibrational circular dichroism (VCD) spectra in the entire 2000-900 cm(-1) region have been recorded, for the first time, for films of carbohydrates prepared from aqueous solutions. Eight different carbohydrates, alpha-D-glucopyranosyl-(1-->4)-D-glucose, cyclomaltohexaose, alpha-D-glucopyranosyl alpha-D-glucopyranoside, beta-D-glucopyranosyl-(1-->6)-D-glucose, beta-D-glucopyranosyl-(1-->4)-D-glucose, D-glucose, and both enantiomers of 6-deoxygalactose and of allose, were investigated. The VCD spectra obtained for films are found to be identical to the corresponding spectra obtained for aqueous solutions of carbohydrates. These measurements demonstrate several advantages of significant importance. The strong infrared absorption of water has prevented, in the past, the pursuit for routine applications of VCD in determining the structures of carbohydrates in aqueous solutions. This limitation is not present for film studies because water solvent is removed in the process of preparing the films. Also, strong infrared absorption of water at 1650 cm(-1) requires the use of very short-pathlength (6 microm) cells for measurements on aqueous solutions. This requirement and concomitant inconveniences (such as laborious assembling of a demountable liquid cell or purchasing an expensive variable pathlength liquid cell) have been eliminated for film measurements. The removal of interfering water absorption in film studies resulted in higher light throughput and better signal-to-noise ratios for VCD measurements. Another point of significance is that the amount of carbohydrate sample required for VCD measurements on films is approximately one to two orders of magnitude smaller than that required for corresponding VCD measurements on aqueous solutions. Since carbohydrate samples can now be studied as films, VCD spectroscopy becomes much more broadly applicable for carbohydrates than previously believed. The present work, in combination with other film measurements in our laboratory, indicate that VCD studies on films can be used more generally, providing a convenient and powerful approach for probing structural information for biologically important compounds.     

Nuclear magnetic resonance studies of amphiphile hydration: Effects of cholesterol on phosphatidyl choline hydration

Water which remains unfrozen at ?25 °C in the presence of phosphatidyl choline (PC) gives rise to a proton magnetic resonance signal which can be used to measure the hydration of single-walled vesicles and multilamellar liposomes of PC. The proton magnetic resonance signal of the unfrozen water in these systems is strongly dependent upon the nature of the molecular domain in which the water is situated. For example, at cholesterol to PC molar ratios below 35 mol%, the vesicle hydration signal consists of a relatively narrow symmetric peak (line width, ~150 Hz). At higher molar ratios, however, rather broad asymmetric signals appear (line widths, ~300–1000 Hz) which indicate that when significant quantities of cholesterol are packed in the bilayer there must be regions in which there is a preferred direction for motion of the unfrozen water. It is possible to solubilize significant quantities of cholesterol by sonicating it in concentrated solutions of sodium dodecyl sulfate. Addition of cholesterol to PC vesicles via these sodium dodecyl sulfate-cholesterol complexes caused hydration changes in the PC, which, at high cholesterol to PC molar ratios, paralleled the effects of cholesterol on PC hydration in homogeneous vesicles in which the cholesterol and PC were simply cosonicated.     

Hydration dynamics of hyaluronan and dextran

Hyaluronan is a polysaccharide, which is ubiquitous in vertebrates and has been reported to be strongly hydrated in a biological environment. We study the hydration of hyaluronan in solution using the rotational dynamics of water as a probe. We measure these dynamics with polarization-resolved femtosecond-infrared and terahertz time-domain spectroscopies. Both experiments reveal that a subensemble of water molecules is slowed down in aqueous solutions of hyaluronan amounting to ～15 water molecules per disaccharide unit. This quantity is consistent with what would be expected for the first hydration shell. Comparison of these results to the water dynamics in aqueous dextran solution, a structurally similar polysaccharide, yields remarkably similar results. This suggests that the observed interaction with water is a common feature for hydrophilic polysaccharides and is not specific to hyaluronan.     

Thermodynamics of carbohydrate-lipid interactions.

Thermodynamic analyses of carbohydrate-lipid interactions were performed by investigating the effects of a series of carbohydrates, including monosaccharides, disaccharides, and trisaccharides, on the phase-transition properties of aqueous dispersions of 1,2-dipalmitoyl phosphatidylcholine (DPPC). The temperature of the lipid's main phase transition from the gel to liquid-crystalline phase is essentially unchanged in the presence of carbohydrate. The change in the free energy (delta G) of the transition is zero when a carbohydrate is added to aqueous dispersions of DPPC, while the enthalpy (delta H) and the entropy of the melting of DPPC are decreased. The thermodynamic information was used to examine carbohydrate-lipid interactions. Such interactions were elucidated according to our knowledge of the specific properties of carbohydrates in aqueous solutions and the previously proposed hydrophobic interaction involving hydrocarbon tails of the lipid in aqueous dispersions.     

Hydration of Cytidine, 2′-Deoxycytidine and their Phosphate Salts in the Aqueous Solutions. A Molecular Dynamics Computer Simulation Study

Abstract

To compare the hydration pattern of the cytidine (Cyd) and 2′-deoxycytidine (dCyd) in the aqueous solutions at the level of microscopic interactions, Molecular Dynamics (MD) computer simulations have been undertaken. The results indicate that the hydration of the heterocyclic base moiety in cytidine and 2′-deoxycytidine has a hydrophobic character. None of the three potential Watson—;Crick base pair centres hydrogen bonds with the water molecules and the formation of something akin to a clathrate cage structure of water around base moieties of nucleosides in the aqueous solution is suggested. In contrast, the hydration of Cyd and dCyd sugar moieties shows a hydrophilic character and the three-dimensional networks of H-bonds involving all hydrophilic centres are formed differently around the ribose and 2′-deoxyribose. The sugar hydroxyl groups participate in the hydrogen bonding with water both as H-donor and as H-acceptor. Their donor-acceptor abilities have been evaluated and compared. The coordination numbers, the geometrical data of the first hydration shell, and the number of hydrogen bonds have been calculated. The changes in the pattern of hydration with the increased concentration of nucleosides and upon nucleoside protonation are discussed. The analysis of the pairwise interaction energies are also presented.     

Nuclear magnetic resonance studies of amphiphile hydration.

The nuclear magnetic resonance signal of water which remains unfrozen at ?25 °C in the presence of phosphatidylcholine has been used to determine the hydration of this amphiphile. The effects of cholesterol and sodium dodecylsulfate on both the area and linewidth of this signal indicate that these molecules cause significant changes in the structure of phosphatidylcholine vesicles in solution. Studies on other amphiphiles indicate that, whereas phosphatidylethanolamine has a hydration similar to phosphatidylcholine, species with just one hydrocarbon chain such as sodium dodecylsulfate and dodecyltrimethylammonium bromide have little, if any, hydration when assayed via the nuclear magnetic resonance experiment.     

Dielectric behavior of water in biological solutions: studies on myoglobin, human low-density lipoprotein, and polyvinylpyrrolidone

The dielectric behavior of the aqueous solutions of three widely differing macromolecules has been investigated: myoglobin, polyvinylpyrrolidone (PVP), and human serum low-density lipoprotein (LDL). It was not possible to interpret unambiguously the dielectric properties of the PVP solution in terms of water structure. The best interpretation of the dielectric data on the myoglobin and LDL solutions was that, in both cases, the macromolecule attracts a layer of water of hydration one or two water molecules in width. For LDL, this corresponds to a hydration factor of only 0.05 g/g, whereas for myoglobin the figure is nearer 0.6 g/g. With myoglobin, part of the water of hydration exhibits its dispersion at frequencies of a few GHz, and the rest disperses at lower frequencies, perhaps as low as 10-12 MHz. The approximate constancy of the width of the hydration shell for two molecules as dissimilar in size as LDL and myoglobin confirms that the proportion of water existing as water of hydration in a biological solution depends critically on the size of the macromolecules as well as on their concentration.     

Hyper-mobile water is induced around actin filaments

When introduced into water, some molecules and ions (solutes) enforce the hydrogen-bonded network of neighboring water molecules that are thus restrained from thermal motions and are less mobile than those in the bulk phase (structure-making or positive hydration effect), and other solutes cause the opposite effect (structure-breaking or negative hydration effect). Using a method of microwave dielectric spectroscopy recently developed to measure the rotational mobility (dielectric relaxation frequency) of water hydrating proteins and the volume of hydration shells, the hydration of actin filament (F-actin) has been studied. The results indicate that F-actin exhibits both the structure-making and structure-breaking effects. Thus, apart from the water molecules with lowered rotational mobility that make up a typical hydration shell, there are other water molecules around the F-actin which have a much higher mobility than that of bulk water. No such dual hydration has been observed for myoglobin studied as the representative example of globular proteins which all showed qualitatively similar dielectric spectra. The volume fraction of the mobilized (hyper-mobile) water is roughly equal to that of the restrained water, which is two-thirds of the molecular volume of G-actin in size. The dielectric spectra of aqueous solutions of urea and potassium-halide salts have also been studied. The results suggest that urea and I(-) induce the hyper-mobile states of water, which is consistent with their well-known structure-breaking effect. The molecular surface of actin is rich in negative charges, which along with its filamentous structure provides a structural basis for the induction of a hyper-mobile state of water. A possible implication of the findings of the present study is discussed in relation to the chemomechanical energy transduction through interaction with myosin in the presence of ATP.     

A Langmuir film balance study of the interactions of ionic and polar solutes with glycolipid monolayers

Using a Langmuir film balance experiments have been conducted to discover if dissolved salts or carbohydrates interact with glycolipid monolayers. Two types of glycolipid were studied, simple glycosides made by ether linking monosaccharides to fatty alcohols and cerebrosides extracted from natural sources. It was found that salts or carbohydrates in the subphase expanded glycolipid monolayers. That is, a monolayer spread on a solution occupied a greater area at a given pressure than it would have spread on pure water. Of the carbohydrates galactose and glucose, galactose caused a markedly greater expansion of monolayers than glucose. However, the magnitude of the expansions measured for stearyl glucoside, mannoside and galactoside films on solutions of a particular sugar were not significantly different, demonstrating that this phenomenon is independent of the glycolipid sugar residue. As with carbohydrates, salts also have differing effects on glycolipid monolayers. Although the effect an individual ion has on a monolayer cannot be directly measured, comparisons between salts indicate that there is a correlation between the size of an ion and the extent of the monolayer expansion it causes. To explain these observations two different mechanisms are proposed. In the case of salts it is suggested that large ions which have a low charge density disrupt water structure in such a way that monolayers spread on the surface of their solutions are expanded. The ability of carbohydrates to expand monolayers is explained in terms of the carbohydrate replacing water molecules bound to the polar groups of the monolayer and in so doing increasing the effective area of the lipid molecules. It is suggested that the molecular mechanisms involved in the interactions of ions and carbohydrates with glycolipid monolayers may also operate in the interactions of glycolipids and glycoproteins with extracellular agents and surfaces.     

Counterion binding and hydration of hyaluronate and chondroitin in solution: An 17O, 23Na,and 25Mg nuclear-magnetic-relaxation study

Nuclear magnetic relaxation rates of H₂¹⁷O, ²³Na⁺, and ²⁵Mg²⁺ have been measured in aqueous hyaluronate solutions. The dependence on solution pH of the relaxation rates has been investigated, as well as the competition behavior of Na⁺ with Ca²⁺ and Mg²⁺. H₂¹⁷O and ²³Na⁺ relaxation rates in chondroitin and hyaluronate solutions have been compared in the interval, 2 ? pH ? 12.5. The ion binding of hyaluronate can be fully accounted for by Coulomb interactions, with no need to involve chemical specificity. The hydration is only slighly pH dependent, and is comparable in magnitude to hydration of synthetic polyelectrolytes and monosaccharides. Ion-binding and hydration properties of hyaluronate and chondroitin are quite similar, except at elevated pH. At alkaline pH, an increase in charge density with pH is seen in hyaluronate and, to a much lesser degree, in chondroitin, possibly due to the titration of hydroxy groups. H₂¹⁷O data indicate an alkali-induced transition in both glycosaminoglycans.     

Mid-infrared spectroscopic analysis of saccharides in aqueous solutions with sodium chloride

The infrared spectral characteristics of three different types of disaccharides (trehalose, maltose, and sucrose) and four different types of monosaccharides (glucose, mannose, galactose, and fructose) in aqueous solutions with sodium chloride (NaCl) were determined. The infrared spectra were obtained using the FT-IR/ATR method and the absorption intensities respected the interaction between the saccharide and water with NaCl were determined. This study also focused on not only the glycosidic linkage position and the constituent monosaccharides, but also the concentration of the saccharides and NaCl and found that they have a significant influence on the infrared spectroscopic characterization of the disaccharides in an aqueous solution with NaCl. The absorption intensities representing the interaction between a saccharide and water with NaCl were spectroscopically determined. Additionally, the applications of MIR spectroscopy to obtain information about saccharide–NaCl interactions in foods and biosystems were suggested.     

Effects of anion substitution on hydration behavior and water uptake of the red-spotted toad, Bufo punctatus: is there an anion paradox in amphibian skin?

Amphibians absorb water osmotically across their skins and rely on chemosensory information from the skin to assess the suitability of hydration sources. The time spent with skin in contact with a moist surface provides a quantitative measure of their ability to perceive the ionic and osmotic properties of aqueous solutions. Dehydrated toads given hyperosmotic (250 mM) solutions of NaCl or Na-gluconate showed significantly longer periods of hydration behavior on the gluconate solution, but they lost water osmotically when immersed in either solution. Similarly, dehydrated toads given 250 mM solutions of NaCl, Na-acetate, Na-phosphate or Na-gluconate showed a progressively greater length of hydration time on solutions with the larger mol. wt anions. These results are consistent with the chemosensory phenomenon previously described in mammalian tongue as 'anion paradox'. On dilute (50 mM) solutions of NaCl or Na-gluconate, the hydration time was not different between anions, despite toads gaining water more rapidly when immersed in dilute NaCl than in Na-gluconate solutions. The differing behavioral results with hyperosmotic and hypoosmotic salt solutions suggest that chemosensory transduction through toad skin involves both transcellular and paracellular pathways.     

Structure and dynamics of primary hydration shell of phosphatidylcholine bilayers at subzero temperatures.

Deuterium NMR relaxation and intensity measurements of the 2H-labeled H2O/dimyristoyl phosphatidylcholine bilayer were performed to understand the molecular origin of the freezing event of phospholipid headgroup and the structure and dynamics of unfrozen water molecules in the interbilayer space at subzero temperatures. The results suggest that about one to two water molecules associated with the phosphate group freeze during the freezing event of phospholipid headgroups, whereas about five to six waters near the trimethylammonium group behave as a water cluster and remain unfrozen at temperatures as low as -70 degrees C. In addition, temperature-dependent T1 and T2 relaxation times suggest that dynamic coupling occurs not only between the phosphate group and its bound water, but also between the methyl group and the adjacent water molecules. Based on these observations, the primary hydration shell of phosphatidylcholine headgroup at subzero temperatures is suggested to consist of two distinct regions: a clathrate-like water cluster, most likely a water pentamer, near the hydrophobic methyl group, and hydration water molecules associated with the phosphate group.     

Optimal conditions for 4-hydroxybenzoyl- and 2-furoylhydrazine as reagents for the determination of carbohydrates, including ketosamines

4- Hydroxybenzoylhydrazine ( PAHBAH ) reacts with glucose in hot aqueous solution when alkali exceeds aroylhydrazine concentration. The related 2- furoylhydrazine ( FAH ) reacts at lower alkali concentrations, making this an attractive alternative carbohydrate reagent since it is (unlike PAHBAH ) freely water soluble. FAH reacts with monosaccharides more slowly than does PAHBAH , giving about half the color. Its specificity and behavior with a bismuth catalyst parallel those of PAHBAH . Solutions which contain 0.05 mol/liter PAHBAH with 1.5 mmol/liter bismuth III (as tartrate complex) and 0.5 mol/liter sodium hydroxide, or 0.01 mol/liter FAH with 1.5 mmol/liter bismuth III and 0.1 mol/liter sodium hydroxide are sensitive reagents for quantitative analysis, giving stable colors with many carbohydrates within 10 min at 75 degrees C. Ketosamines react more rapidly than glucose at lower temperatures and undergo similar reactions in less alkaline solutions. At pH less than 8, the reaction is specific for these 1- aminohexoses , and FAH can be used as a reagent for their assay.