Effects of Addition of a Palmitic Sucrose Ester on Low-Trans-Fat Blends Crystallization in Bulk and in Oil-in-Water Emulsions

The effect of addition of a highly hydrophobic emulsifier, the palmitic sucrose ester P-170, on isothermal crystallization behavior of a high-melting fraction of milk fat (HMF) and its mixtures with 20% or 40% sunflower oil (SFO), both in bulk and in emulsion systems, were studied by nuclear magnetic resonance (NMR) and by time-resolved in situ small-angle synchrotron X-ray scattering (SAXS). NMR studies showed that the effect of P-170 on the overall isothermal crystallization kinetics in bulk phase could be either acceleration or delay. The effect was strongly dependent on supercooling. For HMF, P-170 retarded crystallization at temperatures above 29.0 ± 0.2 °C, while below that temperature, it accelerated the process. For the blends of HMF with 20% or 40% SFO, the temperature at which the behavior changed was 27.0 ± 0.2 or 26.0 ± 0.2 °C, respectively. In emulsion systems, however, the effect was always acceleration for all temperatures selected. With the aid of SAXS, it was possible to study early stage of crystallization during real time and therefore to improve our understanding of the mechanism of P-170 action. The different effects caused by P-170, as described by SFC curves, were strongly related to the effects of P-170 on fat polymorphism, specially to the value of the time interval of coexistence of the α and β′ forms. These events took place at the very beginning of crystallization and thus could not be described by the traditional X-ray techniques used in previous studies of similar systems.     

Freezing of phosphocholine headgroup in fully hydrated sphingomyelin bilayers and its effect on the dynamics of nonfreezable water at subzero temperatures.

Differential scanning calorimetry (DSC) and nuclear magnetic resonance (NMR) spectroscopy are applied to characterize the nonfreezable water molecules in fully hydrated D2O/sphingomyelin at temperatures below 0 degrees C. Upon cooling, DSC thermogram displays two thermal transitions peaked at -11 and -34 degrees C. The high-temperature exothermic transition corresponds to the freezing of the bulk D2O, and the low-temperature transition, which has not previously been reported, can be ascribed to the freezing of the phosphocholine headgroup in the lipid bilayer. The dynamics of nonfreezable water are also studied by 2H NMR T1 (spin-lattice relaxation time) and T2e (spin-spin relaxation time obtained by two pulse echo) measurements at 30.7 MHz and at temperatures down to -110 degrees C. The temperature dependence of the T1 relaxation time is characterized by a distinct minimum value of 2.1 +/- 0.1 ms at -30 degrees C. T2e is discontinuous at temperature around -70 degrees C, indicating another freezing-like event for the bound water at this temperature. Analysis of the relaxation data suggest that nonfreezable water undergoes both fast and slow motions at characteristic NMR time scales. The slow motions are affected when the lipid headgroup freezes.     

Freezing stresses and hydration of isolated cell walls

The hydration of the cell walls of the giant alga Chara australis was measured as a function of temperature using quantitative deuterium nuclear magnetic resonance (NMR) of samples hydrated with D2O. At temperatures 23-5K below freezing, the hydration ratio (the ratio of mass of unfrozen water in microscopic phases in the cell wall to the dry mass) increases slowly with increasing temperature from about 0.2 to 0.4. It then rises rapidly with temperature in the few Kelvin below the freezing temperature. The linewidth of the NMR signal varies approximately linearly with the reciprocal of the hydration ratio, and with the freezing point depression or water potential. These empirical relations may be useful in estimating cell wall water contents in heterogeneous samples.     

Root physiological factors involved in cool-season grass response to high soil temperature

High soil temperature is a critical factor limiting growth of cool-season grasses. This study was designed to examine changes in water, nutritional, and hormonal status in response to high soil temperature for creeping bentgrass (Agrostis stoloniferavar. palustris) and to compare the sensitivity of those parameters to high soil temperatures. Plants of ‘Penncross’ were exposed to 35 °C soil temperature (heat stress) or 20 °C (control) in water baths while air temperature was maintained at 20 °C in growth chambers. Turfgrass quality, shoot growth rate, and root biomass decreased below the control levels at 15, 15, and 10 days of heat stress, respectively, while root mortality increased above the control level at 5 days of heat stress. Relative water content (RWC) of leaves decreased below the control level at 15 days of heat treatment. Root N content increased while P and K content did not change over time at 35 °C. Shoot N, P, and K content decreased below the control level at 15, 15, and 10 days of heat stress, respectively. Root abscisic acid (ABA) content decreased below the control level at 10 days while shoot ABA content increased above the control level at 5 days. The content of cytokinins (zeatin (Z) and zeatin riboside (ZR), dihydrogen zeatin riboside (DHZR), and isopentenyl adenosine (iPA)) decreased below their respective control levels as early as 5 days of heat stress for roots and 10 days for shoots. The decline in cytokinin content was also more dramatic than changes in other parameters. Our results suggested that cytokinin was most sensitive to high soil temperature among parameters examined, suggesting that changes in cytokinins could serve as an early stress indicator for plant responses to high soil temperature; however, decreased water, nutrient (N, P, and K), and cytokinin content, and increased ABA could all contribute to the decline in shoot and root growth for creeping bentgrass exposed to high soil temperatures.     

Hemoglobin senses body temperature

When aspirating human red blood cells (RBCs) into 1.3 μm pipettes (ΔP = −2.3 kPa), a transition from blocking the pipette below a critical temperature T _c = 36.3 ± 0.3°C to passing it above the T _c occurred (micropipette passage transition). With a 1.1 μm pipette no passage was seen which enabled RBC volume measurements also above T _c. With increasing temperature RBCs lost volume significantly faster below than above a T _c = 36.4 ± 0.7 (volume transition). Colloid osmotic pressure (COP) measurements of RBCs in autologous plasma (25°C ≤ T ≤ 39.5°C) showed a T _c at 37.1 ± 0.2°C above which the COP rapidly decreased (COP transition). In NMR T₁-relaxation time measurements, the T₁ of RBCs in autologous plasma changed from a linear (r = 0.99) increment below T _c = 37 ± 1°C at a rate of 0.023 s/K into zero slope above T _c (RBC T₁ transition). In conclusion: An amorphous hemoglobin–water gel formed in the spherical trail, the residual partial sphere of the aspirated RBC. At T _c, a sudden fluidization of the gel occurs. All changes mentioned above happen at a distinct T _c close to body temperature. The T _c is moved +0.8°C to higher temperatures when a D₂O buffer is used. We suggest a mechanism similar to a “glass transition” or a “colloidal phase transition”. At T _c, the stabilizing Hb bound water molecules reach a threshold number enabling a partial Hb unfolding. Thus, Hb senses body temperature which must be inscribed in the primary structure of hemoglobin and possibly other proteins. This article is dedicated to Ludwig Artmann who died on July 21, 2001 on a beautiful summer day during which we performed experiments far away. Ludwig Artmann was a man who encouraged us to be strong and to study hard no matter what were the costs.     

Molecular dynamics simulations of a cyclic-DP-240 amylose fragment in a periodic cell: Glass transition temperature and water diffusion

Molecular dynamics simulations using AMB06C, an in-house carbohydrate force field, (NPT ensembles, 1 atm) were carried out on a periodic cell that contained a cyclic 240 glucose residue amylose fragment (c-DP-240) and TIP3P water molecules. Molecular conformation and movement of the amylose fragment and water molecules at different temperatures were examined. The periodic cell volume, density, and potential energy were determined at temperatures above and below the glass transition temperature (T_g) in 25 K increments_. The amorphous cell is constructed through successive dynamic equilibration steps at temperatures above the assumed T_g value and the temperature successively lowered until several temperature points were obtained below T_g. Molecular dynamics simulations were continued for at least 500 ps or until the volume drift stopped and remained constant for several hundred picoseconds. The T_g values were found by noting the discontinuity in slope of the volume (V), potential energy (PE), or density (ρ) versus 1/T. The changes in flexibility and motion of the amylose chain as well as differences in self diffusion coefficients of water molecules are described. The final average T_g value found (316 K) is in agreement with experimental values, i.e. 320 K.     

Physiological constraints on the life cycle and distribution of the Antarctic fairy shrimp <Emphasis Type="Italic">Branchinecta gaini</Emphasis>

Maturation to adulthood and successful reproduction in the Antarctic fairy shrimp, Branchinecta gaini, must be completed within a physiologically challenging temporal window of ca. 2.5 months in the southern Antarctic Peninsula. Although adults show considerable metabolic opportunism at positive temperatures, little is known of their tolerance of two physiological insults potentially typical to pool life in the maritime Antarctic: sub-zero temperatures and salinity. B. gaini are freeze-avoiding crustaceans with temperatures of crystallisation (T _cs) of −5°C. No antifreeze proteins were detected in the haemolymph. Adults osmoregulate in relation to temperature, but rapid mortality in saline solutions of even low concentration, indicate they cannot osmoregulate in relation to salinity. Survival of ice encasement at temperatures above their T _c was found to be pressure but not time dependent: at severe inoculative ice pressures, there was little immediate survival and none survived after 48 h below −2°C; at mild inoculative ice pressures, immediate survival was ca. 100% at −3°C, but <20% after 48 h. There was no significant difference in survival after 1 and 6 h encasement at −3°C. Observations of ventilation suggest that it is not low temperature per se, but ice that represents the primary cryo-stress, with ventilatory appendages physically handcuffed below the freezing point of pool water. Both sub-zero temperatures and salinity represent real physiological constraints on adult fairy shrimp.     

Water Confined in Cylindrical Pores: A Molecular Dynamics Study

Molecular dynamics simulations of water confined in two hydrophilic cylindrical pores—PH and PL—that differ in their silanol surface concentration (7.6 and 3.0 nm⁻², respectively) have been performed at 300 K. A strong interaction of interfacial water molecules with the pore was systematically found and gives rise to a layering effect, a significant distortion of both the hydrogen bond network (HBN) and the tetrahedral structure of these water molecules, and a corresponding subdiffusive mean square displacement along the main axis of the pores. By contrast, water molecules in the inner part of both pores were found to behave similarly to bulk water. The HBN and the tetrahedral configuration of water were more gradually distorted in the PL pore given the larger heterogeneity and rugosity of the surface, and the number of water–pore hydrogen bonds did not scale linearly with the silanol surface concentration of the pores, in part because of the close proximity between silanols in the PH pore. With the PL pore, the dynamic slowing down of water was found consistent with the experiment, suggesting that it provides a better model for the cylindrical MCM-41 mesopores. The structural and dynamical properties of water vary little with the silica force field.     

Relaxation Behavior of Collagen

The dynamic mechanical properties of purified collagen from bovine tendon were studied using a torsion pendulum in the temperature range of 120°-360°K at 0.3-1 cps. In the temperature range studied, two loss peaks were observed: a β-peak at about 200°K, and an α-peak approximately five times larger at about 280°K. The temperature of the α-transition is shown to be dependent on water content, decreasing with increasing amount of water and shifting to lower temperatures. Broad-line proton magnetic resonance results were also obtained on similar samples. A narrow nuclear magnetic resonance (NMR) line appears at about 250°C. The effects of shrinkage to form gelatin and of cross-linking on the relaxation behavior of collagen were also studied. The motions taking place in collagen over the 120°-360°K range are discussed.     

H NMR study of rehydration/dehydration and water mobility in β-cyclodextrin

The processes of dehydration and rehydration of β-cyclodextrin were studied by analysis of the ¹H NMR (nuclear magnetic resonance) line shape. Dehydration was carried in an open ampoule as a function of temperature and above 400 K total dehydration of β-cyclodextrin was observed. This result was confirmed by the thermogravimetry (TG) measurements. Rehydration was studied as a function of time at room temperature. After 40 days, β-cyclodextrin was found to absorb eight water molecules. The analysis of temperature changes in the shape of the ¹H NMR line of β-cyclodextrin kept in a closed ampoule and its dielectric measurements provided information on the mobility of water molecules. The water molecules were found to perform complex molecular motions, that is, reorientational jumps below 200 K and additionally, translational motion (diffusion) above 200 K.     

H and C NMR studies on the temperature-dependent water and protein dynamics in hydrated elastin,myoglobin and collagen

²H NMR spin-lattice relaxation and line-shape analyses are performed to study the temperature-dependent dynamics of water in the hydration shells of myoglobin, elastin, and collagen. The results show that the dynamical behaviors of the hydration waters are similar for these proteins when using comparable hydration levels of h = 0.25–0.43. Since water dynamics is characterized by strongly nonexponential correlation functions, we use a Cole–Cole spectral density for spin-lattice relaxation analysis, leading to correlation times, which are in nice agreement with results for the main dielectric relaxation process observed for various proteins in the literature. The temperature dependence can roughly be described by an Arrhenius law, with the possibility of a weak crossover in the vicinity of 220 K. Near ambient temperatures, the results substantially depend on the exact shape of the spectral density so that deviations from an Arrhenius behavior cannot be excluded in the high-temperature regime. However, for the studied proteins, the data give no evidence for the existence of a sharp fragile-to-strong transition reported for lysozyme at about 220 K. Line-shape analysis reveals that the mechanism for the rotational motion of hydration waters changes in the vicinity of 220 K. For myoglobin, we observe an isotropic motion at high temperatures and an anisotropic large-amplitude motion at low temperatures. Both mechanisms coexist in the vicinity of 220 K. ¹³C CP MAS spectra show that hydration results in enhanced elastin dynamics at ambient temperatures, where the enhancement varies among different amino acids. Upon cooling, the enhanced mobility decreases. Comparison of ²H and ¹³C NMR data reveals that the observed protein dynamics is slower than the water dynamics.     

The relation of the X-ray B-factor to protein dynamics: insights from recent dynamic solid-state NMR data

In addressing the potential use of B-factors derived from X-ray scattering data of proteins for the understanding the (functional) dynamics of proteins, we present a comparison of B-factors of five different proteins (SH3 domain, Crh, GB1, ubiquitin and thioredoxin) with data from recent solid-state nuclear magnetic resonance experiments reflecting true (rotational) dynamics on well-defined timescales. Apart from trivial correlations involving mobile loop regions and chain termini, we find no significant correlation of B-factors with the dynamic data on any of the investigated timescales, concluding that there is no unique and general correlation of B-factors with the internal reorientational dynamics of proteins.     

Leaf Extension in Zea mays: I. LEAF EXTENSION AND WATER POTENTIAL IN RELATION TO ROOT-ZONE AND AIR TEMPERATURES

The temperature of the roots and shoots of Zea mays plants werevaried independently of each other and the rates of leaf extensionand leaf water potentials were measured. Restrictions of leafextension occurred when root temperatures were lowered from35 to 0 °C, but leaf water potentials were lowered onlyat root temperatures below 5 °C. Similar changes in ratesof leaf extension were measured at air temperatures from 30to 5 °. Between 30 and 35 °C air temperature, in anunsaturated atmosphere, restrictions of leaf extension wereassociated with low leaf water potentials. It was concluded that, at root temperatures 5 to 35 °C,and shoot temperatures 5 to 30 °C, water stress was notthe main factor restricting the extension of Zea mays leaves.     

The relation of the X-ray B-factor to protein dynamics: insights from recent dynamic solid-state NMR data

In addressing the potential use of B-factors derived from X-ray scattering data of proteins for the understanding the (functional) dynamics of proteins, we present a comparison of B-factors of five different proteins (SH3 domain, Crh, GB1, ubiquitin and thioredoxin) with data from recent solid-state nuclear magnetic resonance experiments reflecting true (rotational) dynamics on well-defined timescales. Apart from trivial correlations involving mobile loop regions and chain termini, we find no significant correlation of B-factors with the dynamic data on any of the investigated timescales, concluding that there is no unique and general correlation of B-factors with the internal reorientational dynamics of proteins.     

Replica exchange molecular dynamics simulation of structure variation from α/4β-fold to 3α-fold protein

Replica exchange molecular dynamics (REMD) simulation provides an efficient conformational sampling tool for the study of protein folding. In this study, we explore the mechanism directing the structure variation from α/4β-fold protein to 3α-fold protein after mutation by conducting REMD simulation on 42 replicas with temperatures ranging from 270 K to 710 K. The simulation began from a protein possessing the primary structure of GA88 but the tertiary structure of GB88, two G proteins with “high sequence identity.” Albeit the large Cα-root mean square deviation (RMSD) of the folded protein (4.34 ? at 270 K and 4.75 ? at 304 K), a variation in tertiary structure was observed. Together with the analysis of secondary structure assignment, cluster analysis and principal component, it provides insights to the folding and unfolding pathway of 3α-fold protein and α/4β-fold protein respectively paving the way toward the understanding of the ongoings during conformational variation.     

1H NMR study of rehydration/dehydration and water mobility in β-cyclodextrin

The processes of dehydration and rehydration of β-cyclodextrin were studied by analysis of the (1)H NMR (nuclear magnetic resonance) line shape. Dehydration was carried in an open ampoule as a function of temperature and above 400 K total dehydration of β-cyclodextrin was observed. This result was confirmed by the thermogravimetry (TG) measurements. Rehydration was studied as a function of time at room temperature. After 40 days, β-cyclodextrin was found to absorb eight water molecules. The analysis of temperature changes in the shape of the (1)H NMR line of β-cyclodextrin kept in a closed ampoule and its dielectric measurements provided information on the mobility of water molecules. The water molecules were found to perform complex molecular motions, that is, reorientational jumps below 200K and additionally, translational motion (diffusion) above 200K.     

Impact of two different types of heat stress on chloroplast movement and fluorescence signal of tobacco leaves

Although the chloroplast movement can be strongly affected by ambient temperature, the information about chloroplast movement especially related to high temperatures is scarce. For detailed investigation of the effects of heat stress (HS) on tobacco leaves (Nicotiana tabacum L. cv. Samsun), we used two different HS treatments in dark with wide range of elevated temperatures (25–45°C). The leaf segments were either linearly heated in water bath at heating rate of 2°C min⁻¹ from room temperature up to maximal temperature (T _m) and then linearly cooled down to 25°C or incubated for 5 min in water bath at the same T _m followed by 5 min incubation at 25°C (T-jump). The changes in light-induced chloroplast movement caused by the HS pretreatment were detected after the particular heating regime at 25°C using a method of time-dependent collimated transmittance (CT) and compared with the chlorophyll O–J–I–P fluorescence rise (FLR) measurements. The inhibition of chloroplast movement started at about 40°C while the fluorescence parameters responded generally at higher T _m. This difference in sensitivity of CT and FLR was higher for the T-jump than for the linear HS indicating importance of applied heating regime. A possible influence of chloroplast movement on the FLR measurement and a physiological role of the HS-impaired chloroplast movement are discussed.     

The effect of root temperature on rose plants in relation to air temperature

Rose plants (Rosa hybrida ‘Sonia’=‘Sweet Promise’) were grown in heated (minimum night temperature 17°C), and unheated greenhouses with or without root heating to 21°C. These trials covered 6 growth cycles extending over two winter seasons. In the heated greenhouse, root heating did not increase yield, flower quality or plant development. In the unheated greenhouse, root-heated plants grew as well as those in the air-heated greenhouse as long as the air temperature did not fall below 6°C. When minimum night temperatures fell below 6°C, growth, yield and quality were reduced, irrespective of root temperature. Daytime plant water relations were studied in plants growing at 6 different root temperatures in the unheated greenhouse. Leaf resistance to water diffusion was lowest at optimal root temperature. Total leaf water potential was not significantly affected by root temperature.     

Water exchange through erythrocyte membranes: Biochemical and nuclear magnetic resonance studies re-evaluating the effects of sulfhydryl reagents and of proteolytic enzymes on human membranes

Summary The water permeability of human red blood cell (RBC) membrane has been monitored by a doping nuclear magnetic resonance (NMR) technique on intact cells and resealed ghosts following exposure to various sulfydryl-reacting (SH) reagents and proteolytic enzymes. The main conclusions are the following: (i) When appropriate conditions for exposure of erythrocytes or ghosts to mercury-containing SH reagents (concentration, temperature and duration of incubation) were found, the maximal inhibition of water diffusion could be obtained with all mercurials (including HgCl₂ and mersalyl that failed to show their inhibitory action on RBC water permeability in some investigations). While previous studies claimed that long incubation times are required for the development of maximal inhibition of water diffusion by mercurials, the present results show that it can be induced in a much shorter time (5–15 min at 37°C) if relatively high concentrations of PCMBS (2–4mm) are used and no washings of the inhibitor are performed after incubation. Higher than optimal concentrations of mercurials and/or longer incubation times result in lower values of inhibition, sometimes a loss of inhibition, or can even lead to higher values of permeability compared to control RBCs. (ii) The conditions for inhibition by mercurials are drastically changed by preincubation of erythrocytes with noninhibitory SH reagents (such as NEM or IAM) or by exposure to proteolytic enzymes. If the cells are digested with papain, the duration of incubation with PCMBS should be decreased in order for inhibition to occur. This explains the lack of inhibition reported previously, when a relatively long duration of incubation with PCMBS was used subsequent to papain digestion. (iii) The degree of inhibition of water diffusion induced by mercurials appeared to be dependent upon the temperature of which the water permeability was measured. The values of maximal inhibition ranged from 45–50% at 37°C, increased 10–15% at 20°C and further increased at lower temperatures, reaching values above 75% below 10°C; these results clarify the conflicting reports of various authors. (iv) The inhibition of water diffusion, either reversible, or irreversible, was not accompanied by significant changes in the pattern of RBC membrane polypeptides fractionated by polyacrylamide gel electrophoresis. (v) The mean value of the activation energy of water diffusion (E _a,d) obtained on 42 donors was 25.6 kJ/mol. The values ofE _a,d increased in parallel with the values of the inhibition of water diffusion induced by PCMBS until the maximal inhibition was reached (whenE _a,d=41 kJ/mol) and then both sets of values decreased in parallel.     

Mycelial growth of the snow mold fungus, Sclerotinia borealis, improved at low water potentials: an adaption to frozen environment

The snow mold fungus, Sclerotinia borealis, shows optimal growth at 4°C on potato dextrose agar (PDA) and can grow even at subzero temperature. Its mycelial growth was improved on frozen PDA at −1°C and on PDA containing potassium chloride (KCl) (water potential, −4.27 to −0.85 MPa) or d(−) sorbitol (−3.48 to −0.92 MPa). Its optimal growth temperature shifted from 4 to 10°C on PDA amended with KCl or sorbitol, indicating that inherent optimal growth occurs at high temperatures. These results suggest that S. borealis uses concentrated nutrients in the frozen environment and that such physiologic characteristics are critical for the fungus to prevail at subzero temperatures.