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Formation and retention of organically bound deuterium in rice in deuterium water release experiment.

As a substitute of tritium, deuterated water (D2O) vapor release experiments were performed in a greenhouse to estimate the different formation and subsequent retention of organically bound deuterium in rice plants between daytime and nighttime exposure. Potted rice plants were exposed to D2O vapor in the greenhouse for 8 h, under day or night conditions. Deuterium concentrations in free water and organic matter in rice leaves and ears were investigated until harvest time. The formation of organically bound deuterium in the daytime was higher than during the nighttime by the factors of 2.4 for the ear and 2.9 for the leaf. The decrease of the organically bound deuterium concentration in the ear after the nighttime exposure was faster than that after the daytime exposure. Data analysis was carried out using a compartment model in which different generating processes of organic matter were considered. The calculated organically bound deuterium retention in rice agreed with the measured value.

Deuterium↗

Physical studies of cell surface and cell membrane structure. Deuterium nuclear magnetic resonance investigation of deuterium-labelled N-hexadeconoylgalactosylceramides (cerebrosides).

1. Deuterium Fourier transform nuclear magnetic resonance spectra of a series of N-palmitoylgalactosylceramides (cerebrosides) specifically labelled with deuterium at one of positions 2', 6', 10' and 16' of the acyl chain, or in the C-6 hydroxymethyl group of the galactose residue, have been obtained using a spin-echo technique at 34.1 MHz with a homebuilt superconducting magnet spectrometer. 2. The effects of temperature and cholesterol on the deuterium spectra have been investigated. The results indicate, when compared at the same reduced temperature, that the hydrocarbon chain organization in the liquid crystalline phase of palmitoylgalactosylceramide is essentially identical to that seen in similar chain length glycerophospholipids. In particular, two sets of quadrupole splittings are seen for a 2'-labelled N-palmitoylgalactosylceramide, indicating non-equivalent deuterons as noted previously for phospholipids. 3. Two sets of quadrupole splittings are observed for the headgroup C-6-labelled N-palmitoylgalactosylceramide. It is proposed that these signals arise from the enantiomeric R and S lipids, and that motion of the hydroxymethyl group is slow (greater than 10(-5) S). These results suggest the presence of a hydrogen bond network in the polar headgroup region. 4. The effects of cholesterol on the deuterium spectra of N-palmitoylgalactosylceramide-labelled as C2H3 in the terminal methyl group, at 1:1 mol ratios and in excess water below the crystal to liquid-crystal phase transition temperature (Tc) of the pure lipid (82 degrees C), are different to the effects seen with the phosphatidylcholine-cholesterol system. The spectra below Tc are characterised by two overlapping powder patterns, one with a quadrupole splitting of approx. 6 kHz (fluid liquid-crystalline phase) and one with a quadrupole splitting of about 20--25 kHz (crystal or gel-state lipid). Exchange between these two environments is therefore slow, leading to the possibility of characterising the cerebroside-cholesterol phase diagram using deuterium nuclear magnetic resonance spectroscopy.

Cell Membrane↗

Deuterium isotope effects in carbohydrates revisited. Cryoprobe studies of the anomerization and NH to ND deuterium isotope induced 13C NMR chemical shifts of acetamidodeoxy and aminodeoxy sugars.

Complete 1H and 13C NMR chemical shift assignments have been generated from a series of acetamidodeoxy and aminodeoxy sugar derivatives. For free sugars, the enhanced sensitivity of an NMR cryoprobe allowed simple 1D and 2D NMR spectra to be obtained from essentially single anomers, before significant mutarotation had occurred. The NMR assignments have been used to characterize deuterium isotope effects on 13C chemical shifts measured under conditions of slow NH to ND exchange in single solutions. Within a range of 0 to -0.138 ppm, beta, gamma, delta, and zeta deuterium isotope effects have been observed, thus providing additional reference data for assignment of the 13C NMR spectra of nitrogenous saccharides.

Amination↗

Deuterium enrichment of polycyclic aromatic hydrocarbons by photochemically induced exchange with deuterium-rich cosmic ices.

The polycyclic aromatic hydrocarbon (PAH) coronene (C24H12) frozen in D2O ice in a ratio of less than 1 part in 500 rapidly exchanges its hydrogen atoms with the deuterium in the ice at interstellar temperatures and pressures when exposed to ultraviolet radiation. Exchange occurs via three different chemical processes: D atom addition, D atom exchange at oxidized edge sites, and D atom exchange at aromatic edge sites. Observed exchange rates for coronene (C24H12)-D2O and d12-coronene (C24D12)-H2O isotopic substitution experiments show that PAHs in interstellar ices could easily attain the D/H levels observed in meteorites. These results may have important consequences for the abundance of deuterium observed in aromatic materials in the interstellar medium and in meteorites. These exchange mechanisms produce deuteration in characteristic molecular locations on the PAHs that may distinguish them from previously postulated processes for D enrichment of PAHs.

Deuterium↗

In vivo and in vitro research on the biological effects of deuterium-depleted water: 1. Influence of deuterium-depleted water on cultured cell growth.

Deuterium depleted-water (DDW) is a new available tool for decreasing deuterium concentration in the environment of cells in culture. Several types of established cell lines, both normal and neoplastic were grown in culture media dissolved with DDW and compared with the same strains, in the same amounts, grown in media dissolved with normal distilled water. Naive mice splenocytes were grown, under stimulation with proliferation triggers, like bacterial lipopolysaccharide (LPS) and Concanavalin A (ConA) in the same conditions. The growth and proliferation were estimated using the MTT assay. Both established cell types and explanted splenocytes in the DDW-media had a significantly higher growth rate than cell cultured in normal media. In an attempt to identify the membrane mechanisms involved in the growth stimulation by DDW, the membrane proton transporters Na+/H+ antiporter and H+/K+ATP-ase were inhibited with their selective blockers amiloride and respectively lansoprazole. The results, however incomplete, point towards a lack of involvement of the Na+/H+ antiporter and a possible implication of the H+/K+ATP-ase.

2-Pyridinylmethylsulfinylbenzimidazoles↗

Deuterium-oxide-induced histamine release from basophils of allergic subjects. I. Responsiveness to deuterium oxide requires an activation step.

Basophils from many atopic persons, and especially asthmatic patients, have been shown to release histamine in response to 44% deuterium oxide (D2O), whereas basophils from nonatopic persons do not release histamine. The present experiments analyzed the mechanisms by which D2O mediated release. It was found that although D2O induced release from washed leukocytes, it failed to induce release from whole blood or from leukocytes that had sedimented but had not been washed. The kinetics of release after washing were rapid and were equivalent regardless of the temperature at which cells were sedimented (O degrees or 37 degrees C). Washed cells became desensitized to the action of D2O within 30 to 60 min at 37 degrees C, whereas unwashed leukocytes did not become desensitized. Serum or plasma inhibited D2O-induced release, although high concentrations (1/5) were less inhibitory than lower ones (1/10 to 1/100). Basophils from D2O responders also released histamine in response to a "platelet enhancing factor" (PEF), whereas those from D2O nonresponders did not. As with D2O-mediated release, PEF-mediated release occurred only with washed leukocytes, desensitized within 30 to 60 min at 37 degrees C, and was inhibited by serum. These results suggest that D2O induces histamine release by augmenting the effects of an endogenous activation mechanism, and that PEF acts on the same (D2O-responsive) donors to augment this activation mechanism. Cell activation, as well as desensitization of this activation mechanism, occurs rapidly when basophils are washed free of plasma inhibitors and placed at 37 degrees C.

Asthma↗

Effects of acute hypobaric hypoxia on the appearance of ingested deuterium from a deuterium oxide-labelled carbohydrate beverage in body fluids of humans during prolonged cycling exercise.

To determine whether or not acute hypobaric hypoxia alters the rate of water absorption from a carbohydrate beverage ingested during exercise, six men cycled for 80 min on three randomly assigned different occasions. In one trial, exercise was performed in hypoxia (barometric pressure, P(B) = 594 hPa, altitude 4,400 m) at an exercise intensity selected to elicit 75% of the individual's maximal oxygen uptake (VO2max) previously determined in such conditions. In the two other experiments, the subjects cycled in normoxia (P(B) = 992 hPa) at the same absolute and the same relative intensities as in hypoxia, which corresponded to 55% and 75%, respectively, of their VO2max determined in normoxia. The subjects consumed 400 ml of a 12.5% glucose beverage just prior to exercise, and 250 ml of the same drink at 20, 40 and 60 min from the beginning of exercise. The first drink contained 20 ml of deuterium oxide to serve as a tracer for the entry of water into body fluids. The heart rate (HR) during exercise was higher in hypoxia than in normoxia at the same absolute exercise intensity, whereas it was similar to HR measured in normoxia at the same relative exercise intensity. Both in normoxia and hypoxia, plasma noradrenaline concentrations were related to the relative exercise intensity up to 40 min of exercise. Beyond that duration, when exercise was performed at the highest absolute power in normoxia, the noradrenaline response was higher than in hypoxia at the same relative exercise intensity. No significant differences were observed among experimental conditions, either in temporal profiles of plasma D accumulation or in elimination of water ingested in sweat. Conversely, elimination in urine of the water ingested appeared to be related to the severity of exercise, either high absolute power or the same relative power combined with hypoxia. We concluded that water absorption into blood after drinking a 12.5% glucose beverage is not altered during cycling exercise in acute hypobaric hypoxia. It is suggested that the elimination of water ingested in sweat and urine may be dependent on local circulatory adjustments during exercise.

Absorption↗

Deuterium nuclear magnetic resonance of deuterium-labeled diacetyldeuterohemin incorporated into sperm whale myoglobin.

The heme derivative 2,4-diacetyldeuterohemin deuterated in the methyl groups of the acetyl moieties was reconstituted with sperm whale apomyoglobin and the two labeled methyl groups in the protein environment were observed by deuterium nuclear magnetic resonance spectroscopy. The results were compared to the free hemin form as the dimethyl ester in chloroform and in a pyridine-water mixture, as well as in the zinc complex form. Under most conditions the two methyl resonances overlie each other to a large degree. Resonance width at half-height is of the order of 25 Hz for the protein and approximately one-third as much for the free hemin at 16 degrees and is little affected by conversion to paramagnetic derivatives. Chemical shifts for the oxy- and carbonmonoxymyoglobins are very similar. In cyanoferrimyoglobin a positive pseudo-contact contribution of 3.04 ppm was computed to explain a relative upfield shift offset in part by a small negative contact shift contribution. The cyanoferrimyoglobin resonance was sensitive to the presence of phosphate buffer as well as to cyclopropane. The aquoferrimyoglobin form shows distinct resonances for the two methyl groups, with the downfield resonance considerably broadened. The expected effects of temperature on chemical shift were observed, the paramagnetic derivatives showing an effect and carbonmonoxymyoglobin showing none. The relaxation behavior was gauged from the line widths and from measurements of spin-lattice relaxation time, T1. The effective rotational correlation time is of the order of 50 ps for the liganded myoglobin forms. The temperature dependence of the line widths may imply an increased retational freedom with increasing temperature. The broadening observed in the aquoferrimyoglobin case is indicative of restricted internal rotational motion of one of the methyl groups. The method is suitable for probing the more mobile structures in proteins and retains its value in the neighborhood of paramagnetic centers.

Animals↗

Synthesis of Deuterium-Labeled Plant Sterols and Analysis of Their Side-Chain Mobility by Solid State Deuterium NMR.

The plant sterols sitosterol and stigmasterol exert very different effects on plant model membranes, the first one being a "reinforcer" like cholesterol, the second one not. 25-(2)H-Stigmasterol has been synthesized by coupling of the 22-aldehyde derived from stigmasterol by ozonolysis, with the proper sulfone labeled in position 25. The configuration of the ethyl side chain at C-24 was controlled by separation of the diastereomers introduced via a chiral sulfoxide. This synthetic scheme allowed the introduction of a labeled side chain in plant sterols in eight steps for stigmasterol and nine for sitosterol (overall yield ca. 15%). Using both diastereomers, the 24-epimers of sitosterol (clionasterol) and stigmasterol (poriferasterol) have also been synthesized. Deuterium NMR on oriented lipid bilayers made of soybean phosphatidylcholine and containing these four labeled plant sterols clearly reveals the difference of orientation and mobility of the four side chains.

Journal Article↗