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- Brodelius, Peter, et al.
(författare)
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A Phosphorus-31 Nuclear Magnetic Resonance Study of Phosphate Uptake and Storage in Cultured Catharanthus roseus and Daucus carota Plant Cells
- 1985
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 260, s. 3556-3560
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Tidskriftsartikel (refereegranskat)abstract
- High resolution "P NMR spectra (103.2 MHz) ofoxygenated Catharanthus roseu8 and Daucus carotacells grown in suspension cultures were obtained usinga solenoidal perfusion probe. The spectra showed resonancesfor various phosphorylated metabolites suchas ATP, ADP, NAD(P)(H), nucleoside diphosphoglucose,and sugar phosphates. The relative levels ofthe phosphorylated metabolites remained constantthroughout the growth curve. No resonances for storagecompounds such as polyphosphates, pyrophosphate,or phytates were observed. Two resolved resonancesfor Pi indicated an intracellular pH of 7.3 and5.7 (or below) for the cytoplasm and vacuoles, respectively.The time course of Pi uptake and storage duringgrowth in fresh culture medium was followed by studyingthe level of vacuolar Pi with 31PN MR (145.7 MHz).Simultaneously, the level of Pi in the culture mediumwas followed with radioactive s2P. C. roseus quicklytakes up all the Pi from the culture medium (maximumrate 1.7 pmol min" g" (dry weight of cells)). The Pi isfirst stored in the vacuoles; subsequently, one part ofthis pool is used to keep a constant cytoplasmic Pi levelwhile another part is apparently accumulated as anNMR invisible Pi store, probably in another cell organelle.In contrast, D. carota does not accumulate Pi inthe vacuoles and consequently it takes up Pi from themedium at a much slower rate (0.05 pmol min" g"(dry weight of cells)).
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| 2. |
- Hellstrand, Per, et al.
(författare)
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Phosphagens and intracellular pH in intact rabbit smooth muscle studied by 31P-NMR
- 1985
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Ingår i: American Journal of Physiology: Cell Physiology. - 1522-1563. ; 248:3 Pt 1, s. 320-329
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Tidskriftsartikel (refereegranskat)abstract
- Phosphorus-31 nuclear magnetic resonance (NMR) spectra (103.2 MHz) were obtained from rabbit portal vein, urinary bladder, and taenia coli smooth muscle. The muscles were mounted isometrically as strips in a horizontal probe and superfused with oxygenated medium at 23 degrees C. Under these conditions the preparations could remain in a stable metabolic state for at least 24 h. The resonances observed represented phosphocreatine (PC), adenosine triphosphate (ATP), and phosphomonoester. The PC-ATP ratio in relaxed preparations was approximately 1.9 (bladder) and 1.4 (taenia coli), approximately consistent with chemical analysis of tissue extracts by isotachophoresis. However, the levels of adenosine diphosphate (ADP) and inorganic phosphate (Pi) were considerably lower as estimated by NMR, suggesting intracellular compartmentalization. Contraction for 30 min in high-K+ medium caused a reversible 10-30% decline of PC but no change in ATP. Intracellular pH was 7.0 +/- 0.1 and was unchanged by contraction or inhibition of tissue respiration by cyanide. Treatment with 2-deoxyglucose resulted in accumulation of 2-deoxyglucose 6-phosphate, verifying the pH assignment. However, the phosphomonoester resonance of normal spectra is not glucose 6-phosphate. Treatment with Mg2+-free, high-Mg2+ (10 mM), or hyperosmotic media did not alter the Mg2+ saturation of ATP. The results obtained by the nontissue-destructive 31P-NMR measurements are consistent with the view that metabolic steady-state conditions are maintained during contraction in the smooth muscle.
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| 3. |
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| 4. |
- Vogel, Hans J., et al.
(författare)
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Post-mortem energy metabolism in bovine muscles studied by non-invasive phosphorus-31 nuclear magnetic resonance
- 1985
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Ingår i: Meat Science. - 0309-1740 .- 1873-4138. ; 13:1, s. 1-18
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Tidskriftsartikel (refereegranskat)abstract
- Phosphorus-31 Nuclear Magnetic Resonance ((31)P-NMR) has been utilized to follow non-invasively the post-mortem metabolism of the major phosphorylated metabolites in muscles from beef slaughter carcasses. In addition to adenosine-5'-triphosphate (ATP), creatine phosphate (CP) and inorganic phosphate (P(i)) considerable amounts of glucose- and fructose-6-phosphate (G6P and F6P, respectively) as well as glycerol-3-phosphate (Glyc3P) were detected. ATP was mainly present as a Mg(2+)-ATP complex. Adenosine-5'-diphosphate (ADP) appeared to be mainly bound to muscle proteins. A good quantitative agreement was found for the levels of ATP, CP and sugar phosphates (SP) when estimated by NMR or enzymatic assays. Since the chemical shifts of the P(i) and sugar phosphate resonances are a function of the pH, the intracellular pH could be directly deduced from the NMR spectra. Values obtained in this manner were, within the errors of both methods, the same as those determined in iodoacetate/KCl homogenates. The pH gradients within the tissue never exceeded 0.3 pH units. In a final set of experiments we used (31)P-NMR 10 study the effects of electrical stimulation on the intracellular pH and post-mortem metabolism. It was concluded that (31)P-NMR, due to its non-invasive nature plus the fact that some of the NMR parameters are sensitive to the intracellular environment, provides a useful complement to existing methods for the study of post-mortem metabolism.
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