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J Seelig

Publications and source records attributed to J Seelig.

At least 109 records · Page 6Linked to original sources
PubMed1987

Effects of the anti-cancer drug adriamycin on the energy metabolism of rat heart as measured by in vivo 31P-NMR and implications for adriamycin-induced cardiotoxicity.

In vivo 31P-NMR was used to measure the effects of the anti-tumor drug adriamycin on the energy metabolism of rat heart. The exclusive acquisition of NMR signal from cardiac muscle was assured by positioning a solenoidal radio-frequency NMR coil around the heart. Appropriate control experiments verified that 31P-NMR spectra solely originated from this organ. Acute effects occurring shortly after adriamycin administration are expressed in 31P spectra as a dose-dependent decline in the cardiac levels of phosphocreatine, after which stabilization at a new steady-state level occurs. These acute effects of a single dose are complete in 30-60 min and no significant further changes take place within 150 min after drug introduction. Longer-term effects of single high doses and of multiple lower doses were measured up to a week after the initiation of treatment. It seemed that at a total dose of 20 mg/kg, drug-induced interference with cardiac energy metabolism was more pronounced than at the same dose in the acute phase. These 31P-NMR data demonstrate that adriamycin treatment is accompanied by a decrease of the cardiac phosphocreatine/ATP ratio which might be an expression of the well-established cardiotoxicity of the drug.

Adenosine Triphosphate↗
PubMed1987

2H-NMR studies on ether lipid-rich bacterial membranes: deuterium order profile of Clostridium butyricum.

Palmitic acid specifically deuterated at different carbon atoms, has been incorporated biosynthetically into the membrane lipids of Clostridium butyricum. The lipids of this organism are rich in plasmalogens and their glycerol acetals and exhibit an unusual fatty acyl and alkenyl chain distribution with saturated chains mainly at the sn-2 position and unsaturated chains at the sn-1 position. The ordering of the deuterated hydrocarbon chains in whole cells was measured with deuterium nuclear magnetic resonance and was compared to the order profiles of isolated cell membranes and membranes formed from the total phospholipid extract. The shape of the order profiles was similar for all three membranes, but the absolute values of the order profiles in whole cells and isolated membranes were lower than those of the liposomal lipids. The order profiles have the same characteristic shape as those found for the lamellar liquid-crystalline phases of synthetic diacylphospholipids.

Cell Membrane↗
PubMed1987

Calcium binding to mixed phosphatidylglycerol-phosphatidylcholine bilayers as studied by deuterium nuclear magnetic resonance.

The binding of calcium to bilayer membranes composed of mixtures, in various proportions, of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) plus 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) was investigated by using atomic absorption spectroscopy and deuterium nuclear magnetic resonance. The number of bound calcium ions, X2, was determined in the low calcium concentration range (up to 100 mM) via atomic absorption spectroscopy. Simultaneous measurements of the deuterium magnetic resonance spectra of POPC, specifically deuteriated at the alpha-methylene segment of the choline head group, revealed a linear relationship between the quadrupole splitting, delta vQ, and X2 for each particular proportion of POPC-POPG. The amount of bound calcium was then determined at much greater calcium concentrations, where the atomic absorption spectroscopy measurements were unreliable, using deuterium magnetic resonance. At low Ca2+ concentrations, the amount of bound Ca2+ increased linearly with increasing proportion of POPG, demonstrating an electrostatic contribution to Ca2+ binding. At high Ca2+ concentrations, the calcium binding isotherms exhibited saturation behavior with a maximum binding capacity of 0.5 Ca2+ and 1.0 Ca2+ per phospholipid for pure POPC and mixtures of POPC-POPG, respectively. Simultaneous deuteriation of POPG and POPC showed that both lipids remained in a fluidlike lipid bilayer at all Ca2+ concentrations tested. Any phase separation of quasi-crystalline Ca2+-POPG clusters could be excluded. The residence time of Ca2+ at an individual head group binding site was shorter than 10(-6)-10(-5) s. Thus, Ca2+ ions accumulate near the negatively charged POPG-POPC membrane surface but move freely in a "trough" of the electrical potential. The effective surface charge density, sigma, could be determined from the measured amount of bound Ca2+. Subsequently, the surface potential, psi 0, and the concentration of free Ca2+ ions at the plane of ion binding could be calculated by employing the Gouy-Chapman theory. The availability of these parameters allowed a rigorous evaluation of various models for the chemical contribution to Ca2+ binding. For mixed POPC-POPG bilayers, a simple Langmuir adsorption model yielded the best fit to the experimental data, and the binding constants were 19.5 and 18.8 M-1 for POPG contents of 20 and 50 mol %, respectively. Sodium binding was comparatively weak with a binding constant of 0.6-0.85 M-1.(ABSTRACT TRUNCATED AT 400 WORDS)

Calcium↗
PubMed1987

Structure and dynamics of the phosphatidylcholine and the phosphatidylethanolamine head group in L-M fibroblasts as studied by deuterium nuclear magnetic resonance.

Mouse fibroblast L-M cells were grown in tissue culture medium containing selectively deuterated choline or ethanolamine. Both compounds were incorporated into the corresponding phospholipids at levels greater than 50% thus leading to a selective deuteration of these phospholipid head groups. Choline and ethanolamine were labeled at either the alpha- or the beta-carbon atom and well-resolved deuterium and phosphorus n.m.r. spectra were obtained from intact cells, crude plasma membranes and lipid extracts, leading to the following conclusions. (i) A large fraction, if not all, of the phospholipids in the intact L-M cell membranes were organized in a liquid crystalline bilayer. (ii) The phosphoethanolamine and the phosphocholine head group conformation were found to be remarkably similar in pure lipid bilayers and in intact L-M cell membranes with the head group dipoles being oriented parallel to the membrane surface. (iii) The deuterium T1 spin lattice relaxation times fell in the range of 7-25 ms and were similar in intact L-M cells and in pure lipid model membranes, suggesting that the two head groups are not involved in strong interactions with membrane proteins. The rotational diffusion rate of the two head groups was reduced by at least a factor of 10 compared to molecules of the same size in aqueous solution. (iv) The phosphocholine head group was sensitive to the size and sign of membrane surface charges as verified in mixing experiments with charged lipids. In L-M cell membranes the phosphocholine appeared to sense an electrically neutral environment in spite of the fact that L-M cell membranes contain 10-20% negatively charged lipids.

Animals↗
PubMed1987

A neutron diffraction study of the headgroup conformation of phosphatidylglycerol from Escherichia coli membranes.

By using neutron diffraction, the headgroup conformation of purified phosphatidylglycerol from Escherichia coli membranes has been investigated. Measurements at 25 degrees C and 15% relative humidity on oriented multilayers of lipid selectively deuterated at the sn-3-position of the glycerol backbone and of the gamma-position of the glycerol headgroup show that the labels are at a mean distance of 23.0 A and 27.6 A from the centre of the hydrocarbon chain region. This suggests that the negatively charged headgroup is oriented at about 30 degrees to the membrane surface. The orientation of the phosphatidylglycerol headgroup makes the negatively charged phosphate group easily accessible to cations present in the adjacent water layer.

Cell Membrane↗
PubMed1987

Ketogenesis in the living rat followed by 13C-NMR spectroscopy. Infusion of [1,3-13C]octanoate.

13C-NMR spectroscopy was used as a noninvasive approach to study the metabolism of [1,3-13C]octanoate in rat liver. Using a properly adjusted surface coil a liver selection of better than 90% was achieved in the intact animal without abdominal surgery. After infusion of [1,3-13C]octanoate via the jugular vein different patterns of metabolites were observed depending on the physiological state of the rat. In the fasted animal, the major metabolites were those of the Krebs cycle while in the diabetic animal ketogenic end products were predominant. As a fatty acid of medium chain length octanoate is imported into the inner mitochondrial space without control by the carnitine acyl transferase system. Hence, the metabolic differences observed between diabetic and fasted rats result from an intramitochondrial control mechanism. The in vivo 13C-NMR results therefore support previous biochemical in vitro studies which concluded that a major control of ketone body production occurs in the inner mitochondrial space, presumably via the redox potential of the liver. As an unexpected result, 13C-NMR provides evidence for the transitory esterification of the infused 13C-labeled octanoic acid. The corresponding 13C-NMR chemical shifts are typical for glycerides.

Animals↗
PubMed1986

Metabolic pathways for ketone body production. 13C NMR spectroscopy of rat liver in vivo using 13C-multilabeled fatty acids.

The hormonal regulation of ketogenesis in the liver of living rat has been studied noninvasively with 13C nuclear magnetic resonance. The protocol involved the use of a surface coil that was placed on the skin of the rat, directly over the normal location of the liver. Signals from superficial tissue were suppressed with a 180 degrees pulse at the center of the coil. A resolution of 0.6 ppm was obtained in the 13C NMR spectra at 20.1 MHz, which was equal to or better than that observed in experiments where the liver was surgically exposed and surrounded with radiofrequency coil. The spatial selection for the liver was better than 90%, with extrahepatic adipose tissue contributing only a very small amount of signal. The metabolic activities of the liver were investigated by infusion of 13C-labeled butyrate in the jugular vein of the anesthetized rat. The rate of butyrate infusion was chosen to be close to the maximum oxidative capacity of the rat liver, and the 13C signal intensities were enhanced by using doubly labeled [1,3-13C]butyrate as a substrate. Different 13C NMR spectra and hence different metabolites were observed depending on the hormonal state of the animal. In the fasted rat, the most intense 13C signal came from the end product of the Krebs cycle, namely, HCO3, with additional resonances from glutamine and glutamate. Weak resonances of the ketone bodies 3-hydroxybutyrate and acetoacetate could also be detected and allowed an evaluation of the "redox state" of the in vivo liver.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

Comparative 2H- and 31P-NMR study on the properties of palmitoyllysophosphatidylcholine in bilayers with gramicidin, cholesterol and dipalmitoylphosphatidylcholine.

The stoichiometric palmitoyllysophosphatidylcholine (lysoPC)/gramicidin (4:1, mol/mol) lamellar complex (Killian, J.A., De Kruijff, B., Van Echteld, C.J.A., Verkleij, A.J., Leunissen-Bijvelt, J. and De Gier, J. (1983) Biochim. Biophys. Acta 728, 141-144) is a useful model system to investigate the various aspects of lipid protein interactions. To study the effect of gramicidin on local order and motion of 1-palmitoyl-sn-glycero-3-phosphocholine (lysoPC) we employed 31P and 2H nuclear magnetic resonance (NMR) using selectively deuterated lysoPC's and we compared the results to those obtained for lysoPC in bilayers with cholesterol (1:1, mol/mol) and dipalmitoylphosphatidylcholine (DPPC) (1:4, mol/mol). 2H-NMR experiments on acyl chain deuterated lysoPC showed similar quadrupole splittings in the liquid crystalline state for the lysoPC/DPPC and the lysoPC/gramicidin samples. In the lysoPC/cholesterol sample an increase of the quadrupole splitting was found. T1 measurements showed that gramicidin decreases the lysoPC acyl chain motion, especially at the C12 position. In the lysoPC/cholesterol sample an increase of motion was observed as compared to lysoPC in fluid bilayers of DPPC. 31P-NMR and 2-H-NMR measurements of lysoPC, deuterated at the alpha- and beta-position of the choline moiety, indicated an increase in headgroup flexibility in all samples as compared to the parent compound DPPC. In addition, a change in headgroup conformation was observed. The alpha- and beta-segments in all samples exhibited concerted motion. It was found that also in the polar headgroup gramicidin induces a decrease of the rate of motion.

Cholesterol↗
PubMed1986

Cerebral metabolic studies in situ by 31P-nuclear magnetic resonance after hypothermic circulatory arrest.

Cerebral high energy phosphates were studied in the intact rabbit brain using nuclear magnetic resonance spectroscopy. The effect of hypothermia on degradation kinetics in total ischemia due to circulatory arrest was examined, measuring phosphocreatine, adenosine triphosphate, and inorganic phosphate as a function of time at three different temperatures (35, 24, 21 degrees C). Phosphocreatine- and ATP-decays followed single exponential functions at all three temperatures. The half-life times increased by approximately a factor of three upon lowering the temperature from 35 to 21 degrees C with activation energies of 15-20 kcal/mol, which corresponds to values of Q10 between 2.4 and 3.2. In the temperature range studied, no critical temperature was found below which metabolism would stop completely. We conclude that nuclear magnetic resonance spectroscopy allows, in the intact animal, quantitative assessment of the influence of hypothermia on energy metabolism in the brain. This influence is a major concern in the field of cardiac surgery in infants and children who are often operated in total circulatory arrest under deep hypothermia.

Adenosine Triphosphate↗
PubMed1985

Side-chain dynamics of two aromatic amino acids in pancreatic phospholipase A2 as studied by deuterium nuclear magnetic resonance.

The flexibility of individual amino acid side chains of pancreatic phospholipase A2 in aqueous and micellar solutions was studied with deuterium nuclear magnetic resonance (2H NMR). Bovine pancreatic phospholipase A2 was selectively deuterated at the aromatic ring systems of Trp-3 and Phe-5 and porcine pancreatic phospholipase A2 at Trp-3 only. Solid-state 2H NMR spectra of the lyophilized enzymes exhibited quadrupole splittings on the order of 130 kHz, indicating almost complete immobilization of the aromatic ring systems. Exposure to a water-saturated atmosphere did not remove these steric constraints. However, side-chain mobility could be induced for the tryptophyl residue of the bovine enzyme by dissolving this enzyme in aqueous buffer or micellar solution whereas the phenyl ring always remained immobile and served as a probe for the protein's overall rotation. Typical correlation times for the tryptophyl and phenyl aromatic ring systems in aqueous solution were 7 ps and 13 ns (at 20 degrees C), respectively. The correlation time of the phenyl ring was longer than expected for the monomeric protein (approximately 6 ns), suggesting some aggregation of the protein at the high concentrations used for the NMR measurements. Addition of a micellar solution of oleoylphosphocholine had no influence on the motional freedom of the tryptophyl residue but approximately doubled the correlation time of the phenyl ring, indicating an increase of the effective volume of the tumbling particle due to lipid-protein interaction. A different behavior was observed for the Trp-3 residue of porcine phospholipase A2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Phospholipid composition and organization of cytochrome c oxidase preparations as determined by 31P-nuclear magnetic resonance.

The molecular organization as well as the composition of the phospholipids in cytochrome c oxidase preparations (bovine heart) were investigated by 31P-nuclear magnetic resonance. In the so-called 'lipid-rich' preparation the lipids were found to form a fluid bilayer around the enzyme since the 31P-NMR spectrum was characteristic of a fast, axially symmetric motion of the phosphate groups with a chemical shift anisotropy of delta sigma = -45 ppm. In contrast, the 'lipid-depleted' cytochrome c oxidase gave rise to a broader spectrum where the motion of the phospholipids was no longer axially symmetric. Nevertheless, the total width of the spectrum was still considerably narrower than observed for immobilized phospholipids in solid crystals. Both enzyme preparations were dissolved in 1% detergent solution and used for high-resolution 31P-NMR spectroscopy. Narrow lines of about 20 Hz linewidth were obtained for both types of enzyme preparations, and well-resolved resonances could be assigned to cardiolipin, phosphatidylethanolamin and phosphatidylcholine. The major differences between lipid-rich and lipid-depleted cytochrome c oxidase were the absolute amount of phospholipid associated with the protein and the relative contribution of the individual lipid classes to the 31P-NMR spectrum. For lipid-rich cytochrome c oxidase about 130 molecules phospholipid were bound per enzyme (approx. 11 cardiolipins, 54 phosphatidylethanolamines and 64 phosphatidylcholines). For lipid-depleted cytochrome c oxidase only 6-18 lipids were bound per enzyme (1 or 2 cardiolipins, 3-8 phosphatidylethanolamines and 2-8 phosphatidylcholines). In contrast to earlier suggestions that cardiolipin is the only remaining lipid in lipid-depleted cytochrome c oxidase, the 31P-NMR studies demonstrate that all three lipids remain associated with the protein.

Animals↗
PubMed1985

Phosphocreatine content and intracellular pH of calf muscle measured by phosphorus NMR spectroscopy in occlusive arterial disease of the legs.

Energy metabolism of calf muscle was assessed non-invasively by phosphorus (31P) NMR spectroscopy in eleven patients with symptomatic arterial occlusion and in seven matched controls. Phosphocreatine (PCr) content and pH values decreased during non-ischaemic foot exercise to lower values in severely afflicted patients but in all patients, as a group, they were not significantly decreased compared to controls. In contrast, recovery from ischaemic exercise (arterial occlusion by a tourniquet) demonstrated significant differences between patients and controls. Intracellular pH and PCr recovered more slowly in patients than in controls; PCr recovery proceeded exponentially with a recovery half-time of 203 +/- 74 s in patients compared to 36.7 +/- 5.5 s in controls (P less than 0.02). Phosphocreatine (PCr) recovery after ischaemic exercise correlated significantly with the degree of arterial stenoses as assessed by Doppler ultrasound (r = 0.739, P = 0.019) and by angiography (r = 0.885, P = 0.005), suggesting that the degree of large vessel stenoses limits the postischaemic increase in mitochondrial oxidative phosphorylation. Reactive blood flow after ischaemia failed to correlate with PCr recovery or with the degree of arterial stenoses. Phosphorus (31P) NMR spectroscopy provides, therefore, quantitative parameters of muscle energy metabolism in patients with peripheral arterial occlusions.

Arterial Occlusive Diseases↗
PubMed1985

Extremity bone tumors: evaluation by P-31 MR spectroscopy.

High-resolution P-31 MR spectra were obtained in four patients with bone tumors of their distal extremities. In one case the tumor, a Ewing sarcoma of the tibia, was investigated during clinical remission after radiation therapy and chemotherapy. The other three cases - one low-grade chondrosarcoma of the tibial head, one malignant fibrous histiocytoma of the tibia, and one chondroblastoma of the medial femoral condyle - showed clinically active tumor growth, with corresponding increased metabolism as demonstrated by bone scintigraphy. The spectra of the three active tumors indicated a comparably high adenosine triphosphate content, similar to previously published spectra from animal tumors or human tumors implanted into animals. There were also high resonances of inorganic phosphate and low resonances of phosphocreatine; there were definite peaks in the phosphodiester and phosphomonoester regions, indicating the existence of these metabolites in the tumors. Slight but definite changes in the metabolite content were observed in one tumor after chemotherapy. The spectra of the unaffected leg did not show any well-resolved P-31 signals, which is typical for healthy bone. These are the first P-31 MR spectra of human bone tumors measured in patients to our knowledge.

Adolescent↗
PubMed1984

Ketogenesis in the living rat followed by 13C NMR spectroscopy.

The metabolic fate of 13C1-labeled butyrate in the liver of living rats has been studied by 13C NMR. The formation of the ketone bodies acetoacetate and beta-hydroxybutyrate was observed in vivo as well as resonances from glutamate, glutamine, and carbonate. The observed time course of these metabolites demonstrates the potential of the technique to measure enzyme kinetics in vivo and also to measure the enzyme capacity of a given organ to metabolize a substrate. The in vivo spectra were compared to in vitro spectra of the excised liver and perchloric acid extracts of the liver. Observation of the metabolites and monitoring of their time course in vivo would not have been possible without distinct improvements in the spectral resolution and the spatial localization of the radio-frequency field within the liver. As a novel approach, we have selected the carbonyl region of the 13C NMR spectra for identifying the ketone bodies and other oxidation products in the liver.

3-Hydroxybutyric Acid↗
PubMed1984

Ca2+ binding to phosphatidylcholine bilayers as studied by deuterium magnetic resonance. Evidence for the formation of a Ca2+ complex with two phospholipid molecules.

The binding of Ca2+ to bilayer membranes composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) was investigated with atomic absorption spectroscopy and deuterium magnetic resonance, leading to the following conclusions. Atomic absorption spectroscopy allowed the determination of the amount of Ca2+ bound to the membrane surface (Cb) at low Ca2+ concentrations (3-100 mM). Simultaneous measurements of the deuterium magnetic resonance spectra of POPC with specifically deuterated choline head groups revealed a linear relationship between the quadrupole splitting and the amount of bound Ca2+. With this calibration, the amount of bound Ca2+ could be determined from the deuterium spectra under conditions where atomic absorption spectroscopy was technically not feasible, i.e., in the concentration range of 0.1-5 M CaCl2. The Ca2+ binding isotherm exhibited saturation behavior. The quadrupole splitting at the saturation limit corresponded to a binding stoichiometry of one Ca2+ per two POPC molecules. The surface charge density (sigma) could be evaluated from the amount of bound Ca2+ and the surface area per POPC molecule. By employing the Gouy-Chapman theory, it was then possible to determine the surface potential (psi 0) and the Ca2+ concentration immediately at the lipid-water interface (CI). With this set of experimental parameters, various models for the mode of Ca2+ binding were tested. A simple partition equilibrium or a Langmuir absorption model could be ruled out. However, a very good fit to the experimental data was obtained by applying the law of mass action in the form Cb/(1 - 2Cb)2 = KCI in which K is the only adjustable parameter.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcium↗
PubMed1983

Hydration of Escherichia coli lipids. Deuterium T1 relaxation time studies of phosphatidylglycerol, phosphatidylethanolamine and phosphatidylcholine.

The hydration properties of Escherichia coli lipids (phosphatidylglycerol, phosphatidylethanolamine) and synthetic 1,2-dioleoyl-sn-glycero-3-phosphocholine in H2O/2H2O mixtures (9:1, v/v) were investigated with 2H-NMR. Comparison of the 2H2O spin lattice relaxation time (T1) as a function of the water content revealed a remarkable quantitative similarity of all three lipid-H2O systems. Two distinct hydration regions could be discerned in the T1 relaxation time profile. (1) A minimum of 11-16 water molecules was needed to form a primary hydration shell, characterized by an average relaxation time of T1 approximately equal to 90 ms. (2) Additional water was found to be in exchange with the primary hydration shell. The exchange process could be described in terms of a two-site exchange model, assuming rapid exchange between bulk water with T1 = 500 ms and hydration water with T1 = 80-120 ms. Analysis of the linewidth and the residual quadrupole splitting (at low water content) confirmed the size of the primary hydration layer. However, each lipid-water system exhibited a somewhat different linewidth behavior, and a detailed molecular interpretation appeared to be preposterous.

Cell Membrane↗
PubMed1983

The determination of the separate Ca2+ pump protein and phospholipid profile structures within reconstituted sarcoplasmic reticulum membranes via X-ray and neutron diffraction.

We have previously compared the electron density profiles for several highly-functional reconstituted sarcoplasmic reticulum membranes with that for the isolated sarcoplasmic reticulum membrane (Herbette, L., Scarpa, A., Blasie, J.K., Wang, C.T., Saito, A. and Fleischer, S. (1981) Biophys. J. 36, 47-72). In this paper, we compare the separate calcium pump protein profile within these reconstituted sarcoplasmic reticulum membranes, as derived by X-ray and neutron diffraction methods, with that within isolated sarcoplasmic reticulum membranes. In addition, the time-average perturbation of the lipid bilayer by the incorporated calcium pump protein within these reconstituted sarcoplasmic reticulum membranes has been determined in some detail.

Animals↗