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  • Articles  (48)
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  • Springer  (48)
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  • 1995-1999  (48)
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  • 1
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    Springer
    Journal of inorganic and organometallic polymers and materials 5 (1995), S. 217-236 
    ISSN: 1572-8870
    Keywords: Polysilane ; NMR ; pyrolysis ; randomization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Bimodal molecular weight distributions of alternating copolymers of the type (Me2Si−R2Si−SiMe2) n , where R=hexyl and butyl, were obtained by polymerization of BrSiMe2−R2Si−Me2SiBr, using the Wurtz coupling method. Analysis of the samples by pyrolysis GC MS and solution29Si NMR indicated that some randomization occurred, due to cleavage of the original Si−Si bonds in the monomer. The extent of randomization was significantly greater in the high molecular weight fractions. Based on the nature of the rings from the py MGC/MS traces and the number on nonads found in the29Si spectra, two types of randomization processes have been proposed, involving backbiting followed by (1) ring expulsion or (2) redistribution.
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  • 2
    ISSN: 1572-8927
    Keywords: Hydrogen-1 ; carbon-13 ; nitrogen-15 ; NMR ; magnesiumisothiocyanate complexes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract A hydrogen-1, carbon-13, and nitrogen-15 NMR study of magnesium(II)-isothiocyanate complexation in aqueous mixtures has been completed. At temperatures low enough to slow proton and ligand exchange, separate1H,13C, and15N NMR signals are observed for coordinated and bulk water molecules and anions. The1H NMR spectra reveal signals for the hexahydrate and the mono-through triisothiocyanato complexes, as well as two small signals attributed to [Mg(H2O)5(OH)]1+ and [Mg(H2O)4(OH)(NCS)]. Accurate hydration numbers were obtained from signal area integrations at each NCS− concentration. In the15N NMR spectra, signals also were observed for the mono-through triisothiocyanato complexes, and a small signal believed to be due to [Mg(H2O)4(OH)(NCS)]. Coordination number contributions for NCS− were measured from these spectra and when combined with the hydration numbers they totalled essentially six at each anion concentration. Signals for [Mg(H2O)5(NCS)]1+ through [Mg(H2O)3(NCS)3]1− also were observed in the13C NMR spectra and the area evaluations were comparable to the15N NMR results. An analysis of the magnitude and sign of the coordinated NCS− chemical shifts identified the nitrogen atom as the anion binding site. All spectra indicated [Mg(H2O)5(NCS)]1+ and [Mg(H2O)4(NCS)2] were the dominat isothiocyanato complexes over the entire range of anion concentrations. The inability to detect evidence for complexes higher than the triisothiocyanato reflects the competitive binding ability of water molecules and perhaps the decreased electrostatic interaction between NCS− and negatively charged higher complexes.
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  • 3
    ISSN: 1572-8773
    Keywords: lead ; lipid peroxidation ; pro-oxidant ; anti-oxidant ; rat brain
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Lipid peroxidation in vitro homogenates of brain was examined as sequela of lead toxicity. The levels of malondialdehyde (MDA) in homogenates of rat brain (1 ml, 5% w/v) treated with lead (50 μg) alone or in combination with ascorbic acid (100 μg), alphatocopherol (100 μg) or hydroquinone (100 μg) were evaluated. The levels of MDA were consistently evoked by lead in a dose-related manner. The toxicity of lead was further advanced by the action of the pro-oxidant drug ascorbic acid on the brain. However, the anti-oxidant drugs alphatocopherol and hydroquinone decreased the toxic effect of lead on the brain. These results clearly show that the enhanced lipid peroxidation may provide a basis of lead-induced neurotoxicity.
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  • 4
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    Monatshefte für Chemie 126 (1995), S. 233-239 
    ISSN: 1434-4475
    Keywords: Enaminoketones ; Lupin alkaloid ; Multiflorine ; NMR ; Nucleophilic methylation ; Stereospecificity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Description / Table of Contents: Zusammenfassung Multiflorin (1), ein Lupin-Nebenalkaloid, ergibt bei Umsetzung mit Methyllithium oder Methylmagnesiumiodid 4S-4-Hydroxy-4-methyl-2,3-didehydrospartein (2) und 2S-2-Methyl-4-oxospartein (3) als Hauptprodukte. Ihre NMR-spektroskopisch (1H und13C) aufgeklärte räumliche Struktur weist auf eine Stereoselektivität der erwähnten Reaktionen hin. Die beobachteten nucleophilen 1,2- und 1,4-Additionen zeigen, daß sich die Regiospezifität der Einwirkung von MeLi oder MeMgl auf Multiflorin von jener bis jetzt bekannter Alkylierungen von Enaminoketonen unterscheidet.
    Notes: Summary Multiflorine (1) — a minor lupine alkaloid — treated by methyl lithium or methyl magnesium iodide affords 4S-4-hydroxy-4-methyl-2,3-didehydrosparteine (2) and 2S-2-methyl-4-oxosparteine (3), respectively, as the dominating products. Their steric structure, determined by1H and13C NMR techniques, points to stereospecific preferences of these reactions. The observed nucleophilic 1,2- and 1,4-additions indicate that regiospecificity of the action of MeLi or MeMgI on multiflorine is different from that of the so far known similar alkylation of other enamino ketones.
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  • 5
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    Monatshefte für Chemie 126 (1995), S. 1011-1019 
    ISSN: 1434-4475
    Keywords: Precursors of strigol analogues ; Michael reaction ; NMR ; Molecular modelling
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Description / Table of Contents: Zusammenfassung BasenkatalysierteMichael-Addition von 5-Nitropentan-2-on-ethylenketal (1) und Cyclohex-2-enon (2), anschließende Entfernung der Schutzgruppe und darauffolgende Aldolkondensation liefert isomer 8-Methyl-5-nitro-1-octalone (5a,b). Struktur, relative Konfiguration und Konformation von5a und5b wurden mittels1H- und13C-NMR-Spektroskopie aufgeklärt.
    Notes: Summary Base catalyzedMichael addition of 5-nitropentan-2-one ethylene ketal (1) and cyclohex-2-enone (2), subsequent deprotection, and intramolecular aldol condensation yields the 8-methyl-5-nitro-1-octalone isomers (5a,b). The structure, relative configuration, and conformation of5a and5b were elucidated utilizing the results of1H and13C NMR investigations
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  • 6
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    Catalysis letters 31 (1995), S. 267-272 
    ISSN: 1572-879X
    Keywords: MCM-41 synthesis ; NMR ; FTIR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The synthesis of MCM-41 mesoporous compounds with Si/Al ratios as low as 2 without observing the presence of octahedral Al in27AlMAS NMR is reported. FTIR spectra of chemisorbed pyridine indicated that MCM-41 materials in their protonated form exhibit both Brønsted and Lewis acid sites.
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  • 7
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    Theoretical chemistry accounts 92 (1995), S. 61-65 
    ISSN: 1432-2234
    Keywords: NMR ; Nuclear spin coupling constant ; Maximum bond order hybrid orbital ; Phosphorus-carbon coupling ; Net atomic charge
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The novel generalized correlation of the nuclear spin-spin coupling constants with the atomic hybrids and net charges is employed to give a new relationship for calculating the directly bonded phosphorus-carbon coupling constants by use of the maximum bond order hybrid orbital procedure together with the extended Hückel molecular orbital calculation. The calculated coupling constants of phosphorus-carbon are all in good agreement with the experimental data, which shows that the new relationship obtained in the present paper is quite satisfactory for calculation of the phosphorus-carbon coupling constants.
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  • 8
    ISSN: 1572-9001
    Keywords: NMR ; distance geometry ; cluster analysis ; absolute configuration ; fusaproliferin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The absolute configuration of fusaproliferin, a toxic metabolite produced byFusarium proliferation, was determined by the combined use of1H NMR and distance geometry. The configuration of double bonds has been determined in agreement with NOESY buildup data. An R configuration for C10 was determined using Mosher's method. Processing the constraints obtained from NOESY experiments with a distance geometry program, a limited number (80) of possible structures was derived. An agglomerative nonhierarchical method of clustering was used in order to place these structures into classes suggested by the data, and not previously defined in any way. This statistical method showed that indeed the structures could be grouped in four classes. One of these classes is represented by a single structure, with the highest sum of violations and was discarded. All other structures have the same chirality; respectively S for C14 and R for C15. In solution the overall conformation is quite well defined in the region of the five-member ring and the planes of double bonds C2–C3 and C11–C12, while near to C8 and C9 internal flexibility appears evident.
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  • 9
    ISSN: 1573-4919
    Keywords: ascorbic acid ; microsomes ; NADPH ; superoxide radical ; lipid peroxidation ; oxidative damage of proteins
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract It has recently been indicated that in the absence of free iron, NADPH initiates oxidative damage of proteins in guinea pig liver microsomes and also lipid peroxidation and protein damage in cardiac microsomes and that ascorbic acid specifically inhibits both the lipid peroxidation and protein damage [Mukhopadhyay CK, Chatterjee IB: J Biol Chem 269: 13390–13397, 1994; Mukhopadhyay Met al.: Mol Cell Biochem 126: 69–75, 1993]. In this paper we demonstrate that Fe(III)-independent NADPH-initiated lipid peroxidation and oxidative damage of proteins occur in the microsomes of all the extrahepatic tissues including lung, kidney, adrenal gland and brain and that both the lipid peroxidation and protein damage are specifically prevented by ascorbic acid. We further demonstrate that when NADPH is replaced by $$O_2^{\bar \cdot } $$ as the electron donor, the $$O_2^{\bar \cdot } $$ lipid peroxidation and protein damage are also inhibited by ascorbic acid.
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  • 10
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    Molecular and cellular biochemistry 142 (1995), S. 79-87 
    ISSN: 1573-4919
    Keywords: cyclophosphamide ; curcumin ; bronchoalveolar lavage fluid ; lavage cells ; lipid peroxidation ; antioxidant defense mechanisms
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Cyclophosphamide causes lung injury in rats through its ability to generate free radicals with subsequent endothelial and epithelial cell damage. In order to observe the protective effects of a potent anti-inflammatory antioxidant, curcumin (diferuloyl methane) on cyclophosphamide-induced early lung injury, healthy pathogen free male Wistar rats were exposed to 20 mg/100 g body weight of cyclophosphamide, intraperitoneally as a single injection. Prior to cyclophosphamide intoxication oral administration of curcumin was performed daily for 7 days. At various time intervals (2, 3, 5 and 7 days post insult) serum and lung samples were analyzed for angiotensin converting enzyme, lipid peroxidation, reduced glutathione and ascorbic acid. Bronchoalveolar lavage fluid was analyzed for biochemical constituents. The lavage cells were examined for lipid peroxidation and glutathione content. Excised lungs were analyzed for antioxidant enzyme levels. Biochemical analyses revealed time course increases in lavage fluid total protein, albumin, angiotensin converting enzyme (ACE), lactate dehydrogenase, N-acetyl-β-D-glucosaminidase, alkaline phosphatase, acid phosphatase, lipid peroxide levels and decreased levels of glutathione (GSH) and ascorbic acid 2, 3, 5 and 7 days after cyclophosphamide intoxication. Increased levels of lipid peroxidation and decreased levels of glutathione and ascorbic acid were seen in serum, lung tissue and lavage cells of cyclophosphamide groups. Serum angiotensin converting enzyme activity increased which coincided with the decrease in lung tissue levels. Activities of antioxidant enzymes were reduced with time in the lungs of cyclophosphamide groups. However, a significant reduction in lavage fluid biochemical constituents, lipid peroxidation products in serum, lung and lavage cells with concomitant increase in antioxidant defense mechanisms occurred in curcumin fed cyclophosphamide rats. Therefore, our results suggest that curcumin is effective in moderating the cyclophosphamide induced early lung injury and the oxidant-antioxidant imbalance was partly abolished by restoring the glutathione (GSH) with decreased levels of lipid peroxidation.
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  • 11
    ISSN: 1573-4919
    Keywords: cisplatin ; glutathione ester ; reduced glutathione ; antioxidants ; lipid peroxidation ; nephrotoxicity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The objective of this study was to assess the therapeutic advantage of glutathione ester along with cisplatin. Comparisons were made with renal reduced glutathione, enzymatic antioxidants, and lipid peroxidation levels. Cisplatin caused differential toxic effects on renal antioxidants and lipid peroxidation. However administration of glutathione ester modulates the toxic effects of cisplatin observed in renal antioxidants and lipid peroxidation. The finding that glutathione ester co-administration along with cisplatin is more effective and advantageous in protecting against the nephrotoxicity of cisplatin when it was given alone.
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  • 12
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    Molecular and cellular biochemistry 144 (1995), S. 141-145 
    ISSN: 1573-4919
    Keywords: magnesium ; iron ; lipid peroxidation ; hepatocyte
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract In primary cultures of rat hepatocytes, the effects of extracellular Mg2+ and Fe on lipid peroxidation (LPO) as measured by means of malondialdehyde (MDA) formation were investigated. Incubation of hepatocytes at decreasing extracellular Mg2+ concentration enhanced LPO, depending on extracellular Fe. About 96% of MDA accumulated in the culture medium. Addition of desferrioxamine prevented LPO. Additionally, the formation of oxygen free radicals was determined by fluorescence reduction of cis-parinaric acid. With this method, an immediate decay of fluorescence was found after addition of Fe2+. Fluorescence reduction was completely prevented by desferrioxamine, indicating the function of extracellular Fe. This mechanism may operate additionally to the increase in intracellular Fe and intracellular formation of oxygen free radicals during Mg deficiencyin vivo.
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  • 13
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    Molecular and cellular biochemistry 147 (1995), S. 139-144 
    ISSN: 1573-4919
    Keywords: sarcolemmal Na+−K+ ATPase ; lipid peroxidation ; oxyradicals ; cardiac membrane ; oxidative stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The Na+−K+ ATPase activity and SH group content were decreased whereas malondialdehyde (MDA) content was increased upon treating the porcine cardiac sarcolemma with xanthine plus xanthine oxidase, which is known to generate superoxide and other oxyradicals. Superoxide dismutase either alone or in combination with catalase and mannitol fully prevented changes in SH group content but the xanthine plus xanthine oxidase-induced depression in Na+−K+ ATPase activity as well as increase in MDA content were prevented partially. The Lineweaver-Burk plot analysis of the data for Na+−K+ ATPase activity in the presence of different concentrations of MgATP or Na+ revealed that the xanthine plus xanthine oxidase-induced depression in the enzyme activity was associated with a decrease in Vmax and an increase in Km for MgATP; however, Ka value for Na+ was decreased. Treatment of sarcolemma with H2O2 plus Fe2+, an hydroxyl and other radical generating system, increased MDA content but decreased both Na+−K+ ATPase activity and SH group content; mannitol alone or in combination with catalase prevented changes in SH group content fully but the depression in Na+−K+ ATPase activity and increase in MDA content were prevented partially. The depression in the enzyme activity by H2O2 plus Fe2+ was associated with a decrease in Vmax and an increase in Km for MgATP. These results indicate that the depressant effect of xanthine plus xanthine oxidase on sarcolemmal Na+−K+ ATPase may be due to the formation of superoxide, hydroxyl and other radicals. Furthermore, the oxyradical-induced depression in Na+−K+ ATPase activity may be due to a decrease in the affinity of substrate in the sarcolemmal membrane.
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  • 14
    ISSN: 1573-4919
    Keywords: mitochondria ; oxidative stress ; iron ; lipid peroxidation ; membrane permeability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract It is well established that several iron complexes can induce oxidative damage in hepatic mitochondrial membranes by catalyzing the formation of ·OH radicals and/or by promoting lipid peroxidation. This is a relevant process for the molecular basis of iron overload diseases. The present work demonstrates that Fe(II)ATP complexes (5–50μM) promote an oxygen consumption burst in a suspension of isolated rat liver mitochondria (either in the absence or presence of Antimycin A), caused mainly by lipid peroxidation. Fe(II)ATP alone induced small levels of oxygen uptake but no burst. The time course of Fe(II)ATP oxidation to Fe(III)ATP in the extramitochondrial media also reveals a simultaneous ‘burst phase’. The iron chelator Desferal (DFO) or the chain-break antioxidant butylated hydroxytoluene (BHT) fully prevented both lipid peroxidation (quantified as oxygen uptake burst) and mitochondrial swelling. DFO and BHT were capable of stopping the ongoing process of peroxidation at any point of their addition to the mitochondrial suspension. Conversely, DFO and BHT only halted the Fe(II)ATP-induced mitochondrial swelling at the onset of the process. Fe(II)ATP could also cause the collapse of mitochondrial potential, which was protected by BHT if added at the onset of the damaging process. These results, as well as correlation studies between peroxidation and mitochondrial swelling, suggest that a two phase process is occurring during Fe(II)ATP-induced mitochondrial damage: one dependent and another independent of lipid peroxidation. The involvement of lipid peroxidation in the overall process of mitochondrial membrane injury is discussed.
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  • 15
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    Molecular and cellular biochemistry 151 (1995), S. 33-38 
    ISSN: 1573-4919
    Keywords: preeclampsia ; vitamin E ; lipid peroxidation ; hypertension
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Preeclampsia or pregnancy-induced hypertension is a major cause of both maternal and fetal-neonatal morbidity and mortality. The deficiency of vitamin E can cause accumulation of lipid peroxidation products, which, in turn, can induce vasoconstriction. This study has examined any evidence of increased cellular lipid peroxidation and accumulation of malonydialdehyde (MDA, an end product of lipid peroxidation) in pregnancy-induced hypertension and any relationship between the elevated MDA and lower vitamin E levels with hypertension in pregnant women. EDTA-Blood was collected from pregnant women at the time of delivery. Plasma vitamin E was determined by HPLC; MDA by the thiobarbituric acid-reactivity. Subjects with diastolic blood pressure(DBP) ≥90 mm Hg were considered hypertensive (HT) and with 〈90 mm Hg normotensive (NT). Data (Mean±SE) from 49 NT and 11 HT women show that HT has significantly lower vitamin E (22±1 vs 27±1 nmole/ml, p〈0.03) and elevated MDA levels (0.56±0.06 vs 0.43±0.02 nmole/ml, p〈0.03) compared to NT; the ages and gestational ages of women were similar. Among all women, there was a significant positive relationship between DBP and MDA levels (r=0.27, p〈0.05), and a significant negative relationship between vitamin E levels and DBP (−0.36, p〈0.005), and a significant negative relationship between MDA and vitamin E levels (r=−0.27, p〈0.05). Thus, HT women's plasma has significantly lower E and higher MDA levels, and DBP significantly correlates with the extent of vitamin E deficiency and increased MDA levels. This study suggests a relationship between elevated lipid peroxidation and lower vitamin E levels and hypertension in pregnancy (preeclampsia).
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  • 16
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    Molecular and cellular biochemistry 147 (1995), S. 77-81 
    ISSN: 1573-4919
    Keywords: antioxidants ; redox state ; lipid peroxidation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Various abnormalities have been implicated in the transition of hypertrophy to heart failure but the exact mechanism is still unknown. Thus heart failure subsequent to hypertrophy remains a major clinical problem. Recently, oxidative stress has been suggested to play a critical role in the pathogenesis of heart failure. Here we describe antioxidant changes as well as their significance during hypertrophy and heart failure stages. Heart hypertrophy in rats and guinea pigs, in response to pressure over-load, is associated with an increase in ‘antioxidant reserve’ and a decrease in oxidative stress. Hypertrophied rat hearts show increased tolerance for different oxidative stress conditions such as those imposed by free radicals, hypoxia-reoxygenation and ischemia-reperfusion. On the other hand, heart failure under acute as well as chronic conditions is associated with reduced antioxidant reserve and increased oxidative stress. The latter may have a causal role as suggested by the protection seen with antioxidant treatment in acute as well as in chronic heart failure. It is becoming increasingly apparent that, anytime the available antioxidant reserve in the cell becomes inadequate, myocardial dysfunction is imminent.
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  • 17
    ISSN: 1573-4919
    Keywords: antioxidant reserve ; membrane cholesterol/phospholipid ratio ; lipid peroxidation ; erythrocytes ; Leishmani donovani
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Visceral leishmaniasis has been found to be associated with severe anemia and premature lysis of erythrocytes. Peroxidative damage of red cells has been noted in several hemolytic anemias. Present study shows enhanced formation of methemoglobin in hamsters infected withLeishmania donovani. Increased formation of malonyldialdehyde and diene conjugate has been noted in the erythrocytes of the infected animals with the progress of anemia. Results showed decreased activities of protective enzymes like superoxide dismutase, catalase and glutathione reductase against peroxidative attack. An increase in the membrane cholesterol/phospholipid ratio and a decrease in membrane fluidity of erythrocytes were observed under the diseased condition. Densitometric scan after SDS-PAGE of red cell membrane of the infected animals revealed significant degradation of band 3 and band 4.1 proteins. The results suggest that alteration in the membrane may lead to reduced life span of the red cells in experimental visceral leishmaniasis.
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  • 18
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    Molecular and cellular biochemistry 146 (1995), S. 107-113 
    ISSN: 1573-4919
    Keywords: Marine mussels ; metal accumulation ; lipid peroxidation ; antioxidant enzymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Lipid peroxidation induced by metals at sub-lethal levels, alter physiological and biochemical characteristics of biological systems. To counter the detrimental effects of the prooxidant activity of metals, a group of antioxidant enzyme systems function in the organisms. The present study was performed to investigate into the lipid peroxidation product formation due to the exposure to effects of the metals namely aluminium, lead and cadmium at sub-lethal concentrations and the biological response through protective antioxidant enzyme activity in the marine mussels,Perna viridis Lin. This organism is a known bioindicator and bioconcentrator of metals in the environment. The results of the present study were: (a) accumulation of lead showed a definite linear increase during the period of exposure whereas aluminium and cadmium showed fluctuations. Mantle and gill tissues showed greater accumulation of metals when compared to digestive gland; (b) lead and aluminium induced lipid peroxidation was greater in tissues than the peroxidation induced by cadmium. Cadmium induced peroxidation was observed only after the day 7 of the exposure; (c) anti-oxidant enzymes activity levels were significantly higher in digestive gland and mantle than gills; (d) mantle was observed to significantly contribute to the organismal response to lipid peroxidation as indicated by high activity levels of anti-oxidant enzymes.
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  • 19
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    Molecular and cellular biochemistry 152 (1995), S. 13-21 
    ISSN: 1573-4919
    Keywords: curcumin ; diabetes ; hyperglycemia ; lipid peroxidation ; urinary metabolites
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Effect of feeding 0.5% curcumin diet or 1% cholesterol diet was examined in albino rats rendered diabetic with streptozotocin injection. Diabetic rats maintained on curcumin diet for 8 weeks excreted comparatively less amounts of albumin, urea, creatinine and inorganic phosphorus. Urinary excretion of the electrolytes sodium and potassium were also significantly lowored under curcumin treatment. Dietary curcumin also partially reversed the abnormalities in plasma albumin, urea, creatinine and inorganic phosphorus in diabetic animals. On the other hand, glucose excretion or the fasting sugar level was unaffected by dietary curcumin and so also the body weights were not improved to any significant extent. Diabetic rats fed curcumin diet had a lowered relative liver weight at the end of the study compared to other diabetic rat groups. Diabetic rats fed a curcumin diet also showed lowered lipid peroxidation in plasma and urine when compared to other diabetic groups. The extent of lipid peroxidation on the other hand, was still higher in cholesterol fed diabetic groups compared to diabetic rats fed with control diet. Thus, the study reveals that curcumin feeding improves the metabolic status in diabetic condition, despite no effect on hyperglycemic status or the body weights. The mechanism by which curcumin improves this situation is probably by virtue of its hypocholesterolemic influence, antioxidant nature and free radical scavenging property.
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  • 20
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    Bioscience reports 15 (1995), S. 55-63 
    ISSN: 1573-4935
    Keywords: water transport mechanisms ; NMR ; rat erythrocytes ; tritium effects
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The inhibition of water diffusion across the rat erythrocyte membrane was studied by NMR using two basically different types of inhibitory agents: PCMB andin vivo irradiation. The contribution of lipid and protein to water permeability revealed the inhibitory effect of each pathway. Internal contamination with tritium (25–115 mGy) reduces water permeability due to protein modifications; for doses higher than 100 mGy the lipid mediated mechanism seems also to be impaired. The same procedure enables one to assess the extent to which the higher water permeability of rat, compared to human, erythrocyte is due to one of the two pathways.
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  • 21
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    Journal of sol gel science and technology 4 (1995), S. 151-162 
    ISSN: 1573-4846
    Keywords: sol-gel transition ; rheology ; siloxane ; NMR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Ormosils are well-known organic-inorganic sol-gel derived materials also called heteropolysiloxanes. This paper presents two basic heteropolysiloxane structures where the organic part is either a short organic chain bridging two silicon atoms for the first material or an organic polymer backbone for the second. Their synthesis is detailed and a variety of experimental techniques (IR, 13C and 29Si NMR and CP-MAS NMR, GPC) have been employed to investigate the chemical structure of these new materials. Their mechanical properties, more precisely their viscoelastic behaviour, have been evaluated using dynamic rheological techniques. The storage and loss moduli have been followed during the sol-gel transition at fixed and variable oscillation frequencies. The results have been correlated to the 29Si CP-MAS NMR informations concerning the network polycondensation and compared to a pure inorganic sol-gel material prepared from tetraethoxysilane.
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  • 22
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    The protein journal 14 (1995), S. 115-126 
    ISSN: 1573-4943
    Keywords: NMR ; high-potential iron protein ; 15N assignment ; heteronuclear correlation ; paramagnetic
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The15N resonances in reduced and oxidizedChromatium vinosum high-potential iron protein have been assigned by use of1H-1H COSY spectra and1H-15N HMQC, HMQC-COSY, and HMQC-NOESY spectra. Unambiguous assignment of 70 of 85 backbone15N resonances in the reduced protein and 62 of 85 resonances in the oxidized protein are made, as are 12 of 21 side-chain15N resonances.
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  • 23
    ISSN: 1432-1424
    Keywords: Rat insulinoma cell line, CRI-G1 ; Cyclic nucleotide regulation ; Calcium-activated nonselective cation channel ; Patch clamp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The regulation of a calcium-activated nonselective cation (Ca-NS+) channel by analogues of cyclic AMP has been investigated in the rat insulinoma cell line, CRI-G1. The activity of the channel is modulated by cyclic AMP in a complex way. In the majority of patches (83%) tested concentrations of cyclic AMP of 10 μm and above cause an inhibition of channel activity which is immediately reversible on washing. In contrast, lower concentrations of cyclic AMP, between 0.1 and 1.0 μm, produce a transient activation of channel activity in most patches (63%) tested. One group of analogues, including N6-monobutyryl cyclic AMP and N6, 2′-O-dibutyryl cyclic AMP reduced the activity of the Ca-NS+ channel at all concentrations tested and 2′-O-Monobutyryl cyclic AMP produced inhibition in all patches tested except one, at all concentrations. A second group produced dual concentration-dependent effects on Ca-NS+, low concentrations stimulating and high concentrations inhibiting channel activity. 6-Chloropurine cyclic AMP and 8-bromo cyclic AMP produced effects similar to those of cyclic AMP itself. In contrast, 8-[4-chlorophenylthio] cyclic AMP also showed a dual action, but with a high level of activation at all concentrations tested up to 1mm. Ca-NS+ channel activity was also predominantly activated by low concentrations of Sp-cAMPS. The activating effects of both Sp-cAMPS and cyclic AMP are antagonized by Rp-cAMPS, which by itself only produced a weak inhibition of Ca-NS+ channel activity even at concentrations of 10 μm and above. The results are discussed in terms of a model in which cyclic AMP, and other cyclic nucleotides, modulate the activity of the Ca-NS+ channel by binding to two separate sites.
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  • 24
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    The journal of membrane biology 146 (1995), S. 73-84 
    ISSN: 1432-1424
    Keywords: Gustation ; Taste ; Sensory transduction ; Sodium current ; Patch clamp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Voltage-dependent sodium currents were analyzed in detail from dissociated mammalian taste receptor cells using the whole-cell patch clamp technique. Approximately 50–75% of all taste receptor cells expressed sodium currents. These currents activated close to −50 mV (holding potential = −80 mV) with maximal currents most often occurring at −10 mV. The distribution of maximal inward currents across all cells appeared to display two peaks, at −254 pA and −477 pA, possibly due to differences in sodium channel density. Inward currents were eliminated by replacing 90% of external sodium with N-methyl-D-glucamine. The currentvoltage relationship of the activated current, as measured by a tail current analysis, was linear, suggesting an ohmic nature of the open channel conductance. The relationship between the time to the peak activated current and the step potential was well fit by a double exponential curve (τ1 = 6.18, τ2 = 37.8 msec). Development of inactivation of the sodium current was dependent upon both voltage- and temporal-parameters. The voltage dependence of the time constant (τ) obtained from removal of inactivation, development of inactivation, and decay of the sodium current displayed a bell-shaped curve with a maximum of 55 msec at −70 mV. In addition to fast inactivation (half maximal at −50 mV), these currents also displayed a slow inactivation (half maximal at −65 mV). Voltage-dependent sodium currents were reversibly inhibited by nanomolar concentrations of tetrodotoxin (Kd = 10−8 m). There was no evidence of a TTX-insensitive sodium current. This description broadens our understanding of gustatory transduction mechanisms with a particular relevance to the physiological role of receptor cell action potentials.
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  • 25
    ISSN: 1432-1424
    Keywords: Alpha toxin ; Ion channel ; Lettre cell ; Patch clamp ; Planar bilayer ; Staphylococcus aureus
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    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The conductance of pores induced by Staphylococcus aureus α-toxin in Lettre cells has been compared to that in bilayers composed of synthetic lipids or Lettre cell membrane constituents. Previously described characteristics of toxin-induced conductance changes in lipid bilayers, namely rectification, voltage-dependent closure, and closure at low pH or in the presence of divalent cations (Menestrina, 1986) are displayed also in bilayers prepared from Lettre cell membranes and in patch clamped Lettre cells. It is concluded that endogenous proteins do not affect the properties of α-toxininduced channels significantly and that the relative lack of ion channels in Lettre cells makes them ideal for studies of pore-forming toxins by the patch clamp technique.
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  • 26
    ISSN: 1432-1424
    Keywords: Cortical collecting duct ; Flufenamic acid ; Amiloride ; Adenine nucleotides ; cGMP dependent protein kinase ; Patch clamp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract We recently reported that M-1 mouse cortical collecting duct cells show nonselective cation (NSC) channel activity (Proc. Natl. Acad. Sci. USA 89:10262–10266, 1992). In this study, we further characterize the M-1 NSC channel using single-channel current recordings in excised inside-out patches. The M-1 NSC channel does not discriminate between Na+, K+, Rb+, Cs+, and Li+. It has a linear I-V relation with a conductance of 22.7±0.5 pS (n=78) at room temperature. The Pcation/ Panion ratio is about 60 and there is no measurable conductance for NMDG, Ca2+, Ba2+, and Mn2+. Cytoplasmic calcium activates the M-1 NSC channel at a threshold of 10−6 m and depolarization increases channel activity (NP o ). Cytoplasmic application of adenine nucleotides inhibits the M-1 NSC channel. At doses of 10−4 m and 10−3 m, ATP reduces NP o by 23% and 69%, respectively. Furthermore, since ADP (10−3 m) reduces NP o by 93%, the inhibitory effect of adenine nucleotides is not dependent on the presence of a γ-phosphoryl group and therefore does not involve protein phosphorylation. The channel is not significantly affected by 8-Br-cGMP (10−4 m) or by cGMP-dependent protein kinase (10−7 m) in the presence of 8-Br-cGMP (10−5 m) and ATP (10−4 m). The NSC channel is not sensitive to amiloride (10−4 m cytoplasmic and/or extracellular) but flufenamic acid (10−4 m) produces a voltage-dependent block, reducing NP o by 35% at depolarizing voltages and by 80% at hyperpolarizing voltages. We conclude that the NSC channel of M-1 mouse cortical collecting duct cells belongs to an emerging family of calcium-activated and nucleotide-sensitive nonselective cation channels. It does not contribute to amiloride-sensitive sodium absorption and is unlikely to be a major route for calcium entry. The channel is normally quiescent but may be activated under special physiological conditions, e.g., during volume regulation.
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  • 27
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    The journal of membrane biology 146 (1995), S. 59-71 
    ISSN: 1432-1424
    Keywords: Plantago media ; Whole cell ; Root cells ; Channel kinetics ; Patch clamp ; Hodgkin-Huxley
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Two different, simultaneously activated outward rectifying K+ currents were analyzed in the plasmalemma of root cortex protoplasts of Plantago media. Their gating is dependent on the diffusion potential for K+ (E K ).The threshold potential was more negative than E K allowing small inward currents at potentials below E K thereby keeping cells with little pump activity in the K state (Vogelzang & Prins, 1994). Time and voltage dependence of the outward rectifying K+ currents have been analyzed with Hodgkin-Huxley-like (HH) models. Dynamic responses of whole cell currents to pulse potentials were analyzed with two voltage dependent functions, the Boltzmann distribution for open probability per gate and the transition rate towards the open state (α). The transition rate in the opposite direction (β), was calculated from a and the Boltzmann distribution. These functions were used for an integral analysis of activation and deactivation currents measured over a range of pulse potentials. Both whole cell and single channel data were used for the determination of the number of closed and open states. The effects of single channel flickering on time response and amplitude of tail currents were added to the model. The dominant K+ channel present in the plasmalemma of P. media has a characteristic nonlinear single channel I-V curve reducing the amplitude of whole cell currents at positive potentials. To compensate for this nonlinearity, a four state translocator model was added to the whole cell open probability model. The analysis presented here provides a general basis for the study and comparison of K+ channel kinetics in plant protoplasts.
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  • 28
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    The journal of membrane biology 146 (1995), S. 273-282 
    ISSN: 1432-1424
    Keywords: Anion channel GCAC1 ; Guard cells ; Patch clamp ; Voltage-dependent kinetics ; Hodgkin-Huxley equations
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract A quantitative analysis of the time- and voltage-dependent kinetics of the guard cell anion channel (GCAC1) current in guard cell protoplasts from Vicia faba was analyzed using the whole-cell patch clamp technique. The voltage-dependent steady-state activation of GCAC1 current followed a Boltzmann distribution. For the corresponding steady-state value of the activation variable a power of two was derived which yielded suitable fits of the time course of voltage-dependent current activation. The GCAC1 mediated chloride current could successfully be described in terms of the Hodgkin-Huxley equations commonly evoked for the Na channel in nerve. After step depolarizations from a potential in the range of the resting potential to potentials above the equilibrium potential for chloride an activation and also an inactivation could be described. The gating of both processes exhibited an inverse relationship on the polarity of the applied step potentials in the order of milliseconds. Deactivating tail currents decline exponentially. The presented analysis contributes to the understanding of the rising phase of the observed action potentials in guard cells of V. faba. Evidence is presented that the voltage-dependent kinetic properties of the GCAC1 current are different from those properties described for the excitable anion currents in the plasmalemma of Chara corallina (Beilby & Coster, 1979a).
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  • 29
    ISSN: 1432-1424
    Keywords: Potassium channel ; Patch clamp ; Cyto-skeleton ; Cytokine ; Mechanosensitivity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract A variety of stimuli, including cytokines and adhesion to surfaces and matrix proteins, can regulate macrophage function, in part through changes in Ca2+-dependent second messengers. While fluctuation in in-tracellular Ca2+ is an important modulator of cellular activation, little attention has been paid to the roles of other ions whose cytoplasmic concentrations can be rapidly regulated by ion channels. To examine the role of ion channels in macrophage function, we undertook patch clamp studies of human culture-derived macrophages grown under serum-free conditions. The major ionic current in these cells was carried by an outwardly rectifying K+ channel, which had a single-channel conductance of 229 pS in symmetrical K+-rich solution and macroscopic whole-cell conductance of 9.8 nS. These channels opened infrequently in resting cells but were activated immediately by (i) adhesion of mobile cells onto a substrate, (ii) stretch applied to isolated membrane patches in Ca2+-free buffers, (iii) intracellular Ca2+ (EC50 of 0.4 μm), and (iv) the cytokine IL-2. Furthermore, barium and 4-aminopyridine, blockers of this channel, altered the organization and structure of the cytoskeletal proteins actin, tubulin and vimentin. These cytoskeletal changes were associated with reversible alteration to the morphology of the cells. Thus, we have identified an outwardly rectifying K+ channel that appeared to be involved in cytokine and adherence-mediated macrophage activation, and in the maintenance of cytoskeletal integrity and cell shape.
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  • 30
    ISSN: 1432-1424
    Keywords: ATP receptor ; G protein ; Patch clamp ; Ca2+-permeable channels ; Rat macrophages
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    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Outside-out configuration of the patch clamp technique was used to test whether an intracellular application of G protein activator (GTPγS) affects ATP-activated Ca2+-permeable channels in rat macrophages without any agonist in the bath solution. With 145 mm K+ (pCa 8.0) in the pipette solution, activity of channels permeable to a variety of divalent cations and Na+ was observed and general channel characteristics were found to be identical to those of ATP-activated ones. Absence of extracellular ATP makes it possible to avoid the influence of ATP receptor desensitization and to study the channel selectivity using a number of divalent cations (105 mm) and Na+ (145 mm) as the charge carriers. Permeability sequence estimated by extrapolated reversal potential measurements was: Ca2+ ∶ Ba2+ ∶ Mn2+ ∶ Sr2+ ∶ Na+ ∶ K+ = 68 ∶ 30 ∶ 26 ∶ 10 ∶ 3.5 ∶ 1. Slope conductances (in pS) for permeant ions rank as follows: Ca2+ ∶ Sr2+ ∶ Na+ ∶ Mn2+ ∶ Ba2+ = 19 ∶ 18 ∶ 14 ∶ 12 ∶ 10. Unitary Ca2+ currents display a tendency to saturate with the Ca2+ concentration increase with apparent dissociation constant (K d ) of 10 mm. No block of Na+ permeation by extracellular Ca2+ in millimolar range was found. The data obtained suggest that (i) activation of some G protein is sufficient to gate the channels without the ATP receptor being occupied, (ii) the ATP receptor activation results in the gating of a special channel with the properties that differ markedly from those of the receptoroperated or voltage-gated Ca2+-permeable channels on the other cell types.
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  • 31
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    The journal of membrane biology 148 (1995), S. 193-202 
    ISSN: 1432-1424
    Keywords: Baculovirus ; Expression ; Ion channel ; Desensitization ; Patch clamp ; Rapid perfusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract α1 and β1 subunits of human GABAA receptors were expressed in Sf9 cells using the Sf9-baculovirus system. Better expression was obtained by manipulating the system. Cell growth phase at the time of infection determined the practical range of virus titre, the period postinfection during which cells were useful for signal detection and the maximal current obtained. Cells in the early exponential phase were relatively insensitive to multiplicity of infection (MOI) whereas cells in the midto late-exponential phase were highly dependent on MOI and they responded with the largest Cl−} current generated by GABA. Channels activated by GABA were chloride-selective. Half the maximum peak whole-cell current was obtained with 11 μm GABA. The time course of Cl−} currents activated by saturating GABA concentrations in cells infected with α1β1recombinant viruses was examined employing a rapid perfusion system which allowed whole-cell solution exchange in less than 1 msec. The current decay could be fitted by 3 to 4 exponentials for the first 8 sec. The initial fast current decrease had a time constant of about 23 msec. No voltage dependence of time constants was detected but the whole-cell IV relation showed outward rectification. Currents were depressed by bicuculline, penicillin and picrotoxin and potentiated by pentobarbitone.
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  • 32
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    The journal of membrane biology 148 (1995), S. 255-262 
    ISSN: 1432-1424
    Keywords: K channel ; Heterologous expression ; Electrical development ; Muscle ; Patch clamp ; Embryo ; Xenopus laevis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The development of excitable cells is characterized by highly organized patterns of expression of ion channels. During the terminal differentiation Xenopus muscle somites, potassium currents are expressed first just after Stage 15 (early-mid neurula), following a long period during which no voltage-dependent currents can be detected in any cell in the dorsal embryo. We have investigated whether early expression of a foreign delayed rectifier potassium channel may affect this endogenous pattern of electrical development. We injected the purified cRNA of the mammalian brain Shaker-like potassium channel, Kv1.1, into fertilized Xenopus eggs. The resulting currents were analyzed in blastomeres during a 12-hr period prior to Stage 15 and in differentiating muscle cells after Stage 15. In injected embryos, a high fraction of blastomeres expressed a delayed rectifier-type current. The Kv1.1 current could be distinguished from the endogenous muscle delayed potassium current (I K,X) by its very different voltage dependence. Separation of currents based on this difference indicated that, in injected embryos, I K,X appeared much earlier in development than in control embryos. Furthermore, even in cells which expressed solely Kv1.1-type current, the sensitivity of the current to dendrotoxin declined dramatically during development, approaching that of I K,X . These data suggest an interaction between Kv1.1 and endogenous channel subunits, and/or modification of the Kv1.1 protein by the embryonic cells in ways not seen in Xenopus oocytes or mammalian cell lines.
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  • 33
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    The journal of membrane biology 144 (1995), S. 257-265 
    ISSN: 1432-1424
    Keywords: Paramecium ; Patch clamp ; Calmodulin ; Ion channels ; Channel regulation ; Ca2+ sensor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Paramecium Na+ channels, which were Ca2+-calmodulin activated, were studied in the inside-out mode of patch clamp. After excision of the membrane patch, they were active in the presence of 10−5 to 10−3 m Ca2+ in the bath. They became much less active in the presence of 10−6 m Ca2+, and their activity subsided completely at 10−8 m Ca2+. A Hill plot showed a dissociation constant of 6 μm for Ca2+ binding. This dissociation constant shifted to a submicromolar range in the presence of 1 mm Mg2+. The channels also exhibited a mild voltage dependence. When exposed to 10−8 m Ca2+ for an extended period of 2–4 min, channels were further inactivated even after bath Ca2+ was restored to 10−4 m. Whereas neither high voltage (+100 mV) nor high Ca2+ (10−3 m) was effective in reactivation of the inactive channels, addition of Paramecium wild-type calmodulin together with high Ca2+ to the bath restored channel activity without a requirement of additional Mg2+ and metabolites such as ATP. The channels reactivated by calmodulin had the same ion conductance, ion selectivity and Ca2+ sensitivity as those prior to inactivation. These inactivation and reactivation of the channels could be repeated, indicating that the direct calmodulin effect on the Na+ channel was reversible. Thus, calmodulin is a physiological factor critically required for Na+ channel activation, and is the Ca2+ sensor of the Na+-channel gating machinery.
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  • 34
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    The journal of membrane biology 146 (1995), S. 253-261 
    ISSN: 1432-1424
    Keywords: Nuclear pore complex ; Nuclear ion channels ; Gene activity ; Control of gene expression ; Transcription factors ; Oncogenes ; Proto-oncogenes ; AP-1 ; c-Jun ; NF-κB ; SP1 ; Patch clamp ; Cardiac myocytes ; Cell nucleus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Transcription factors (TFs) are cytoplasmic proteins that play an essential role in gene expression. These proteins form multimers and this phenomenon is thought to be one of the mechanisms that regulate transcription. TF molecules reach their DNA binding sites through the large central channel of the nuclear pore complex (NPC). However, the NPC channel is known to restrict the translocation of molecules ⩾20–70 kD. Therefore, during their translocation, TF molecules and/or their multimers may plug the NPC channel and thus, interrupt ion flow through the channel, with a concomitant reduction in the ion conductance of the channel (γ). Here we show with patch clamp that γ is reduced during translocation of three major TFs: c-Jun (40 kD), NF-κB (≈50 kD), and SP1 (≈100 kD). Within a minute, femtomolar concentrations of these proteins reduced γ suggesting a purely mechanical interaction between single TF molecules and the inner wall of the NPC channel. NPCs remained plugged for 0.5–3 hr in the absence of ATP but when ATP was added, channel plugging was shortened to 〈5 min. After unplugging, channel closures were rarely observed and the number of functional channels increased. The transcription factors also stabilized the NPCs as shown by the extended duration of the preparations which allowed recordings for up to 72 hr. These observations are the first direct demonstration of the important role of NPCs in mediating nuclear translocation of TFs and, therefore, in forming part of the mechanisms regulating gene expression. The studies also demonstrate the potential of the patch clamp technique in quantifying TF translocation to the nucleus, mRNA export, and other processes governing gene expression.
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  • 35
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    The journal of membrane biology 146 (1995), S. 239-251 
    ISSN: 1432-1424
    Keywords: Nuclear pore complex ; Nuclear ion channels ; Gene activity ; Control of gene expression ; Patch clamp ; Cardiac myocytes ; Cell nucleus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Macromolecule-conducting pores have been recently recognized as a distinct class of ion channels. The poor role of macromolecules as electrical charge carriers can be used to detect their movement along electrolyte-filled pores. Because of their negligible contribution to electrical ion currents, translocating macromolecules reduce the net conductivity of the medium inside the pore, thus decreasing the measured pore ion conductance. In the extreme case, a large translocating macromolecule can interrupt ion flow along the pore lumen, reflected as a negligible pore conductance. Therefore, ion conductance serves as a measurement of macromolecular transport, with lesser values indicating greater macromolecular translocation (in size and/or number). Such is the principle of operation of the widely used Coulter counter, an instrument for counting and sizing particles. It has long been known that macromolecules translocate across the central channel of nuclear pore complexes (NPCs). Recently, large conductance ion channel activity (100–1000 pS) was recorded from the nuclear envelope (NE) of various preparations and it was suggested that NPCs may be the source of this activity. Despite its significance to understanding the regulation of transcription, replication, mRNA export, and thus gene expression of normal and pathological states, no report has appeared demonstrating that this channel activity corresponds to ion flow along the central channel of the NPC. Here we present such a demonstration in adult mouse cardiac myocyte nuclei. In agreement with concepts introduced for macromolecule-conducting channels, our patch clamp experiments showed that ion conductance is reduced, and thus that ion flow is restricted during translocation of macromolecules containing nuclear targeting signals. Ion flow was blocked by mAb414, a monoclonal antibody raised against a major NPC glycoprotein and known to localize on the NPC channel where it blocks macromolecular transport. These results also establish patch clamp as a useful technique for the measurement of macromolecular translocation along the large central channel of the NPC and provide a basis for the design of future investigations of nuclear signaling for control of gene activity, mRNA export for gene expression, as well as other processes subservient to NPC-mediated nucleocytoplasmic exchange.
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  • 36
    ISSN: 1432-1424
    Keywords: Nuclear pore complex ; Nuclear ion channels ; Gene activity ; Control of gene expression ; TATA-binding protein ; TBP ; Patch clamp ; Atomic force microscopy ; Cell nucleus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The universal TATA-binding protein, TBP, is an essential component of the multiprotein complex known as transcription factor IID (TFIID). This complex, which consists of TBP and TBP-associated factors (TAFs), is essential for RNA polymerase II-mediated transcription. The molecular size of human TBP (37.7 kD) is close to the passive diffusion limit along the transport channel of the nuclear pore complex (NPC). Therefore, the possibility exists that NPCs restrict TBP translocation to the nuclear interior. Here we show for the first time, with patch-clamp and atomic force microscopy (AFM), that NPCs regulate TBP movement into the nucleus and that TBP (10−15–10−10 m) is capable of modifying NPC structure and function. The translocation of TBP was ATP-dependent and could be detected as a transient plugging of the NPC channels, with a concomitant transient reduction in single NPC channel conductance, γ, to a negligible value. NPC unplugging was accompanied by permanent channel opening at concentrations greater than 250 pm. AFM images demonstrated that the TBP molecules attached to and accumulated on the NPC cytosolic side. NPC channel activity could be recorded for more than 48 hr. These observations suggest that three novel functions of TBP are: to stabilize NPC, to force the NPC channels into an open state, and to increase the number of functional channels. Since TBP is a major component of transcription, our observations are relevant to the understanding of the gene expression mechanisms underlying normal and pathological cell structure and function.
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  • 37
    ISSN: 1432-1424
    Keywords: Regulatory volume decrease ; Slowly activating K+ channel ; Vestibular Labyrinth ; Vibrating probe ; Micro-Ussing chamber ; Patch clamp
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Volume regulation of vestibular dark cells from the gerbilline inner ear in response to a hypoosmotic challenge depends on the presence of cytosolic K+ and Cl−. The present study addresses the questions: (i) whether and by what mechanism K+ is released during volume regulation, (ii) whether the osmolarity of the basolateral medium has an effect on the steady-state rate of transepithelial K+ transport and (iii) whether there is cross-talk between the basolateral membrane responsible for K+ uptake and the apical membrane responsible for K+ release. K+ secretion $$(J_{{\text{K}}^{\text{ + }} ,{\text{probe}}} )$$ and current density (I sc,probe) were measured with vibrating probes in the vicinity of the apical membrane and the transepithelial potential (V t) and resistance (R t) were measured in a micro-Ussing chamber. The equivalent short-circuit current (I sc) was calculated. The current (I IsK), conductance (g IsK) and inactivation time constant (τIsK) of the I sK channel and the apparent reversal potential of the apical membrane (V r) were obtained with the cell-attached macropatch technique. V rwas corrected (V rc) for the membrane voltage (V m) measured separately with microelectrodes. A hypo-osmotic challenge (294 to 154 mosm by removal of 150 mm mannitol) on the basolateral side of the epithelium increased $$J_{K^{\text{ + }} ,{\text{probe}}} $$ and I sc,probe by a factor of 2.7 and 1.6. When this hypo-osmotic challenge was applied to both sides of the epithelium V tand I sc increased from 5 to 14 mV and from 189 to 824 μA/cm2 whereas R tdecreased from 27 to 19 Ω-cm2. With 3.6 mm K+ in the pipette I IsK was outwardly directed, τIsK was 267 msec and the hypo-osmotic challenge caused I IsK and g IsK to increase from 14 to 37 pA and from 292 to 732 pS. V rc hyperpolarized from −44 to −76 mV. With 150 mm K+ in the pipette I IsK was inwardly directed, τIsK was 208 msec and the hypo-osmotic challenge caused I IsK and g IsK to increase in magnitude from 0 to −21 pA and from 107 to 1101 pS. V rc remained unchanged (−2 vs. 1 mV). These data demonstrate that a hypo-osmotic challenge stimulates transepithelial K+ secretion and activates the apical I IsK channel. The hypoosmotically-induced increase in K+ secretion exceeded the estimated amount of K+ release necessary for the maintenance of constant cell volume, suggesting that the rate of basolateral K+ uptake was upregulated in the presence of the hypo-osmotic challenge and that cross-talk exists between the apical membrane and the basolateral membrane.
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  • 38
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    The journal of membrane biology 148 (1995), S. 143-156 
    ISSN: 1432-1424
    Keywords: Anion channel ; Channel gating ; Patch clamp ; Thylakoid membrane ; Nitellopsis obtusa
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Measurements of single channel currents were performed on isolated membrane patches from osmotically swollen thylakoids of the Charophyte alga Nitellopsis obtusa. A channel with a high selectivity for anions over cations and a conductance of 100 to 110 pS (114 mM Cl−) was revealed. The channel has a bells-haped voltage-dependence of the open probability, with a maximum at about 0 mV. This dependence was explained by two gating processes, one causing channel closure at positive and one at negative potentials. The steepness of the voltage-dependence corresponded to approximately 2 elementary charges to be transferred across the entire membrane in each of the two gating processes. The analysis of the anion channel kinetics in the millisecond time domain revealed an e-fold increase of mean open and decrease of mean closed times when the membrane voltage was made more positive by 20 and 36 mV, respectively. Concert transitions of two identical anion channels between open and long inactivated states were observed, while the millisecond closed-open transitions of the two channels within a burst of activity were kinetically independent.
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  • 39
    ISSN: 1573-8353
    Keywords: NMR ; 1H NMR ; 13C NMR ; 17O NMR ; 15N NMR ; aziridine-carboxylates ; configurational analysis ; conformational analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract A confrgurational and conformational study of NH, N-acetyl- and N-sulfonylaziridine carboxylates is performed by1H ,13C,17O, and15N NMR spectroscopy. The presence of acetyl and su fonyl groups on the ring nitrogen atom seems to reduce greatly the configurational stability at nitrogen.
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  • 40
    ISSN: 1573-904X
    Keywords: xanthonolignoids ; xanthones ; rat hepatocytes ; hepatoprotective activity ; tert-butylhydroperoxide ; lipid peroxidation ; glutathione
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Purpose. Synthesize and evaluate the protective activity against tert-butylhydroperoxide-induced toxicity in freshly isolated rat hepatocytes of trans-kielcorin, trans-isokielcorin B, as well as their respective building blocks 3,4-dihydroxy-2-methoxyxanthone and 2,3-dihydroxy-4-methoxyxanthone. Methods. Wistar rats, weighing 200-250g were used. Hepatocyte isolation was performed by collagenase perfusion. Incubations were performed at 37°C, using 1 million cells per milliliter in modified Krebs—Henseleit buffer. The protective activity was evaluated by measuring reduced and oxidized glutathione, lipid peroxidation and cell viability after inducing toxicity with tert-butylhydroperoxide (1.0 mM, 30 min), with or without the studied compounds in the concentrations of 0.025, 0.050, 0.100 and 0.200 mM. Silybin was tested in the same experimental conditions to serve as a positive control. Results. Using these concentrations, the tested compounds prevented tert-butylhydroperoxide-induced lipid peroxidation and cell death in freshly isolated rat hepatocytes. All compounds were also effective in preventing perturbation of cell glutathione homeostasis in some extent. 3,4-Dihydroxy-2-methoxyxanthone and 2,3-dihydroxy-4-methoxyxanthone were more effective than trans-kielcorin and trans-isokielcorin B respectively. Silybin was less effective in protecting cells against lipid peroxidation and loss of cell viability than the four xanthonic derivatives. Conclusions. The tested compounds protected the freshly isolated rat hepatocytes against tert-butylhydroperoxide-induced toxicity.
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  • 41
    ISSN: 1573-904X
    Keywords: 2′,3′,5′-triacetyl-6-azauridine ; prodrug ; hydrolysis ; pH-profile ; arrhenius plots ; CI-MS ; NMR ; liquid chromatography
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Purpose. The purposes were to study the kinetics of hydrolysis of 2′,3′,5′-triacetyl-6-azauridine ( 1 ) in aqueous solution (µ = 0.5) and to identify the main intermediates and products of the reaction. Methods. A stability indicating isocratic LC assay was used to study the rate of degradation of 1 A gradient LC assay was used to study the time courses of the degradants. The products of hydrolysis were isolated by preparative liquid chromatography and identified by 1H-NMR and CI-MS. The pKa value was obtained by potentiometric titration. Results. At 36.8°C, the pH-rate profile of 1 in water was adequately described by a four-term rate equation. The intermediates were identified as the primary and secondary di-acetates, and the primary and secondary mono-acetates. The final product was 6-azauridine. Conclusions. A simplified kinetic scheme could be used to describe the concentration-time profiles of 1, the intermediates and the final product.
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  • 42
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    Journal of chemical ecology 21 (1995), S. 1191-1215 
    ISSN: 1573-1561
    Keywords: Ourebia ourebi ; Bovidae ; mammalian semiochemicals ; mammalian pheromones ; exocrine secretion ; preorbital secretion ; dimethyl disulfide derivatization ; skipped dienes ; chemical-ionization mass spectrometry ; NMR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Using gas chromatography-mass spectrometry in conjunction with ancillary techniques such as chemical ionization with different reactant gases, determination of the position of double bonds by means of dimethyl disulfide derivatization, and finally gas chromatographic and mass spectrometric comparison with authentic synthetic material, 75 constituents were identified in the preorbital secretion of the male oribi,Ourebia ourebi. The secretion contains compounds with long-chain, unbranched structures similar to those found in many other preorbital secretions but with a finite volatility range, in contrast to the seemingly endlessly increasing chain lengths typical of other preorbital secretions.
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  • 43
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    Journal of chemical ecology 21 (1995), S. 1271-1285 
    ISSN: 1573-1561
    Keywords: Allelochemicals ; lipid peroxidation ; sulfhydryl groups ; leakage ; plasma membrane ; superoxide dismutase ; catalase ; peroxidase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Benzoic (BEN) and cinnamic (CIN) acids are commonly found in soils and are considered as strong allelochemicals. Published information suggest that BEN and CIN and other phenolic acids decrease plant growth in part by suppressing nutrient absorption. However, studies on the mechanism of action were not conclusive. We examined the effects of BEN and CIN on the cell plasma membrane in intact soybean (Glycine max L. cv. Maple Bell) seedlings. Treating intact root systems with BEN or CIN rapidly increased electrolyte leakage and ultraviolet absorption of materials into the surrounding solution. After 12 hr of treatment, BEN and CIN lowered the extracellular sulfhydryl group content in roots. The two allelochemicals induced lipid peroxidation, which resulted from free radical formation in plasma membranes, inhibition of catalase and peroxidase activities, and sulfhydryl group depletion. Oxidation or cross-linking of plasma membrane sulfhydryl groups is the first mode of action of both compounds. The BEN- and CIN-induced decrease in soybean nutrient absorption may be a consequence of damage to cell membrane integrity caused by a decrease in sulfhydryl groups followed by lipid peroxidation.
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  • 44
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    International journal of peptide research and therapeutics 2 (1995), S. 121-124 
    ISSN: 1573-3904
    Keywords: Backbone cyclization ; Substance P ; NMR ; Molecular dynamics ; Conformational analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Structure-activity relationships of cyclic substance P (SP) analogs were extensively studied in our laboratories utilizing the new concept of backbone cyclization. Employing the C-terminal hexapeptide SP6–11, we examined the influence of chemical changes in peptides containing backbone-to-amino-end cyclization. These changes in the ring have significant influence on activity, and should be carefully designed in order to optimize pharmacological feature.
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  • 45
    ISSN: 1573-3904
    Keywords: Biosurfactant ; Lipopeptide ; NMR ; Hydrophobic substitution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Bacillus subtilis coproduces several surfactin variants that are powerful biosurfactants and have potential applications in biology and industry. A single amino acid substitution in the heptapeptide moiety of surfactins strongly modifies their properties. To better establish structure-activity relationships and to search new variants with enhanced properties, Bacillus subtilis was grown into two modified culture media. Two new variants were isolated by chromatographic methods and studied by NMR spectroscopy. As planned, modifications consisted in the substitution of the l-valine residue at the fourth position by a more hydrophobic residue, i.e., leucine or isoleucine. These [Leu4]- and [Ile4]surfactins have a higher affinity for hydrophobic solvents and a twice improved surfactant power. Structure-property correlations were confirmed by analysis of the hydrophobic residue distribution in the three-dimensional model of the structure of surfactin in solution.
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  • 46
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    International journal of peptide research and therapeutics 2 (1995), S. 59-70 
    ISSN: 1573-3904
    Keywords: Opioid peptide ; NMR ; Enkephalins ; Micelles ; Dodecylphosphocholine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Opioid peptides are thought to interact with the cell membrane in their biological journey to the membrane-bound receptor. Both organic solvents and model membranes have been used previously to determine the stable solution conformations of peptide hormones. Leucine enkephalin has been studied in a number of different environments, but with limited resolution. Here it is shown that leucine enkephalin forms a stable type IV β-turn structure in dodecylphosphocholine micelles. We have observed a highly solvent-shielded amide proton with no evidence for a complementary hydrogen bond acceptor. The structural details of the peptide as determined by NMR spectroscopy in solution are described.
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  • 47
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    International journal of peptide research and therapeutics 2 (1995), S. 161-164 
    ISSN: 1573-3904
    Keywords: Cyclised peptide ; Dioxopiperazine ; Thiazolidine ; NMR ; Computer modelling
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Conformationally constrained peptidomimetics of the Lys-Pro sequence have been obtained using a new polycyclic structure. Bicyclic compounds containing a dioxopiperazine-thiazolidine fused ring have been synthesised starting from N-lysyl-4-ethoxycarbonylthiazolidine-2-carboxylic acid. The carboxyl group in either position 2 or 4 of the thiazolidine ring was reacted with the N-terminal amino group, giving different regioisomers. NMR and computer modelling were used to study the configuration of isomers and the lysine side-chain orientation with respect to the pseudoproline ring.
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  • 48
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    International journal of peptide research and therapeutics 2 (1995), S. 239-242 
    ISSN: 1573-3904
    Keywords: Pseudopeptide ; Triphosgene ; Beta-turn ; NMR ; FT-IR ; X-ray diffraction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary For the sake of improving synthetic methods and evaluating the conformational perturbation induced by the substitution of AzAsx for the Asx residue in the cognate dipeptide R-CO-Asx-Pro-NHR′, we prepared the AzAsx-dipeptide sequence by making use of the crystalline triphosgene. This reagent allows, in situ and under very mild experimental conditions, both the carbonylation and the activation of the properly substituted and N-protected hydrazine before coupling with the proline partner. With regard to conformational behaviour, the azadipeptide sequence displays a β-fold, unlike the cognate dipeptide which adopts an Asx-turn.
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