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A nitro radical anion formation from nifedipine: an electrochemical approach.

The cyclic voltammetric behaviour of nifedipine was studied. The addition of three aprotic solvents to nifedipine in an aqueous citrate buffer system was examined. Qualitatively they result in separation of the initial irreversible 4 electron reduction into two stages, the NO2/RNO2.- and RNO2.-, 4H/RNHOH, H2O couples, respectively. Particular attention was directed to the 1-electron RNO2/RNO2.- couple as measured by the cyclic voltammetric mode in mixed media. Analysis of the cyclic voltammetric response as a function of scan rate and non-aqueous solvent content yields information on the stability of the radical anion. The chemical forward reaction of the radical anion follows a second order kinetics with a stability constant of 1.1 x 10(-3) l mol-1 s-1 and a half-life time of 0.09 s for 1 mM of nifedipine in aqueous citrate buffer, pH 7.4/DMF; 50:50.

Electrochemistry↗

The simultaneous determination of neurotensin and its major fragments by on-line trace enrichment HPLC with electrochemical detection.

An HPLC assay using on-line cation exchange trace enrichment and acetonitrile gradient elution, ion pair reverse phase separation with electrochemical detection (EC) is described for the simultaneous determination of the tridecapeptide neurotensin (NT) and six of its fragments. Cyclic voltammetric analysis indicated that the oxidative electrochemical properties of NT and its fragments is not merely a function of the sum of its electroactive amino acids (i.e. tyrosine) but reflects the presence and association of other amino acids (e.g. the arginine-arginine pair at position 8-9). Using the described method, NT1-6, NT1-8, NT1-10, NT1-11, NT8-13, NT9-13 and NT1-13 were baseline resolved within 20 min with a limit of detection varying from 1 to 5 ng peptide/injection. Other structurally similar or quantitatively significant neuropeptides (e.g. substance P, somatostatin, bombesin) did not interfere. Initial application of this on-line trace enrichment HPLC-EC assay to the question of the molecular nature of NT in unprocessed human CSF indicated the predominance of NT1-13 with an apparent formation of NT1-8 and NT9-13 resulting from more vigorous sample preparation techniques. The improvements in assay specificity, signal-to-noise ratios, biomatrix compatibility and assayable sample volume compared to non-enrichment HPLC-EC are discussed.

Amino Acid Sequence↗

Calmodulin in mouse male germ cells: a qualitative and quantitative study.

Isolated male germ cells of the mouse possess a heat-stable stimulatory activity of Ca2+-dependent, calmodulin-free phosphodiesterase. Ionic exchange chromatography allowed partial purification of the activator and the isolation of multiple forms of phosphodiesterase stimulation inhibitor. The activator has been identified as calmodulin on the basis of chromatographic behaviour and electrophoretic mobility. Quantitative analysis showed variations of calmodulin levels at different stages of spermatogenesis. Quantitative analysis of cyclic nucleotide hydrolysis in germ cell cytosol showed that the activity of Ca2+-dependent phosphodiesterase is different in meiotic and post-meiotic mouse male germ cells. These data suggest that calcium-dependent pathway and a Ca2+-dependent regulation of cyclic nucleotides are present in developing germ cells.

Animals↗

An analysis of inhibitory effect of cyclic GMP on arachidonic acid- or collagen-induced aggregation in rabbit platelets.

Aggregation of washed rabbit platelets induced by arachidonic acid (AA) or collagen was inhibited by nitroprusside (NP) and 8-bromo cyclic GMP in a concentration-dependent manner. Although NP and 8-bromo cyclic GMP inhibited the AA-induced aggregation, these agents did not affect the conversion of exogenous AA to PG endoperoxides and TXA2 (which were observed as TXB2). On the other hand, collagen caused release of AA from phospholipids and sequential formation of TXB2 in [14C]AA prelabeled platelets. In contrast with the case in which exogenous AA was used, NP and 8-bromo cyclic GMP inhibited the collagen-induced formation of TXB2 by preventing the liberation of endogenous AA. These results indicate that cyclic GMP has at least two different inhibitory actions in platelets; one is the inhibition of AA release from phospholipids and the other is the inhibition of the action of TXA2 in platelets.

Animals↗

Excitatory amino acid antagonists, lamotrigine and BW 1003C87 as anticonvulsants in the genetically epilepsy-prone rat.

Sound-induced seizures in genetically epilepsy-prone rats were used to compare the anticonvulsant effect of phenytoin and diazepam with compounds which decrease glutamatergic neurotransmission including excitatory amino acid antagonists acting at N-methyl-D-aspartate (NMDA) receptors: D(-)CPPene, CGP 37849 and MK 801 or at the glycine/NMDA site: ACPC (1-aminocyclopropane-dicarboxylic acid) (partial agonist) or non-NMDA receptors: NBQX (2,3-dihydroxy-6-nitro-7-sulfamoylbenzo[f]-quinoxaline.Li) and GYKI 52466 (1-(aminophenyl)-4-methyl-7,8-methylene-dioxy-5H-2,3-benzodiazepin e.HCl) or acting at sodium channels to decrease glutamate release: lamotrigine and BW 1003C87 (5(2,3,5-trichlorophenyl)-2,4-diaminopyrimidine ethane sulphonate). ED50 values against clonic seizures (in mumol/kg at the time of peak anticonvulsant effect) were: phenytoin 30.5 (2 h), diazepam 0.5 (0.5 h), MK 801 0.01 (4 h), D(-)CPPene 1.9 (4 h), CGP 37849 2 (1 h), GYKI 52466 24 (0.25 h), NBQX 40 (0.5 h), ACPC 1053 (0.5 h), BW 1003C87 2.2 (1 h), lamotrigine 4.8 (4 h). BW 1003C87, lamotrigine, MK 801, phenytoin, diazepam and CGP 37849 had the most favourable therapeutic indices (rotarod locomotor deficit ED50/anticonvulsant ED50).

2-Amino-5-phosphonovalerate↗

Living patches engineered from human umbilical cord derived fibroblasts and endothelial progenitor cells.

OBJECTIVE: A major shortcoming in contemporary congenital heart surgery is the lack of viable replacement materials with the capacity of growth and regeneration. Here we focused on living autologous patches engineered from human umbilical cord derived fibroblasts and endothelial progenitor cells (EPCs) as a ready-to-use cell source for paediatric cardiovascular tissue engineering. METHODS: EPCs were isolated from 20 ml fresh umbilical cord blood by density gradient centrifugation and myofibroblasts were harvested from umbilical cord tissue. Cells were differentiated and expanded in vitro using nutrient media containing growth factors. Before seeding, cell-phenotypes were assessed by immuno-histochemistry. Biodegradable patches fabricated from synthetic polymers (PGA/P4HB) were seeded with myofibroblasts followed by endothelialization with EPCs. All patches were cultured in a perfusion bioreactor. A subgroup of patches was additionally stimulated by cyclic strain. Analysis of the neo-tissues comprised histology, immuno-histochemistry, extracellular matrix (ECM) analysis and biomechanical testing. RESULTS: Endothelial phenotypes of EPCs before seeding were confirmed by Ac-Dil-LDL, CD 31, von-Willebrand-Factor and eNOS staining. Histology of the seeded patches demonstrated layered viable tissue formation in all samples. The cells in the newly formed tissues expressed myofibroblast markers, such as desmin and alpha-SMA. The EPCs derived neo-endothelia showed constant endothelial phenotypes (CD 31, vWF). major constituents of ECM such as collagen and proteoglycans were biochemically detected. Stress-strain properties of the patches showed features of native-analogous tissues. CONCLUSIONS: Living tissue engineered patches can be successfully generated from human umbilical cord derived myofibroblasts and EPCs. This new cell source may enable the tissue engineering of versatile, living, autologous replacement materials for congenital cardiac interventions.

Biocompatible Materials↗

Modification of indium-tin oxide (ITO) glass with aziridine provides a surface of high amine density.

The surface of indium-tin oxide (ITO) substrates was successfully modified with aziridine. Modification of the surface was achieved through facile ring-opening, and hyperbranching polymerization of the ring-strained heterocycle initiated from the reactive group on the surface. Amine density of the aziridine-modified ITO measured with UV-vis spectrophotometry is 10 amines/nm2. Cyclic voltammetric analysis showed that the aziridine-modified electrode was less active for Ru(NH3)6(3+) in comparison with the pristine electrode, while no difference was observed for Fe(CN)6(4-). Electrochemical impedance spectroscopic experiments unveiled that the modified electrode was more efficient for electron transfer to the latter species than to the former.

Amines↗

Electrochemical treatment of Procion Black 5B using cylindrical flow reactor--a pilot plant study.

The paper presents the results of an efficient electrochemical treatment of Procion Black 5B--a pilot plant study. Experiments were conducted at different current densities and selected electrolyte medium using Ti/RuO2 as anode, stainless-steel as cathode in a cylindrical flow reactor. By cyclic voltammetric analysis, the best condition for maximum redox reaction rate was found to be in NaCl medium. During the various stages of electrolysis, parameters such as COD, colour, FTIR, UV-vis spectra studies, energy consumption and mass transfer coefficient were computed and presented. The experimental results showed that the electrochemical oxidation process could effectively remove colour and the chemical oxygen demand (COD) from the synthetic dye effluent. The maximum COD reduction and colour removal efficiencies were 74.05% and 100%, respectively. Probable theory, reaction mechanism and modeling were proposed for the oxidation of dye effluent. The results obtained reveal the feasibilities of application of electrochemical treatment for the degradation of Procion Black 5B.

Electrodes↗

A new format of electrodes for the electrochemical reduction of cytochromes P450.

New approach to the electrochemical reduction of cytochromes P450 (P450s, CYPs) at electrodes chemically modified with appropriate substrates for P450s ("reverse" electrodes) was proposed. The method is based on the analysis of cyclic voltammograms, square-wave voltammograms and amperograms with subsequent determination of electrochemical characteristics such as catalytic current and redox potential. The sensitivity of proposed method is 0.2-1 nmol P450/electrode. The changes of maximal current and of redox potentials in square-wave voltammograms as well as the changes of catalytic current in amperometric experiments proved to be informative and reliable. Planar regime of screen-printed electrodes (strip-type sensors) enabled to utilise 20-60 microl of electrolyte volume. The enzyme-substrate pairs P450 2B4/benzphetamine and P450scc/cholesterol were investigated. Electrochemical parameters of electrodes with unspecific P450 substrates differed considerably from electrodes with appropriate substrates.

Biosensing Techniques↗

Single cell analysis of activity-dependent cyclic AMP-responsive element-binding protein phosphorylation during long-lasting long-term potentiation in area CA1 of mature rat hippocampal-organotypic cultures.

Phosphorylation of the transcription factor cyclic AMP (cAMP)-response element-binding protein (CREB) has been implicated in long-term synaptic plasticity and memory, and its activation has been proposed to be required for the maintenance of long-term potentiation (LTP). The previously described temporal dynamics of CREB phosphorylation during the maintenance of LTP showed differences between experimental models. In the present study the level of CREB phosphorylation was evaluated in organotypic hippocampal slices from young adult rats (P25-30) after long-lasting LTP was induced. Immunohistochemistry and confocal imaging were used to determine the ratio between non-phosphorylated and phosphorylated CREB at a single cell resolution, revealing not only the temporal dynamics but also the extent of CREB phosphorylation. The activation of CREB after LTP-induction was compared with cAMP-activation after bath application of forskolin. An increase in cAMP by forskolin resulted in a persistent, uniform increase of the phosphorylated CREB (pCREB/CREB immunofluorescence ratio) in all hippocampal principal neurons. In contrast, the induction of long-lasting LTP in CA1 was accompanied by a local increase in the pCREB/CREB ratio. Both CREB activation and LTP induction in mature cultured slices required N-methyl-D-aspartate (NMDA) receptor activation. CREB phosphorylation continued to increase for 4 h during LTP maintenance. This sustained activation is in contrast to previous observations in acutely prepared slices and supports the hypothesis that CREB plays an important role during the late phases of LTP.

Animals↗

Dopamine enhances the phosphaturic effect of PTH during acute respiratory alkalosis.

The phosphaturic response to parathyroid hormone (PTH) is blunted during acute respiratory alkalosis. The objective of the present study was to determine the effect of dopamine on the blunted phosphaturic response to PTH during acute respiratory alkalosis. The phosphaturic response to PTH was determined in thyroparathyroidectomized (TPTX) normocapnic and respiratory alkalotic rats in the absence and presence of the infusion of exogenous dopamine (25 microg/kg/min) or of 3,4-dihydroxyphenylalanine (L-DOPA, 250 microg/kg/min) to increase endogenous dopamine synthesis. In normocapnic rats, PTH infusion (33 U/kg plus 1 U/kg/min) significantly increased the fractional excretion of phosphate (FE(Pi)), from 1.5%+/-0.5% to 28.4%+/-4.0%, (deltaFE(Pi) 26.9%+/-4.1%, n = 11, P<.05); in respiratory alkalotic rats, the increase was from 0.4%+/-0.1% to 11.4%+/-1.7% (deltaFE(Pi) 11.0%+/-1.8%, n = 13, P<.05). However, the phosphaturic response to PTH was attenuated in respiratory alkalotic rats (deltaFE(Pi) 26.9%+/-4.1% vs 11.0%+/-1.9%, P<.05). In normocapnic rats, in the presence of dopamine or L-DOPA infusions, PTH infusion significantly increased the FE(Pi) from 6.1%+/-2.3% to 33.4%+/-8.0% (deltaFE(Pi) 27.3%+/-7.0%, n = 5) and from 3.2%+/-0.6% to 32.5%+/-3.3% (deltaFE(Pi) 29.3%+/-3.2%, n = 7), respectively. In respiratory alkalotic rats, in the presence of dopamine infusion, PTH significantly increased the FE(Pi), from 0.6%+/-0.2% to 19.3%+/-3.3% (deltaFE(Pi) 18.7%+/-3.3%, n = 6); in the presence of L-DOPA infusion it increased from 1.0%+/-0.3% to 20.5%+/-2.8% (deltaFE(Pi) 19.5%+/-2.9%, n = 8, P<.05 as compared with PTH alone). Thus the phosphaturic effect of PTH that was attenuated in respiratory alkalotic rats was enhanced by stimulation of endogenous dopamine synthesis by the infusion of L-DOPA.

Acute Disease↗

Synthesis, characterization and antitumor activity of copper(II) complex with nicotinamido-4-bis(2-chloroethyl)aminobenzaldimine.

Nicotinamido-4-bis(2-chloroethyl)aminobenzaldimine (NBAB) was synthesized and characterized by elemental analysis, IR and 1H NMR spectra. The complex of Cu(NBAB)2(NO3)2 was prepared in ethanol and characterized by elemental analysis, conductivity, cyclic voltammetry, IR, UV-Vis, fluorescence, CD and EPR spectra. The characteristic data suggest that the complex has an elongated octahedral structure, and NBAB behaves as bidentate in the keto form. The antitumor activities of NBAB and the complex against L1210 murine leukemia and K562 were investigated with both the MTT method and the colony formation test. The results in vitro indicate that antitumor activities of NBAB are superior to 2,2'-chlorodiethylamine hydrochloride (nitrogen mustard) for L1210, and inferior to nitrogen mustard for K562, but the antitumor activities of the complex for both cell lines are superior to nitrogen mustard.

Animals↗

Synthesis, solution structure analysis and antibody binding of cyclic epitope peptides from glycoprotein D of Herpes simplex virus type I.

Two cyclic peptides with a thioether bond have been synthesised corresponding to the 9-22 (9LKMADPNRFRGKDL(22)) sequence of glycoprotein D (gD-1) of Herpes simplex virus. The role of the secondary structure in protein-specific monoclonal antibody recognition was investigated. The sequence selected for this study comprises a strongly antigenic site adopting a beta-turn at residues 14Pro-(15)Asn. Thioether bond was formed between the free thiol group of cysteine or homocysteine inserted in position 11 and the chloroacetylated side chain of lysine in position 18. We report here the preparation of cyclic peptides containing Cys or Hcy in position 11, differing only in one methylene group. The linear precursor peptides were synthesised by Boc/Bzl strategy on MBHA resin, and the cyclisation was carried out in alkaline solution. The secondary structure of the peptides was studied by CD, FT-IR and NMR spectroscopy. The CD and FT-IR data have revealed fundamental changes in the solution conformation of the two compounds. The CH(2) group difference significantly resulted in the altered turn structure at the 12Ala and 13Asp as identified by NMR spectroscopy. The antibody binding properties of the cyclopeptides studied by gD-specific monoclonal antibody (A16) in direct and competition enzyme-linked immunosorbent assay (ELISA) were also not the same. We found that peptide LK[HcyADPNRFK]GKDL exhibited higher affinity to Mab A16 than peptide LK[CADPNRFK]GKDL, however, their reactivity was significantly lower compared to the linear ones. Our results clearly show the importance of secondary structure in an antibody binding and demonstrate that even a slight modification of the primary structure dramatically could influence the immune recognition of the synthetic antigens.

Amino Acid Sequence↗

Changes of the power coefficient in the 'metabolism-mass' relationship in the evolutionary process of animals.

The power coefficient k decreases along evolution in an allometric relationship between the oxygen consumption rate and the body mass of animals. This theoretical study investigated the role of the power coefficient k and its behavior along evolution. The animals were organized in three groups according to the values of the power coefficient k as follows: (I) from unicellular Prokaryotes to Eukaryotes; (II) from Mytilus and Annelida to Pisces; (III) from Reptilia to Mammals and Aves. At the beginning of each animal group (stage), the value of k was close to 0.9-1.0 and at the end of the stage it was close to 0.67-0.70. Exponential sharp increase of the power coefficient k was observed during the biological transition from Protozoa to simply organized Metazoa and in the transition from Poikylothermic to Homothermic organisms (e.g. from Pisces to Reptilia). Also, when using the periodogram regression analysis, a cyclic (periodic) pattern in this increase was observed (i.e. period T approximately 8-11 units, P<0.05). It was postulated that the power coefficient k, as with the coefficient a, might represent the increase of complexity of animal organization within each group.

Animals↗

Synthesis and characterization of trans-[Os(en)2py(H)]2+ and related studies.

trans-[Os(en)(2)pyH](Otf)(2), 2, is recovered from an acidic solution of trans-[Os(en)(2)py(H(2))](OTf)(2), 1, which has been subjected to one electron oxidation. The structures of both 1 and 2 have been determined by single crystal X-ray analysis. In cyclic voltammetry, 2 shows a one electron oxidation wave at 0.95 V and a one electron reduction wave at -1.2 V, neither accompanied by a signal for the reverse process. Reduction of 2 by Zn/Hg in methanol solution leads to quantitative formation of [Os(en)(2)(py)H(2))](2+), predominantly in the trans-form. In aqueous solution, species 2 reacts rapidly with N-methylacridium ion, [MAH](+), by hydride transfer. One electron chemical oxidation of 2 to the corresponding Os(IV) is slower than that of 1 to 2 owing to the increase in coordination number when Os(IV) is produced. Treatment of 1, or the cis-form, 1', in DMSO by sodium t-butoxide produces mainly the corresponding isomers of the monohydrides of Os(II), that derived from 1' is deep red in color while the trans-monohydride is colorless. Both react with [MAH](+) to form [MAH](2), and both disappear rapidly in acetone or acetonitrile, presumably by reducing the solvents. Reaction of trans-[Os(NH(3))(4)(H(2))H(2)O](BPh(4))(2), 4, in acetone-d(6) as solvent with either CH(3)CHO or styrene leads to hydrogenation of the substrate. Reactions which compete with trans-[Os(en)(2)(eta(2)-H(2))(CF(3)SO)(3)]CF(3)SO(3) release of substrate from the trans-complex before isomerization to the cis-form, required for hydrogenation to occur, result in the trans-derivative of the added solute. When H(2)C=CH-CH(2)-SCH(3) is the substrate, binding takes place at sulfur. Complete conversion to the cis-substrate isomer is observed, without hydrogenation occurring even though contact between dihydrogen and the double bond is possible.

Journal Article↗

High-valent imido complexes of manganese and chromium corroles.

The oxidation reaction of M(tpfc) [M = Mn or Cr and tpfc = tris(pentafluorophenyl)corrole] with aryl azides under photolytic or thermal conditions gives the first examples of mononuclear imido complexes of manganese(V) and chromium(V). These complexes have been characterized by NMR, mass spectrometry, UV-vis, EPR, elemental analysis, and cyclic voltammetry. Two X-ray structures have been obtained for Mn(tpfc)(NMes) and Cr(tpfc)(NMes) [Mes = 2,4,6-(CH(3))(3)C(6)H(2)]. Short metal-imido bonds (1.610 and 1.635 Angstroms) as well as nearly linear M-N-C angles are consistent with triple M triple-bond NR bond formation. The kinetics of nitrene [NR] group transfer from manganese(V) corroles to various organic phosphines have been defined. Reduction of the manganese(V) corrolato complex affords phosphine imine and Mn(III) with reaction rates that are sensitive to steric and electronic elements of the phosphine substrate. An analogous manganese complex with a variant corrole ligand containing bromine atoms in the beta-pyrrole positions, Mn(Br(8)tpfc)(NAr), has been prepared and studied. Its reaction with PEt(3) is 250x faster than that of the parent tpfc complex, and its Mn(V/IV) couple is shifted by 370 mV to a more positive potential. The EPR spectra of chromium(V) imido corroles reveal a rich signal at ambient temperature consistent with Cr(V) triple-bond NR (d(1), S = 1/2) containing a localized spin density in the d(xy) orbital, and an anisotropic signal at liquid nitrogen temperature. Our results demonstrate the synthetic utility of organic aryl azides in the preparation of mononuclear metal imido complexes previously considered elusive, and suggest strong sigma-donation as the underlying factor in stabilizing high-valent metals by corrole ligands.

Journal Article↗

Substituent effects on nitrosyl iron corrole complexes Fe(Ar3C)(NO).

A series of nitrosyl tris(5,10,15-aryl)corrolate complexes of iron(III) Fe(Ar3C)(NO) with different substituents on the aryl groups have been prepared, and certain spectroscopic and reaction properties were compared. The cyclic voltammetric analysis of the various Fe(Ar3C)(NO) complexes demonstrated that both the one-electron oxidation and one-electron reduction potentials respond in systematic and nearly identical trends relative to the electron-donor properties of the substituents. A similar pattern was seen in the nitrosyl stretching frequency, nu(NO), which modestly decreased with the stronger donor substituents. Flash photolysis of Fe(Ar3C)(NO) solutions in toluene leads to NO dissociation followed by rapid [NO]-dependent decay of the transients formed (presumably Fe(Ar3C)) to regenerate the original spectra. As was seen in an earlier flash photolysis study of Fe(TNPC)(NO) (TNPC3- = 5,10,15-tris(4-nitro-phenyl)corrolate; Joseph, C.; Ford, P. C. J. Am. Chem. Soc. 2005, 127, 6737-6743), the second-order rate constants, k(NO), are all much faster ((1-9) x 10(8) M(-1) s(-1) at 298 K) than those for analogous iron(III) complexes of porphyrins. However, on a more microscopic level there is no obvious pattern in these rates with respect to the donor properties of the aryl ring substituents. The high reactivity of the ferric triarylcorrolates with NO data is interpreted in terms of the strongly electron-donating character of the Ar3C3- ligand and the quartet electronic configuration of the Fe(Ar3C) intermediate.

Electrochemistry↗

Iridium luminophore complexes for unimolecular oxygen sensors.

In this study, a series of novel luminescent cyclometalated Ir(III) complexes has been synthesized and evaluated for use in unimolecular oxygen-sensing materials. The complexes Ir(C6)(2)(vacac), 1, Ir(ppy)(2)(vacac), 2, fac-Ir(ppy)(2)(vppy), 3, and mer-Ir(ppy)(2)(vppy), 4, where C6 = Coumarin 6, vacac = allylacetoacetate, ppy = 2-phenylpyridine, and vppy = 2-(4-vinylphenyl)pyridine, all have pendent vinyl or allyl groups for polymer attachment via the hydrosilation reaction. These luminophore complexes were characterized by NMR, absorption, and emission spectroscopy, luminescence lifetime and quantum yield measurements, elemental analysis, and cyclic voltammetry. Complex 1 was structurally characterized using X-ray crystallography, and a series of 1-D ((1)H, (13)C) and 2-D ((1)H-(1)H, (1)H-(13)C) NMR experiments were used to resolve the solution structure of 4. Complexes 1 and 3 displayed the longest luminescence lifetimes and largest quantum efficiencies in solution (tau = 6.0 micros, phi = 0.22 for 1; tau = 0.4 micros, phi = 0.2 for 3) and, as result, are the most promising candidates for future luminescence-quenching-based oxygen-sensing studies.

Journal Article↗