Osmolytic Effect of Sucrose on Thermal Denaturation of Pea Seedling Copper Amine Oxidase.

PubMed

Amani, Mojtaba; Barzegar, Aboozar; Mazani, Mohammad

2017-04-01

Protein stability is a subject of interest by many researchers. One of the common methods to increase the protein stability is using the osmolytes. Many studies and theories analyzed and explained osmolytic effect by equilibrium thermodynamic while most proteins undergo an irreversible denaturation. In current study we investigated the effect of sucrose as an osmolyte on the thermal denaturation of pea seedlings amine oxidase by the enzyme activity, fluorescence spectroscopy, circular dichroism, and differential scanning calorimetry. All experiments are in agreement that pea seedlings amine oxidase denaturation is controlled kinetically and its kinetic stability is increased in presence of sucrose. Differential scanning calorimetry experiments at different scanning rates showed that pea seedlings amine oxidase unfolding obeys two-state irreversible model. Fitting the differential scanning calorimetry data to two-state irreversible model showed that unfolding enthalpy and T * , temperature at which rate constant equals unit per minute, are increased while activation energy is not affected by increase in sucrose concentration. We concluded that osmolytes decrease the molecular oscillation of irreversible proteins which leads to decline in unfolding rate constant.

Mutation at a strictly conserved, active site tyrosine in the copper amine oxidase leads to uncontrolled oxygenase activity.

PubMed

Chen, Zhi-Wei; Datta, Saumen; Dubois, Jennifer L; Klinman, Judith P; Mathews, F Scott

2010-08-31

The copper amine oxidases carry out two copper-dependent processes: production of their own redox-active cofactor (2,4,5-trihydroxyphenylalanine quinone, TPQ) and the subsequent oxidative deamination of substrate amines. Because the same active site pocket must facilitate both reactions, individual active site residues may serve multiple roles. We have examined the roles of a strictly conserved active site tyrosine Y305 in the copper amine oxidase from Hansenula polymorpha kinetically, spetroscopically (Dubois and Klinman (2006) Biochemistry 45, 3178), and, in the present work, structurally. While the Y305A enzyme is almost identical to the wild type, a novel, highly oxygenated species replaces TPQ in the Y305F active sites. This new structure not only provides the first direct detection of peroxy intermediates in cofactor biogenesis but also indicates the critical control of oxidation chemistry that can be conferred by a single active site residue.

SPERMINE OXIDASE: AN AMINE OXIDASE WITH SPECIFICITY FOR SPERMINE AND SPERMIDINE

PubMed Central

Hirsch, James G.

1953-01-01

Sheep serum and bovine serum contain an enzyme which brings about a rapid oxidative deamination of certain biological amines. This enzyme differs from previously described amine oxidases in several regards and especially in its substrate specificity. Studies thus far indicate that only spermine and the closely related compound spermidine serve as substrates for the enzyme in sheep serum. For this reason, the enzyme has been named spermine oxidase. Spermine oxidase is active in a variety of fluids of various ionic strength and buffer composition. The reaction takes place between pH 6.0 and pH 8.0 with an optimal rate in the vicinity of neutrality. Under certain conditions, the rate of oxygen consumption during the initial phase of the reaction is independent of the concentration of substrate. The diminution in rate observed during the latter phase of the enzymatic attack appears to be due to an alteration in the kinetics at low concentrations of substrate, or to competitive inhibition by a product of the reaction. Carbonyl reagents almost completely block the action of spermine oxidase, while certain amines and the cyanide ion bring about partial inhibition. Thiol reagents and sequestering compounds do not alter the course of the oxidative process. In the presence of low concentrations of mercuric chloride, the sheep serum-spermine system consumes approximately twice as much oxygen as controls containing no mercuric ion. The mechanism by which the mercuric ion stimulates additional oxygen uptake is obscure. PMID:13052805

A novel proteolytic processing of prolysyl oxidase

PubMed Central

Atsawasuwan, Phimon; Mochida, Yoshiyuki; Katafuchi, Michitsuna; Tokutomi, Kentaro; Mocanu, Viorel; Parker, Carol E.; Yamauchi, Mitsuo

2012-01-01

Lysyl oxidase (LOX) is an amine oxidase that is critical for the stability of connective tissues. The secreted proLOX is enzymatically quiescent and is activated through proteolytic cleavage between residue Gly162 and Asp163 (residue numbers according to the mouse LOX) by bone morphogenetic protein (BMP)-1 gene products. Here we report a novel processing of proLOX identified in vitro and in vivo. Two forms of mature LOX were identified and characterized by their immunoreactivity to specific antibodies, amine oxidase activity and mass spectrometry. One form was identified as a well characterized BMP-1 processed LOX protein. Another was found to be a truncated form of LOX (tLOX) resulting from the cleavage at the carboxy terminus of Arg192. The tLOX still appeared to retain amine oxidase activity. The results from the proLOX gene deletion and mutation experiments indicated that the processing occurs independent of the cleavage of proLOX by BMP-1 gene products and likely requires the presence of LOX propeptide. These results indicate that proLOX could be processed by two different mechanisms producing two forms of active LOX. PMID:21591931

A novel proteolytic processing of prolysyl oxidase.

PubMed

Atsawasuwan, Phimon; Mochida, Yoshiyuki; Katafuchi, Michitsuna; Tokutomi, Kentaro; Mocanu, Viorel; Parker, Carol E; Yamauchi, Mitsuo

2011-01-01

Lysyl oxidase (LOX) is an amine oxidase that is critical for the stability of connective tissues. The secreted proLOX is enzymatically quiescent and is activated through proteolytic cleavage between residues Gly(162) and Asp(163) (residue numbers according to the mouse LOX) by bone morphogenetic protein (BMP)-1 gene products. Here we report a novel processing of proLOX identified in vitro and in vivo. Two forms of mature LOX were identified and characterized by their immunoreactivity to specific antibodies, amine oxidase activity, and mass spectrometry. One form was identified as a well-characterized BMP-1 processed LOX protein. Another was found to be a truncated form of LOX resulting from the cleavage at the carboxy terminus of Arg(192). The truncated form of LOX still appeared to retain amine oxidase activity. The results from the proLOX gene deletion and mutation experiments indicated that the processing occurs independent of the cleavage of proLOX by BMP-1 gene products and likely requires the presence of LOX propeptide. These results indicate that proLOX could be processed by two different mechanisms producing two forms of active LOX.

Bienzyme biosensors for glucose, ethanol and putrescine built on oxidase and sweet potato peroxidase.

PubMed

Castillo, Jaime; Gáspár, Szilveszter; Sakharov, Ivan; Csöregi, Elisabeth

2003-05-01

Amperometric biosensors for glucose, ethanol, and biogenic amines (putrescine) were constructed using oxidase/peroxidase bienzyme systems. The H(2)O(2) produced by the oxidase in reaction with its substrate is converted into a measurable signal via a novel peroxidase purified from sweet potato peels. All developed biosensors are based on redox hydrogels formed of oxidases (glucose oxidase, alcohol oxidase, or amine oxidase) and the newly purified sweet potato peroxidase (SPP) cross-linked to a redox polymer. The developed electrodes were characterized (sensitivity, stability, and performances in organic medium) and compared with similarly built ones using the 'classical' horseradish peroxidase (HRP). The SPP-based electrodes displayed higher sensitivity and better detection limit for putrescine than those using HRP and were also shown to retain their activity in organic phase much better than the HPR based ones. The importance of attractive or repulsive electrostatic interactions between the peroxidases and oxidases (determined by their isoelectric points) were found to play an important role in the sensitivity of the obtained sensors.

Mapping the primary structure of copper/topaquinone-containing methylamine oxidase from Aspergillus niger.

PubMed

Lenobel, R; Sebela, M; Frébort, I

2005-01-01

The amino acid sequence of methylamine oxidase (MeAO) from the fungus Aspergillus niger was analyzed using mass spectrometry (MS). First, MeAO was characterized by an accurate molar mass of 72.4 kDa of the monomer measured using MALDI-TOF-MS and by a pI value of 5.8 determined by isoelectric focusing. MALDI-TOF-MS revealed a clear peptide mass fingerprint after tryptic digestion, which did not provide any relevant hit when searched against a nonredundant protein database and was different from that of A. niger amine oxidase AO-I. Tandem mass spectrometry with electrospray ionization coupled to liquid chromatography allowed unambiguous reading of six peptide sequences (11-19 amino acids) and seven sequence tags (4-15 amino acids), which were used for MS BLAST homology searching. MeAO was found to be largely homologous to a hypothetical protein AN7641.2 (EMBL/GenBank protein-accession code EAA61827) from Aspergillus nidulans FGSC A4 with a theoretical molar mass of 76.46 kDa and pI 6.14, which belongs to the superfamily of copper amine oxidases. The protein AN7641.2 is only little homologous to the amine oxidase AO-I (32% identity, 49 % similarity).

Bioconversion of Airborne Methylamine by Immobilized Recombinant Amine Oxidase from the Thermotolerant Yeast Hansenula polymorpha

PubMed Central

Sigawi, Sasi; Nitzan, Yeshayahu

2014-01-01

Aliphatic amines, including methylamine, are air-pollutants, due to their intensive use in industry and the natural degradation of proteins, amino acids, and other nitrogen-containing compounds in biological samples. It is necessary to develop systems for removal of methylamine from the air, since airborne methylamine has a negative effect on human health. The primary amine oxidase (primary amine : oxygen oxidoreductase (deaminating) or amine oxidase, AMO; EC 1.4.3.21), a copper-containing enzyme from the thermotolerant yeast Hansenula polymorpha which was overexpressed in baker's yeast Saccharomyces cerevisiae, was tested for its ability to oxidize airborne methylamine. A continuous fluidized bed bioreactor (CFBR) was designed to enable bioconversion of airborne methylamine by AMO immobilized in calcium alginate (CA) beads. The results demonstrated that the bioreactor with immobilized AMO eliminates nearly 97% of the airborne methylamine. However, the enzymatic activity of AMO causes formation of formaldehyde. A two-step bioconversion process was therefore proposed. In the first step, airborne methylamine was fed into a CFBR which contained immobilized AMO. In the second step, the gas flow was passed through another CFBR, with alcohol oxidase from the yeast H. polymorpha immobilized in CA, in order to decompose the formaldehyde formed in the first step. The proposed system provided almost total elimination of the airborne methylamine and the formaldehyde. PMID:24672387

New recombinant cyclohexylamine oxidase variants for deracemization of secondary amines by orthogonally assaying designed mutants with structurally diverse substrates

NASA Astrophysics Data System (ADS)

Li, Guangyue; Yao, Peiyuan; Cong, Peiqian; Ren, Jie; Wang, Lei; Feng, Jinhui; Lau, Peter C. K.; Wu, Qiaqing; Zhu, Dunming

2016-05-01

To further expand the substrate range of the cyclohexylamine oxidase (CHAO) from Brevibacterium oxydans, a library of diverse mutants was created and assayed toward a group of structurally diverse substrates. Among them, mutants T198A and M226A exhibited enhanced activity relative to wt CHAO for most (S)-enantiomers of primary amines and some secondary amines. While mutants T198I, L199I, L199F, M226I and M226T were more active than wt CHAO toward the primary amines, mutants T198F, L199T, Y321A, Y321T, Y321I and Y321F enhanced the enzyme activity toward the secondary amines. In particular, mutant Y321I displayed an enhanced catalytic efficiency toward 1-(4-methoxybenzyl)-1, 2, 3, 4, 5, 6, 7, 8-octahydroisoquinoline (13). Whereas a double mutant, Y321I/M226T, acted on (S)-N-(prop-2-yn-1-yl)-2, 3-dihydro-1H-inden-1-amine [(S)-8]. Since (R)-8 is an irreversible inhibitor of monoamine oxidase and (S)-13 is an intermediate of dextromethorphan, a cough suppressant drug, deracemizations of 8 and 13 were carried out with crude enzyme extracts of the respective mutants. This resulted in 51% and 78% isolated yields of (R)-8 and (S)-13, respectively, each with high enantiomeric excess (93% and 99% ee). The results demonstrated the application potential of the evolved CHAO mutants in drug synthesis requiring chiral secondary amines.

New recombinant cyclohexylamine oxidase variants for deracemization of secondary amines by orthogonally assaying designed mutants with structurally diverse substrates

PubMed Central

Li, Guangyue; Yao, Peiyuan; Cong, Peiqian; Ren, Jie; Wang, Lei; Feng, Jinhui; Lau, Peter C.K.; Wu, Qiaqing; Zhu, Dunming

2016-01-01

To further expand the substrate range of the cyclohexylamine oxidase (CHAO) from Brevibacterium oxydans, a library of diverse mutants was created and assayed toward a group of structurally diverse substrates. Among them, mutants T198A and M226A exhibited enhanced activity relative to wt CHAO for most (S)-enantiomers of primary amines and some secondary amines. While mutants T198I, L199I, L199F, M226I and M226T were more active than wt CHAO toward the primary amines, mutants T198F, L199T, Y321A, Y321T, Y321I and Y321F enhanced the enzyme activity toward the secondary amines. In particular, mutant Y321I displayed an enhanced catalytic efficiency toward 1-(4-methoxybenzyl)-1, 2, 3, 4, 5, 6, 7, 8-octahydroisoquinoline (13). Whereas a double mutant, Y321I/M226T, acted on (S)-N-(prop-2-yn-1-yl)-2, 3-dihydro-1H-inden-1-amine [(S)-8]. Since (R)-8 is an irreversible inhibitor of monoamine oxidase and (S)-13 is an intermediate of dextromethorphan, a cough suppressant drug, deracemizations of 8 and 13 were carried out with crude enzyme extracts of the respective mutants. This resulted in 51% and 78% isolated yields of (R)-8 and (S)-13, respectively, each with high enantiomeric excess (93% and 99% ee). The results demonstrated the application potential of the evolved CHAO mutants in drug synthesis requiring chiral secondary amines. PMID:27138090

A comparison of cell proliferation in normal and neoplastic intestinal epithelia following either biogenic amine depletion or monoamine oxidase inhibition.

PubMed

Tutton, P J; Barkla, D H

1976-08-11

Epithelial cell proliferation was studied in the jejunum and in the colon of normal rats, in the colon of dimethylhydrazine-treated rats and in dimethylhydrazine-induced adenocarcinoma of the colon using a stathmokinetic technique. Estimates of cell proliferation rates in these four tissues were then repeated in animals which had been depleted of biogenic animes by treatment with reserpine and in animals whose monoamine oxidase was inhibited by treatment with nialamide. In amine-depleted animals cell proliferation essentially ceased in all four tissues examined. Inhibition of monoamine oxidase did not significantly influence cell proliferation in nonmalignant tissues but accelerated cell division in colonic tumours.

Anti-inflammatory effects of LJP 1586 [Z-3-fluoro-2-(4-methoxybenzyl)allylamine hydrochloride], an amine-based inhibitor of semicarbazide-sensitive amine oxidase activity.

PubMed

O'Rourke, Anne M; Wang, Eric Y; Miller, Andrew; Podar, Erika M; Scheyhing, Kelly; Huang, Li; Kessler, Christina; Gao, Hongfeng; Ton-Nu, Huong-Thu; Macdonald, Mary T; Jones, David S; Linnik, Matthew D

2008-02-01

Semicarbazide-sensitive amine oxidase (SSAO, amine oxidase, copper-containing 3, and vascular adhesion protein-1) is a copper-containing enzyme that catalyzes the oxidative deamination of primary amines to an aldehyde, ammonia, and hydrogen peroxide. SSAO is also involved in leukocyte migration to sites of inflammation, and the enzymatic activity of SSAO is essential to this role. Thus, inhibition of SSAO enzyme activity represents a target for the development of small molecule anti-inflammatory compounds. Here, we have characterized the novel SSAO inhibitor, Z-3-fluoro-2-(4-methoxybenzyl)allylamine hydrochloride (LJP 1586), and assessed its anti-inflammatory activity. LJP 1586 is a potent inhibitor of rodent and human SSAO activity, with IC(50) values between 4 and 43 nM. The selectivity of LJP 1586 was confirmed with a broad panel of receptors and enzymes that included the monoamine oxidases A and B. Oral administration of LJP 1586 resulted in complete inhibition of rat lung SSAO, with an ED(50) between 0.1 and 1 mg/kg, and a pharmacodynamic half-life of greater than 24 h. In a mouse model of inflammatory leukocyte trafficking oral dosing with LJP 1586 resulted in significant dose-dependent inhibition of neutrophil accumulation, with an effect comparable to that of anti-leukocyte function-associated antigen-1 antibody. In a rat model of LPS-induced lung inflammation, administration of 10 mg/kg LJP 1586 resulted in a 55% significant reduction in transmigrated cells recovered by bronchoalveolar lavage. The results demonstrate that a selective, orally active small molecule inhibitor of SSAO is an effective anti-inflammatory compound in vivo and provide further support for SSAO as a therapeutic anti-inflammatory target.

Enzymatic oxidation of 2-phenylethylamine to phenylacetic acid and 2-phenylethanol with special reference to the metabolism of its intermediate phenylacetaldehyde.

PubMed

Panoutsopoulos, Georgios I; Kouretas, Demetrios; Gounaris, Elias G; Beedham, Christine

2004-12-01

2-phenylethylamine is an endogenous constituent of the human brain and is implicated in cerebral transmission. This bioactive amine is also present in certain foodstuffs such as chocolate, cheese and wine and may cause undesirable side effects in susceptible individuals. Metabolism of 2-phenylethylamine to phenylacetaldehyde is catalysed by monoamine oxidase B but the oxidation to its acid is usually ascribed to aldehyde dehydrogenase and the contribution of aldehyde oxidase and xanthine oxidase, if any, is ignored. The objective of this study was to elucidate the role of the molybdenum hydroxylases, aldehyde oxidase and xanthine oxidase, in the metabolism of phenylacetaldehyde derived from its parent biogenic amine. Treatments of 2-phenylethylamine with monoamine oxidase were carried out for the production of phenylacetaldehyde, as well as treatments of synthetic or enzymatic-generated phenylacetaldehyde with aldehyde oxidase, xanthine oxidase and aldehyde dehydrogenase. The results indicated that phenylacetaldehyde is metabolised mainly to phenylacetic acid with lower concentrations of 2-phenylethanol by all three oxidising enzymes. Aldehyde dehydrogenase was the predominant enzyme involved in phenylacetaldehyde oxidation and thus it has a major role in 2-phenylethylamine metabolism with aldehyde oxidase playing a less prominent role. Xanthine oxidase does not contribute to the oxidation of phenylacetaldehyde due to low amounts being present in guinea pig. Thus aldehyde dehydrogenase is not the only enzyme oxidising xenobiotic and endobiotic aldehydes and the role of aldehyde oxidase in such reactions should not be ignored.

Biodegradation of Cyclohexylamine by Brevibacterium oxydans IH-35A

PubMed Central

Iwaki, Hiroaki; Shimizu, Masatake; Tokuyama, Tai; Hasegawa, Yoshie

1999-01-01

A bacterial strain capable of growing on cyclohexylamine (CHAM) was isolated by using enrichment and isolation techniques. The strain isolated, strain IH-35A, was classified as a member of the genus Brevibacterium. The results of growth and enzyme studies are consistent with degradation of CHAM via cyclohexanone (CHnone), 6-hexanolactone, 6-hydroxyhexanoate, and adipate. Cell extracts obtained from this strain grown on CHAM contained CHAM oxidase, and the model for CHAM oxidation by this enzyme was similar to the model for deamino oxidation of amine by amine oxidase. PMID:10224025

Sites and Regulation of Polyamine Catabolism in the Tobacco Plant. Correlations with Cell Division/Expansion, Cell Cycle Progression, and Vascular Development1

PubMed Central

Paschalidis, Konstantinos A.; Roubelakis-Angelakis, Kalliopi A.

2005-01-01

We previously gave a picture of the homeostatic characteristics of polyamine (PA) biosynthesis and conjugation in tobacco (Nicotiana tabacum) plant organs during development. In this work, we present the sites and regulation of PA catabolism related to cell division/expansion, cell cycle progression, and vascular development in the tobacco plant. Diamine oxidase (DAO), PA oxidase (PAO), peroxidases (POXs), and putrescine N-methyltransferase expressions follow temporally and spatially discrete patterns in shoot apical cells, leaves (apical, peripheral, and central regions), acropetal and basipetal petiole regions, internodes, and young and old roots in developing plants. DAO and PAO produce hydrogen peroxide, a plant signal molecule and substrate for POXs. Gene expression and immunohistochemistry analyses reveal that amine oxidases in developing tobacco tissues precede and overlap with nascent nuclear DNA and also with POXs and lignification. In mature and old tissues, flow cytometry indicates that amine oxidase and POX activities, as well as pao gene and PAO protein levels, coincide with G2 nuclear phase and endoreduplication. In young versus the older roots, amine oxidases and POX expression decrease with parallel inhibition of G2 advance and endoreduplication, whereas putrescine N-methyltransferase dramatically increases. In both hypergeous and hypogeous tissues, DAO and PAO expression occurs in cells destined to undergo lignification, suggesting a different in situ localization. DNA synthesis early in development and the advance in cell cycle/endocycle are temporally and spatially related to PA catabolism and vascular development. PMID:16040649

An Inner-Sphere Mechanism for Molecular Oxygen Reduction Catalyzed by Copper Amine Oxidases

PubMed Central

Mukherjee, Arnab; Smirnov, Valeriy V.; Lanci, Michael P.; Brown, Doreen E.; Shepard, Eric M.; Dooley, David M.; Roth, Justine P.

2008-01-01

Copper and topaquinone (TPQ) containing amine oxidases utilize O2 for the metabolism of biogenic amines while concomitantly generating H2O2 for use by the cell. The mechanism of O2 reduction has been the subject of long-standing debate due to the obscuring influence of a proton-coupled electron transfer between the tyrosine-derived TPQ and copper, a rapidly established equilibrium precluding assignment of the enzyme in its reactive form. Here we show that substrate-reduced pea seedling amine oxidase (PSAO) exists predominantly in the CuI, TPQ semiquinone state. A new mechanistic proposal for O2 reduction is advanced on the basis of thermodynamic considerations together with kinetic studies (at varying pH, temperature and viscosity), the identification of steady-state intermediates and the analysis of competitive oxygen kinetic isotope effects: 18O KIEs, [kcat/KM(16,16O2)]/[kcat/KM(16,18O2)]. The 18O KIE = 1.0136 ± 0.0013 at pH 7.2 is independent of temperature from 5 to 47°C and insignificantly changed to 1.0122 ± 0.0020 upon raising the pH to 9, thus indicating the absence of kinetic complexity. Using density functional methods, the effect is found to be precisely in the range expected for reversible O2 binding to CuI to afford a superoxide, [CuII(η1-O2)−I]+, intermediate. Electron transfer from the TPQ semiquinone follows in the first irreversible step to form a peroxide, CuII(η1-O2)−II, intermediate driving the reduction of O2. The similar 18O KIEs reported for copper amine oxidases from other sources raise the possibility that all enzymes react by related inner-sphere mechanisms although additional experiments are needed to test this proposal. PMID:18582059

Stability of spermine oxidase to thermal and chemical denaturation: comparison with bovine serum amine oxidase.

PubMed

Cervelli, Manuela; Leonetti, Alessia; Cervoni, Laura; Ohkubo, Shinji; Xhani, Marla; Stano, Pasquale; Federico, Rodolfo; Polticelli, Fabio; Mariottini, Paolo; Agostinelli, Enzo

2016-10-01

Spermine oxidase (SMOX) is a flavin-containing enzyme that specifically oxidizes spermine to produce spermidine, 3-aminopropanaldehyde and hydrogen peroxide. While no crystal structure is available for any mammalian SMOX, X-ray crystallography showed that the yeast Fms1 polyamine oxidase has a dimeric structure. Based on this scenario, we have investigated the quaternary structure of the SMOX protein by native gel electrophoresis, which revealed a composite gel band pattern, suggesting the formation of protein complexes. All high-order protein complexes are sensitive to reducing conditions, showing that disulfide bonds were responsible for protein complexes formation. The major gel band other than the SMOX monomer is the covalent SMOX homodimer, which was disassembled by increasing the reducing conditions, while being resistant to other denaturing conditions. Homodimeric and monomeric SMOXs are catalytically active, as revealed after gel staining for enzymatic activity. An engineered SMOX mutant deprived of all but two cysteine residues was prepared and characterized experimentally, resulting in a monomeric species. High-sensitivity differential scanning calorimetry of SMOX was compared with that of bovine serum amine oxidase, to analyse their thermal stability. Furthermore, enzymatic activity assays and fluorescence spectroscopy were used to gain insight into the unfolding process.

Comparative study of the conformational lock, dissociative thermal inactivation and stability of euphorbia latex and lentil seedling amine oxidases.

PubMed

Amani, M; Moosavi-Movahedi, A A; Floris, G; Longu, S; Mura, A; Moosavi-Nejad, S Z; Saboury, A A; Ahmad, F

2005-04-01

The thermal stability of copper/quinone containing amine oxidases from Euphorbia characias latex (ELAO) and lentil seedlings (LSAO) was measured in 100 mM potassium phosphate buffer (pH 7.0) following changes in absorbance at 292 nm. ELAO was shown to be about 10 degrees C more stable than LSAO. The dissociative thermal inactivation of ELAO was studied using putrescine as substrate at different temperatures in the range 47-70 degrees C, and a "conformational lock" was developed using the theory pertaining to oligomeric enzyme. Moreover ELAO was shown to be more stable towards denaturants than LSAO, as confirmed by dodecyl trimethylammonium bromide denaturation curves. A comparison of the numbers of contact sites in inter-subunits of ELAO relative to LSAO led us to conclude that the higher stability of ELAO to temperature and towards denaturants was due to the presence of larger number of contact sites in the conformational lock of the enzyme. This study also gives a putative common mechanism for thermal inactivation of amine oxidases and explains the importance of C-terminal conserved amino acids residues in this class of enzymes.

Semicarbazide-sensitive amine oxidase substrates potentiate hydralazine hypotension: possible role of hydrogen peroxide.

PubMed

Vidrio, Horacio; Medina, Martha; González-Romo, Pilar; Lorenzana-Jiménez, Marte; Díaz-Arista, Patricia; Baeza, Alejandro

2003-11-01

The relation between inhibition of semicarbazide-sensitive amine oxidase (SSAO) and vasodilation by hydralazine (HYD) was evaluated in chloralose/urethane-anesthetized rats pretreated with various substrates of the enzyme and subsequently administered a threshold hypotensive dose of the vasodilator. The SSAO substrates benzylamine, phenethylamine, and methylamine potentiate the hypotensive response to HYD. Methylamine, which was studied in greater detail because of its status as a possible endogenous SSAO substrate, does not influence the response to the reference vasodilator pinacidil; it does enhance HYD relaxation in aortic rings obtained from pretreated rats. Experiments designed to identify the product of SSAO activity responsible for potentiation by methylamine suggest involvement of hydrogen peroxide (H2O2), as evidenced by the findings that such potentiation is abolished by additional pretreatment with the H2O2-metabolizing enzyme catalase, and that the plasma concentration of H2O2 is increased by methylamine and decreased by HYD. These results are interpreted as a substantiation of the relation between the known SSAO inhibitory effect of HYD and its vasodilator activity. Pretreatment with the SSAO substrates would increase production of H2O2 in vascular smooth muscle and thus magnify the influence of this vasoconstrictor agent on vascular tone. In these conditions, the decrease in H2O2 production and hence in vascular tone caused by SSAO inhibition by HYD would also be magnified. It is speculated that inhibition of vascular SSAO could represent a novel mechanism of vasodilation.

Effects on amine oxidase of substances which antagonize 5-hydroxytryptamine more than tryptamine on the rat fundus strip

PubMed Central

Barlow, R. B.

1961-01-01

Certain substances, 2-bromolysergic acid diethylamide, dimethyltryptamine (3-(2-dimethylaminoethyl)indole), 2-methyldimethyltryptamine (3-(2-dimethylaminoethyl)-2-methylindole), and 5-benzyloxydimethyltryptamine (5-benzyloxy-3-(2-dimethylaminoethyl)indole), antagonize the effects of 5-hydroxytryptamine on the rat fundus strip more than those of tryptamine. These substances have been tested for their ability to inhibit the oxidation of tryptamine and 5-hydroxytryptamine by suspensions of guinea-pig liver and rat fundus. 2-Bromolysergic acid diethylamide has virtually no inhibitory activity and it is doubtful if the others produce any significant inhibition of amine oxidase in the concentrations which antagonize the effects of 5-hydroxytryptamine more than those of tryptamine. It seems that the differential character of the blocking action of these compounds should be ascribed either to interference with the transport of tryptamine (but not 5-hydroxytryptamine) through the cell wall, coupled with the block of a receptor common to both tryptamine and 5-hydroxytryptamine, or to the existence of separate tryptamine and 5-hydroxytryptamine receptors. The amine oxidases of the guinea-pig liver and rat fundus appear to be a mixture of at least two types of enzyme, one of which has a higher affinity for 5-hydroxytryptamine than the other and is more susceptible to inhibition by 2-methyldimethyltryptamine. PMID:13687054

Analgesic effects of the novel semicarbazide-sensitive amine oxidase inhibitor SZV 1287 in mouse pain models with neuropathic mechanisms: Involvement of transient receptor potential vanilloid 1 and ankyrin 1 receptors.

PubMed

Horváth, Ádám; Tékus, Valéria; Bencze, Noémi; Szentes, Nikolett; Scheich, Bálint; Bölcskei, Kata; Szőke, Éva; Mócsai, Attila; Tóth-Sarudy, Éva; Mátyus, Péter; Pintér, Erika; Helyes, Zsuzsanna

2018-05-01

Semicarbazide-sensitive amine oxidase (SSAO) produces tissue irritants by deamination of primary amines, which activate transient receptor potential ankyrin 1 (TRPA1) and vanilloid 1 (TRPV1) receptors expressed predominantly on nociceptors. Since there are no data about its functions in pain, we studied the effects and mechanisms of action of our novel SSAO inhibitor and dual TRPA1/TRPV1 antagonist multi-target drug SZV 1287 in different pain models. Acute chemonociception was induced by TRPV1 and TRPA1 activation (resiniferatoxin and formalin, respectively), chronic arthritis by K/BxN serum transfer, traumatic mononeuropathy by sciatic nerve ligation. SZV 1287 (20 mg/kg i.p.) was investigated in C57BL/6J wildtype (WT), TRPA1- (TRPA1 -/- ) and TRPV1-deficient (TRPV1 -/- ) mice. Paw mechanonociception was measured by aesthesiometry, thermonociception by hot plate, nocifensive behavior by licking duration, volume by plethysmometry, myeloperoxidase activity by luminescence and plasma extravasation by fluorescence imaging, glia activation in pain-related brain regions by immunohistochemistry. SZV 1287 significantly inhibited both TRPA1 and TRPV1 activation-induced acute chemonociception and hyperalgesia. In K/BxN arthritis, daily SZV 1287 injections significantly decreased hyperalgesia, L4-L6 spinal dorsal horn microgliosis, edema and myeloperoxidase activity. SZV 1287-evoked antihyperalgesic and anti-edema effects were absent in TRPV1 -/- , and remarkably reduced in TRPA1 -/- mice. In contrast, myeloperoxidase-inhibitory effect was absent in TRPA1 -/-, but not in TRPV1 -/- animals. Acute SZV 1287 administration resulted in approximately 50% significant reduction of neuropathic hyperalgesia 7 days after nerve ligation, which was not observed in either TRPA1 -/- or TRPV1 -/- mice. SZV 1287 inhibits chronic inflammatory and neuropathic pain via TRPV1 and TRPA1/TRPV1 activation, respectively, highlighting its drug developmental potential. Copyright © 2018 Elsevier Ltd. All rights reserved.

Environmentally friendly chemoselective oxidation of primary aliphatic amines by using a biomimetic electrocatalytic system.

PubMed

Largeron, Martine; Chiaroni, Angèle; Fleury, Maurice-Bernard

2008-01-01

Environmentally friendly oxidation of primary aliphatic amines to imines has been successfully achieved, under metal-free conditions, by the use of diverse electrogenerated o-azaquinone mediators. High catalytic performance, together with high chemoselectivity, were observed with electron-poor o-azaquinone catalysts generated from 2-aminoresorcinol derivatives. Similar to copper amine oxidase enzymes, these mediators exhibited lower reactivity toward alpha-branched primary amines and no reactivity toward secondary amines. In the case of 3,4-aminophenol derivatives lacking a 2-hydroxy group, the generated o-azaquinone species failed to catalyze the oxidation of the amine to the corresponding imine. Further mechanistic considerations allowed a rationalization of the crucial role of the 2-hydroxy group in converting a catalytically inert species into a highly effective biomimetic catalyst.

Engineering an enantioselective amine oxidase for the synthesis of pharmaceutical building blocks and alkaloid natural products.

PubMed

Ghislieri, Diego; Green, Anthony P; Pontini, Marta; Willies, Simon C; Rowles, Ian; Frank, Annika; Grogan, Gideon; Turner, Nicholas J

2013-07-24

The development of cost-effective and sustainable catalytic methods for the production of enantiomerically pure chiral amines is a key challenge facing the pharmaceutical and fine chemical industries. This challenge is highlighted by the estimate that 40-45% of drug candidates contain a chiral amine, fueling a demand for broadly applicable synthetic methods that deliver target structures in high yield and enantiomeric excess. Herein we describe the development and application of a "toolbox" of monoamine oxidase variants from Aspergillus niger (MAO-N) which display remarkable substrate scope and tolerance for sterically demanding motifs, including a new variant, which exhibits high activity and enantioselectivity toward substrates containing the aminodiphenylmethane (benzhydrylamine) template. By combining rational structure-guided engineering with high-throughput screening, it has been possible to expand the substrate scope of MAO-N to accommodate amine substrates containing bulky aryl substituents. These engineered MAO-N biocatalysts have been applied in deracemization reactions for the efficient asymmetric synthesis of the generic active pharmaceutical ingredients Solifenacin and Levocetirizine as well as the natural products (R)-coniine, (R)-eleagnine, and (R)-leptaflorine. We also report a novel MAO-N mediated asymmetric oxidative Pictet-Spengler approach to the synthesis of (R)-harmicine.

Development of 2-(Substituted Benzylamino)-4-Methyl-1, 3-Thiazole-5-Carboxylic Acid Derivatives as Xanthine Oxidase Inhibitors and Free Radical Scavengers.

PubMed

Ali, Md Rahmat; Kumar, Suresh; Afzal, Obaid; Shalmali, Nishtha; Sharma, Manju; Bawa, Sandhya

2016-04-01

A series of 2-(substituted benzylamino)-4-methylthiazole-5-carboxylic acid was designed and synthesized as structural analogue of febuxostat. A methylene amine spacer was incorporated between the phenyl ring and thiazole ring in contrast to febuxostat in which the phenyl ring was directly linked with the thiazole moiety. The purpose of incorporating methylene amine was to provide a heteroatom which is expected to favour hydrogen bonding within the active site residues of the enzyme xanthine oxidase. The structure of all the compounds was established by the combined use of FT-IR, NMR and MS spectral data. All the compounds were screened in vitro for their ability to inhibit the enzyme xanthine oxidase as per the reported procedure along with DPPH free radical scavenging assay. Compounds 5j, 5k and 5l demonstrated satisfactory potent xanthine oxidase inhibitory activities with IC50 values, 3.6, 8.1 and 9.9 μm, respectively, whereas compounds 5k, 5n and 5p demonstrated moderate antioxidant activities having IC50 15.3, 17.6 and 19.6 μm, respectively, along with xanthine oxidase inhibitory activity. Compound 5k showed moderate xanthine oxidase inhibitory activity as compared with febuxostat along with antioxidant activity. All the compounds were also studied for their binding affinity in active site of enzyme (PDB ID-1N5X). © 2015 John Wiley & Sons A/S.

A Bioinspired Organocatalytic Cascade for the Selective Oxidation of Amines under Air.

PubMed

Largeron, Martine; Fleury, Maurice-Bernard

2017-05-17

A bioinspired organocatalytic cascade reaction for the selective aerobic oxidative cross-coupling of primary amines to imines is described. This approach takes advantages of commercially available pyrogallol monomeric precursor to deliver low loadings of natural purpurogallin in situ, under air. This is further engaged in a catalytic process with the amine substrate affording, under single turnover, the active biomimetic quinonoid organocatalyst and the homocoupled imine intermediate, which is then converted into cross-coupled imine after dynamic transimination. This organocatalytic cascade inspired by both purpurogallin biosynthesis and copper amine oxidases allows the aerobic oxidation of non-activated primary amines that non-enzymatic organocatalysts were not able to accomplish alone. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Biogenic amines in dry fermented sausages: a review.

PubMed

Suzzi, Giovanna; Gardini, Fausto

2003-11-15

Biogenic amines are compounds commonly present in living organisms in which they are responsible for many essential functions. They can be naturally present in many foods such as fruits and vegetables, meat, fish, chocolate and milk, but they can also be produced in high amounts by microorganisms through the activity of amino acid decarboxylases. Excessive consumption of these amines can be of health concern because their not equilibrate assumption in human organism can generate different degrees of diseases determined by their action on nervous, gastric and intestinal systems and blood pressure. High microbial counts, which characterise fermented foods, often unavoidably lead to considerable accumulation of biogenic amines, especially tyramine, 2-phenylethylamine, tryptamine, cadaverine, putrescine and histamine. However, great fluctuations of amine content are reported in the same type of product. These differences depend on many variables: the quali-quantitative composition of microbial microflora, the chemico-physical variables, the hygienic procedure adopted during production, and the availability of precursors. Dry fermented sausages are worldwide diffused fermented meat products that can be a source of biogenic amines. Even in the absence of specific rules and regulations regarding the presence of these compounds in sausages and other fermented products, an increasing attention is given to biogenic amines, especially in relation to the higher number of consumers with enhanced sensitivity to biogenic amines determined by the inhibition of the action of amino oxidases, the enzymes involved in the detoxification of these substances. The aim of this paper is to give an overview on the presence of these compounds in dry fermented sausages and to discuss the most important factors influencing their accumulation. These include process and implicit factors as well as the role of starter and nonstarter microflora growing in the different steps of sausage production. Moreover, the role of microorganisms with amino oxidase activity as starter cultures to control or reduce the accumulation of biogenic amines during ripening and storage of sausages is discussed.

Bioinspired aerobic oxidation of secondary amines and nitrogen heterocycles with a bifunctional quinone catalyst.

PubMed

Wendlandt, Alison E; Stahl, Shannon S

2014-01-08

Copper amine oxidases are a family of enzymes with quinone cofactors that oxidize primary amines to aldehydes. The native mechanism proceeds via an iminoquinone intermediate that promotes high selectivity for reactions with primary amines, thereby constraining the scope of potential biomimetic synthetic applications. Here we report a novel bioinspired quinone catalyst system consisting of 1,10-phenanthroline-5,6-dione/ZnI2 that bypasses these constraints via an abiological pathway involving a hemiaminal intermediate. Efficient aerobic dehydrogenation of non-native secondary amine substrates, including pharmaceutically relevant nitrogen heterocycles, is demonstrated. The ZnI2 cocatalyst activates the quinone toward amine oxidation and provides a source of iodide, which plays an important redox-mediator role to promote aerobic catalytic turnover. These findings provide a valuable foundation for broader development of aerobic oxidation reactions employing quinone-based catalysts.

The amine oxidase inhibitor phenelzine limits lipogenesis in adipocytes without inhibiting insulin action on glucose uptake.

PubMed

Carpéné, Christian; Grès, Sandra; Rascalou, Simon

2013-06-01

The antidepressant phenelzine is a monoamine oxidase inhibitor known to inhibit various other enzymes, among them semicarbazide-sensitive amine oxidase (currently named primary amine oxidase: SSAO/PrAO), absent from neurones but abundant in adipocytes. It has been reported that phenelzine inhibits adipocyte differentiation of cultured preadipocytes. To further explore the involved mechanisms, our aim was to study in vitro the acute effects of phenelzine on de novo lipogenesis in mature fat cells. Therefore, glucose uptake and incorporation into lipid were measured in mouse adipocytes in response to phenelzine, other hydrazine-based SSAO/PrAO-inhibitors, and reference agents. None of the inhibitors was able to impair the sevenfold activation of 2-deoxyglucose uptake induced by insulin. Phenelzine did not hamper the effect of lower doses of insulin. However, insulin-stimulated glucose incorporation into lipids was dose-dependently inhibited by phenelzine and pentamidine, but not by semicarbazide or BTT2052. In contrast, all these SSAO/PrAO inhibitors abolished the transport and lipogenesis stimulation induced by benzylamine. These data indicate that phenelzine does not inhibit glucose transport, the first step of lipogenesis, but inhibits at 100 μM the intracellular triacylglycerol assembly, consistently with its long-term anti-adipogenic effect and such rapid action was not found with all the hydrazine derivatives tested. Therefore, the alterations of body weight control consecutive to the use of this antidepressant drug might be not only related to central effects on food intake/energy expenditure, but could also depend on its direct action in adipocytes. Nonetheless, phenelzine antilipogenic action is not merely dependent on SSAO/PrAO inhibition.

Plasma treatment of polymers for improved adhesion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kelber, J.A.

1988-01-01

A variety of plasma treatments of polymer surfaces for improved adhesion are reviewed: noble and reactive gas treatment of fluoropolymers; noble and reactive treatment of polyolefins, and plasma-induced amination of polymer fibers. The plasma induced surface chemical and morphological changes are discussed, as are the mechanisms of adhesion to polymeric adhesives, particularly epoxy. Noble gas plasma etching of flouropolymers produces a partially defluorinated, textured surface. The mechanical interlocking of this textured surface is the primary cause of improved adhesion to epoxy. Reactive gas plasmas also induce defluorination, but oxygen containing gases cause continual ablation of the fluoropolymer surface. Noble andmore » reactive gas (exept for hydrogen) etching of polyolefins results in surface oxidation and improved adhesion via hydrogen bonding of these oxygen containing groups across the interface. The introduction of amine groups to a polymer surface by amonia or amine plasma treatment generally results in improved adhesion to epoxy. However, amine-epoxy ring interactions can be severely effected by steric factors due to chemical groups surrounding the amine. 41 refs.« less

The role of protein crystallography in defining the mechanisms of biogenesis and catalysis in copper amine oxidase.

PubMed

Klema, Valerie J; Wilmot, Carrie M

2012-01-01

Copper amine oxidases (CAOs) are a ubiquitous group of enzymes that catalyze the conversion of primary amines to aldehydes coupled to the reduction of O(2) to H(2)O(2). These enzymes utilize a wide range of substrates from methylamine to polypeptides. Changes in CAO activity are correlated with a variety of human diseases, including diabetes mellitus, Alzheimer's disease, and inflammatory disorders. CAOs contain a cofactor, 2,4,5-trihydroxyphenylalanine quinone (TPQ), that is required for catalytic activity and synthesized through the post-translational modification of a tyrosine residue within the CAO polypeptide. TPQ generation is a self-processing event only requiring the addition of oxygen and Cu(II) to the apoCAO. Thus, the CAO active site supports two very different reactions: TPQ synthesis, and the two electron oxidation of primary amines. Crystal structures are available from bacterial through to human sources, and have given insight into substrate preference, stereospecificity, and structural changes during biogenesis and catalysis. In particular both these processes have been studied in crystallo through the addition of native substrates. These latter studies enable intermediates during physiological turnover to be directly visualized, and demonstrate the power of this relatively recent development in protein crystallography.

The Role of Protein Crystallography in Defining the Mechanisms of Biogenesis and Catalysis in Copper Amine Oxidase

PubMed Central

Klema, Valerie J.; Wilmot, Carrie M.

2012-01-01

Copper amine oxidases (CAOs) are a ubiquitous group of enzymes that catalyze the conversion of primary amines to aldehydes coupled to the reduction of O2 to H2O2. These enzymes utilize a wide range of substrates from methylamine to polypeptides. Changes in CAO activity are correlated with a variety of human diseases, including diabetes mellitus, Alzheimer’s disease, and inflammatory disorders. CAOs contain a cofactor, 2,4,5-trihydroxyphenylalanine quinone (TPQ), that is required for catalytic activity and synthesized through the post-translational modification of a tyrosine residue within the CAO polypeptide. TPQ generation is a self-processing event only requiring the addition of oxygen and Cu(II) to the apoCAO. Thus, the CAO active site supports two very different reactions: TPQ synthesis, and the two electron oxidation of primary amines. Crystal structures are available from bacterial through to human sources, and have given insight into substrate preference, stereospecificity, and structural changes during biogenesis and catalysis. In particular both these processes have been studied in crystallo through the addition of native substrates. These latter studies enable intermediates during physiological turnover to be directly visualized, and demonstrate the power of this relatively recent development in protein crystallography. PMID:22754303

Cyanide as a copper and quinone-directed inhibitor of amine oxidases from pea seedlings ( Pisum sativum) and Arthrobacter globiformis: evidence for both copper coordination and cyanohydrin derivatization of the quinone cofactor.

PubMed

Shepard, Eric M; Juda, Gregory A; Ling, Ke-Qing; Sayre, Lawrence M; Dooley, David M

2004-04-01

The interactions of cyanide with two copper-containing amine oxidases (CuAOs) from pea seedlings (PSAO) and the soil bacterium Arthrobacter globiformis (AGAO) have been investigated by spectroscopic and kinetic techniques. Previously, we rationalized the effects of azide and cyanide for several CuAOs in terms of copper coordination by these exogenous ligands and their effects on the internal redox equilibrium TPQ(amr)-Cu(II) right harpoon over left harpoon TPQ(sq)-Cu(I). The mechanism of cyanide inhibition was proposed to occur through complexation to Cu(I), thereby directly competing with O(2) for reoxidation of TPQ. Although cyanide readily and reversibly reacts with quinones, no direct spectroscopic evidence for cyanohydrin derivatization of TPQ has been previously documented for CuAOs. This work describes the first direct spectroscopic evidence, using both model and enzyme systems, for cyanohydrin derivatization of TPQ. K(d) values for Cu(II)-CN(-) and Cu(I)-CN(-), as well as the K(i) for cyanide inhibition versus substrate amine, are reported for PSAO and AGAO. In spite of cyanohydrin derivatization of the TPQ cofactor in these enzymes, the uncompetitive inhibition of amine oxidation is determined to arise almost exclusively through CN(-) complexation of Cu(I).

Bioinspired Aerobic Oxidation of Secondary Amines and Nitrogen Heterocycles with a Bifunctional Quinone Catalyst

PubMed Central

Wendlandt, Alison E.; Stahl, Shannon S.

2014-01-01

Copper amine oxidases are a family of enzymes with quinone cofactors that oxidize primary amines to aldehydes. The native mechanism proceeds via an iminoquinone intermediate that promotes high selectivity for reactions with primary amines, thereby constraining the scope of potential biomimetic synthetic applications. Here, we report a novel bioinspired quinone catalyst system, consisting of 1,10-phenanthroline-5,6-dione/ZnI2, that bypasses these constraints via an abiological pathway involving a hemiaminal intermediate. Efficient aerobic dehydrogenation of non-native secondary amine substrates, including pharmaceutically relevant nitrogen heterocycles, is demonstrated. The ZnI2 cocatalyst activates the quinone toward amine oxidation and provides a source of iodide, which plays an important redox-mediator role to promote aerobic catalytic turnover. These findings provide a valuable foundation for broader development of aerobic oxidation reactions employing quinone-based catalysts. PMID:24328193

Gravity Responsive NADH Oxidase of the Plasma Membrane

NASA Technical Reports Server (NTRS)

Morre, D. James (Inventor)

2002-01-01

A method and apparatus for sensing gravity using an NADH oxidase of the plasma membrane which has been found to respond to unit gravity and low centrifugal g forces. The oxidation rate of NADH supplied to the NADH oxidase is measured and translated to represent the relative gravitational force exerted on the protein. The NADH oxidase of the plasma membrane may be obtained from plant or animal sources or may be produced recombinantly.

Putting together a plasma membrane NADH oxidase: a tale of three laboratories.

PubMed

Löw, Hans; Crane, Frederick L; Morré, D James

2012-11-01

The observation that high cellular concentrations of NADH were associated with low adenylate cyclase activity led to a search for the mechanism of the effect. Since cyclase is in the plasma membrane, we considered the membrane might have a site for NADH action, and that NADH might be oxidized at that site. A test for NADH oxidase showed very low activity, which could be increased by adding growth factors. The plasma membrane oxidase was not inhibited by inhibitors of mitochondrial NADH oxidase such as cyanide, rotenone or antimycin. Stimulation of the plasma membrane oxidase by iso-proterenol or triiodothyronine was different from lack of stimulation in endoplasmic reticulum. After 25 years of research, three components of a trans membrane NADH oxidase have been discovered. Flavoprotein NADH coenzyme Q reductases (NADH cytochrome b reductase) on the inside, coenzyme Q in the middle, and a coenzyme Q oxidase on the outside as a terminal oxidase. The external oxidase segment is a copper protein with unique properties in timekeeping, protein disulfide isomerase and endogenous NADH oxidase activity, which affords a mechanism for control of cell growth by the overall NADH oxidase and the remarkable inhibition of oxidase activity and growth of cancer cells by a wide range of anti-tumor drugs. A second trans plasma membrane electron transport system has been found in voltage dependent anion channel (VDAC), which has NADH ferricyanide reductase activity. This activity must be considered in relation to ferricyanide stimulation of growth and increased VDAC antibodies in patients with autism. Copyright © 2012 Elsevier Ltd. All rights reserved.

Dysregulated hepatic expression of glucose transporters in chronic disease: contribution of semicarbazide-sensitive amine oxidase to hepatic glucose uptake.

PubMed

Karim, Sumera; Liaskou, Evaggelia; Fear, Janine; Garg, Abhilok; Reynolds, Gary; Claridge, Lee; Adams, David H; Newsome, Philip N; Lalor, Patricia F

2014-12-15

Insulin resistance is common in patients with chronic liver disease (CLD). Serum levels of soluble vascular adhesion protein-1 (VAP-1) are also increased in these patients. The amine oxidase activity of VAP-1 stimulates glucose uptake via translocation of transporters to the cell membrane in adipocytes and smooth muscle cells. We aimed to document human hepatocellular expression of glucose transporters (GLUTs) and to determine if VAP-1 activity influences receptor expression and hepatic glucose uptake. Quantitative PCR and immunocytochemistry were used to study human liver tissue and cultured cells. We also used tissue slices from humans and VAP-1-deficient mice to assay glucose uptake and measure hepatocellular responses to stimulation. We report upregulation of GLUT1, -3, -5, -6, -7, -8, -9, -10, -11, -12, and -13 in CLD. VAP-1 expression and enzyme activity increased in disease, and provision of substrate to hepatic VAP-1 drives hepatic glucose uptake. This effect was sensitive to inhibition of VAP-1 and could be recapitulated by H2O2. VAP-1 activity also altered expression and subcellular localization of GLUT2, -4, -9, -10, and -13. Therefore, we show, for the first time, alterations in hepatocellular expression of glucose and fructose transporters in CLD and provide evidence that the semicarbazide-sensitive amine oxidase activity of VAP-1 modifies hepatic glucose homeostasis and may contribute to patterns of GLUT expression in chronic disease. Copyright © 2014 the American Physiological Society.

Structures and Mechanism of the Monoamine Oxidase Family

PubMed Central

Gaweska, Helena; Fitzpatrick, Paul F.

2011-01-01

Members of the monoamine oxidase family of flavoproteins catalyze the oxidation of primary and secondary amines, polyamines, amino acids, and methylated lysine side chains in proteins. The enzymes have similar overall structures, with conserved FAD-binding domains and varied substrate-binding sites. Multiple mechanisms have been proposed for the catalytic reactions of these enzymes. The present review compares the structures of different members of the family and the various mechanistic proposals. PMID:22022344

Sensitive amperometric biosensor for the determination of biogenic and synthetic amines using pea seedlings amine oxidase: a novel approach for enzyme immobilisation.

PubMed

Wimmerová, M; Macholán, L

1999-12-01

We prepared a new inorganic sorbent based on modified triazine (2-[4,6-bis (aminoethylamine)-1,3,5-triazine]-Silasorb; BAT-Silasorb) which binds pea seedlings amine oxidase (PSAO) very tightly without loss of its catalytic activity. This unique feature as well as the wide substrate specificity of PSAO was successfully utilised in the construction of an amperometric biosensor based on a carbon paste electrode for the fast and sensitive detection of various amines at a formal potential 0 mV versus Ag/AgCl reference electrode. The reaction layer of the biosensor is created by the direct immobilisation of PSAO at the electrode surface via affinity carrier BAT-Silasorb. Used arrangement facilitates a simple restoration of the inactive biosensor. An amperometric signal results from horseradish peroxidase catalysed reduction of H2O2, a secondary product of the oxidative deamination of amines, catalysed by PSAO. The sensor was used for the basic characterisation of 55 biogenic and synthetic amines, from numerous mono-, di- and polyamines to various hydroxy-, thio-, benzyl- and aromatic derivatives in order to establish its suitability as a postcolumn detector. Its high sensitivity to putrescine 20.0 +/- 0.64 mA l-1 per mol (636.9 +/- 2.03 mA l-1 per mol per cm2), a limit of detection of 10 nmol l-1 (determined with respect to a signal-to-noise ratio 3:1), a linear range of current response to 0.01-100 mumol l-1 concentration of substrate and good reproducibility all indicate that the sensor could be applied to future industrial and clinical analyses.

Nitric oxide production and monoamine oxidase activity in cancer patients during interferon-alpha therapy.

PubMed

Fekkes, Durk; Van Gool, Arthur R; Bannink, Marjolein; Sleijfer, Stefan; Kruit, Wim H J; van der Holt, Bronno; Eggermont, Alexander M M; Hengeveld, Michiel W; Stoter, Gerrit

2009-10-01

Both increased and decreased nitric oxide (NO) synthesis have been reported in patients treated with interferon-alpha (IFN-alpha). Animal studies showed that IFN-alpha administration results in increased levels of biogenic amines, subsequent activation of monoamine oxidases (MAOs), and finally in a change in NO production due to the H(2)O(2) generated by MAOs. We examined the potential relationship between NO production in plasma and MAO-B activity in platelets of 43 cancer patients during 8 weeks of treatment with IFN-alpha. NO synthesis was quantitated by measuring both the ratio of citrulline and arginine (CIT/ARG-ratio) and total nitrite/nitrate (NOx) levels. Compared to baseline, MAO activity and NOx increased, while the CIT/ARG-ratio decreased. No associations were found between NOx, MAO and CIT/ARG-ratio. Only few associations were observed between changes in the biochemical parameters and changes in psychopathology induced by IFN-alpha, of which the association between changes in CIT and lassitude was the most consistent. The results suggest that peripheral NO production and MAO activity are unrelated to each other, and that peripheral changes in these biochemical parameters induced by IFN-alpha are unlikely to contribute to definite psychiatric disturbance.

Norepinephrine metabolism in neuron: dissociation between 3,4-dihydroxyphenylglycol and 3,4-dihydroxymandelic acid pathways.

PubMed

Dong, Wen-Xin; Ni, Xiang-Lian

2002-01-01

To investigate the pre-synaptic metabolism of norepinephrine (NE), judged by variations in plasma concentration of 3,4-dihydroxyphenylglycol (DHPG) and 3,4-dihydroxymandelic acid (DOMA). Pithed and electrically stimulated (2.5 Hz) rats were given intravenous infusion of exogenous NE (6 nmol . kg-1 . min-1). Plasma NE, DHPG, DOMA, and the activities of mono- amine oxidases (MAO) were measured with the radio-enzymatic assay. Exogenous NE induces an about 100-fold increase in plasma NE concentration while blood pressure remained within normal limits. A 12-fold increase in plasma DHPG and 1.2-fold increase for DOMA were observed. When NE transportation across the pre-synaptic membrane was inhibited by desipramine (2 mg/kg, iv), a great reduction in plasma DHPG concentration (about 25 % of control) was observed while DOMA remained unchanged. When MAO-A activity was inhibited to 25 % of control by clorgyline (2 mg/kg, iv) and MAO-B to 30 % by deprenyl, the plasma DHPG and DOMA concentrations were reduced to 15 % and 70 % of controls, and to 26 % and 76 % of controls, respectively. When clorgyline and deprenyl were combined, plasma DHPG was vanished (less than 2 % of control) while plasma DOMA remained in the same range (72 % of control). The metabolizing system of NE in pre-synapse, associating with the pre-synaptic reuptake plus oxidative deamination on the external membrane of mitochondria, is predominant for the reduction to DHPG.

Methylamine-Sensitive Amperometric Biosensor Based on (His)6-Tagged Hansenula polymorpha Methylamine Oxidase Immobilized on the Gold Nanoparticles

PubMed Central

Stasyuk, Nataliya Ye.; Smutok, Oleh V.; Zakalskiy, Andriy E.; Zakalska, Oksana M.; Gonchar, Mykhailo V.

2014-01-01

A novel methylamine-selective amperometric bienzyme biosensor based on recombinant primary amine oxidase isolated from the recombinant yeast strain Saccharomyces cerevisiae and commercial horseradish peroxidase is described. Two amine oxidase preparations were used: free enzyme (AMO) and covalently immobilized on the surface of gold nanoparticles (AMO-nAu). Some bioanalytical parameters (sensitivity, selectivity, and storage stability) of the developed biosensors were investigated. The sensitivity for both sensors is high: 1450 ± 113 and 700 ± 30 A−1 ·M−1 ·m−2 for AMO-nAu biosensor, respectively. The biosensors exhibit the linear range from 15 μM to 150 μM (AMO-nAu) and from 15 μM to 60 μM (AMO). The developed biosensor demonstrated a good selectivity toward methylamine (MA) (signal for dimethylamine and trimethylamine is less than 5% and for ethylamine 15% compared to MA output) and reveals a satisfactory storage stability. The constructed amperometric biosensor was used for MA assay in real samples of fish products in comparison with chemical method. The values obtained with both approaches different methods demonstrated a high correlation. PMID:25136590

Control of Biogenic Amines in Food—Existing and Emerging Approaches

PubMed Central

Naila, Aishath; Flint, Steve; Fletcher, Graham; Bremer, Phil; Meerdink, Gerrit

2010-01-01

Biogenic amines have been reported in a variety of foods, such as fish, meat, cheese, vegetables, and wines. They are described as low molecular weight organic bases with aliphatic, aromatic, and heterocyclic structures. The most common biogenic amines found in foods are histamine, tyramine, cadaverine, 2-phenylethylamine, spermine, spermidine, putrescine, tryptamine, and agmatine. In addition octopamine and dopamine have been found in meat and meat products and fish. The formation of biogenic amines in food by the microbial decarboxylation of amino acids can result in consumers suffering allergic reactions, characterized by difficulty in breathing, itching, rash, vomiting, fever, and hypertension. Traditionally, biogenic amine formation in food has been prevented, primarily by limiting microbial growth through chilling and freezing. However, for many fishing based subsistence populations, such measures are not practical. Therefore, secondary control measures to prevent biogenic amine formation in foods or to reduce their levels once formed need to be considered as alternatives. Such approaches to limit microbial growth may include hydrostatic pressures, irradiation, controlled atmosphere packaging, or the use of food additives. Histamine may potentially be degraded by the use of bacterial amine oxidase or amine-negative bacteria. Only some will be cost-effective and practical for use in subsistence populations. PMID:21535566

Chronic benzylamine administration in the drinking water improves glucose tolerance, reduces body weight gain and circulating cholesterol in high-fat diet-fed mice.

PubMed

Iffiú-Soltész, Zsuzsa; Wanecq, Estelle; Lomba, Almudena; Portillo, Maria P; Pellati, Federica; Szöko, Eva; Bour, Sandy; Woodley, John; Milagro, Fermin I; Alfredo Martinez, J; Valet, Philippe; Carpéné, Christian

2010-04-01

Benzylamine is found in Moringa oleifera, a plant used to treat diabetes in traditional medicine. In mammals, benzylamine is metabolized by semicarbazide-sensitive amine oxidase (SSAO) to benzaldehyde and hydrogen peroxide. This latter product has insulin-mimicking action, and is involved in the effects of benzylamine on human adipocytes: stimulation of glucose transport and inhibition of lipolysis. This study examined whether chronic, oral administration of benzylamine could improve glucose tolerance and the circulating lipid profile without increasing oxidative stress in overweight and pre-diabetic mice. The benzylamine diffusion across the intestine was verified using everted gut sacs. Then, glucose handling and metabolic markers were measured in mice rendered insulin-resistant when fed a high-fat diet (HFD) and receiving or not benzylamine in their drinking water (3600micromol/(kgday)) for 17 weeks. HFD-benzylamine mice showed lower body weight gain, fasting blood glucose, total plasma cholesterol and hyperglycaemic response to glucose load when compared to HFD control. In adipocytes, insulin-induced activation of glucose transport and inhibition of lipolysis remained unchanged. In aorta, benzylamine treatment partially restored the nitrite levels that were reduced by HFD. In liver, lipid peroxidation markers were reduced. Resistin and uric acid, surrogate plasma markers of metabolic syndrome, were decreased. In spite of the putative deleterious nature of the hydrogen peroxide generated during amine oxidation, and in agreement with its in vitro insulin-like actions found on adipocytes, the SSAO-substrate benzylamine could be considered as a potential oral agent to treat metabolic syndrome. Copyright 2010 Elsevier Ltd. All rights reserved.

Real-Time In Vivo Monitoring of Reactive Oxygen Species in Guard Cells.

PubMed

Park, Ky Young; Roubelakis-Angelakis, Kalliopi A

2018-01-01

The intra-/intercellular homeostasis of reactive oxygen species (ROS), and especially of superoxides (O 2 .- ) and hydrogen peroxide (O 2 .- ) participate in signalling cascades which dictate developmental processes and reactions to biotic/abiotic stresses. Polyamine oxidases terminally oxidize/back convert polyamines generating H 2 O 2 . Recently, an NADPH-oxidase/Polyamine oxidase feedback loop was identified to control oxidative burst under salinity. Thus, the real-time localization/monitoring of ROS in specific cells, such as the guard cells, can be of great interest. Here we present a detailed description of the real-time in vivo monitoring of ROS in the guard cells using H 2 O 2 - and O 2 .- specific fluorescing probes, which can be used for studying ROS accumulation generated from any source, including the amine oxidases-dependent pathway, during development and stress.

Plasma diamine oxidase levels in pregnancy complicated by threatened abortion.

PubMed

Legge, M; Duff, G B

1981-02-01

Plasma diamine oxidase levels were assayed in 66 patients who presented with pregnancy complicated by threatened abortion. Levels within the normal range were associated with continuing pregnancies, whereas levels below the normal range were associated with subsequent abortion. Among those patients in whom gestation was greater than eight weeks, 66.6% of diamine oxidase levels correctly predicted the pregnancy outcome. Assay of the diamine oxidase levels at eight weeks of gestation or less gave little useful information.

The role of monamine oxidase inhibition in the acute toxicity of chlordimeform.

DOT National Transportation Integrated Search

1977-08-01

This paper presents data from experiments on male rats performed to determine whether drugs which interfere with central amine mechanisms would decrease the lethality of the acaricide chlordimeform (and thus be of potential value as antidotes for acc...

Histamine 50-skin-prick test: a tool to diagnose histamine intolerance.

PubMed

Kofler, Lukas; Ulmer, Hanno; Kofler, Heinz

2011-01-01

Background. Histamine intolerance results from an imbalance between histamine intake and degradation. In healthy persons, dietary histamine can be sufficiently metabolized by amine oxidases, whereas persons with low amine oxidase activity are at risk of histamine toxicity. Diamine oxidase (DAO) is the key enzyme in degradation. Histamine elicits a wide range of effects. Histamine intolerance displays symptoms, such as rhinitis, headache, gastrointestinal symptoms, palpitations, urticaria and pruritus. Objective. Diagnosis of histamine intolerance until now is based on case history; neither a validated questionnaire nor a routine test is available. It was the aim of this trial to evaluate the usefullness of a prick-test for the diagnosis of histamine intolerance. Methods. Prick-testing with 1% histamine solution and wheal size-measurement to assess the relation between the wheal in prick-test, read after 20 to 50 minutes, as sign of slowed histamine degradation as well as history and symptoms of histamine intolerance. Results. Besides a pretest with 17 patients with HIT we investigated 156 persons (81 with HIT, 75 controls): 64 out of 81 with histamine intolerance(HIT), but only 14 out of 75 persons from the control-group presented with a histamine wheal ≥3 mm after 50 minutes (P < .0001). Conclusion and Clinical Relevance. Histamine-50 skin-prickt-test offers a simple tool with relevance.

Histamine 50-Skin-Prick Test: A Tool to Diagnose Histamine Intolerance

PubMed Central

Kofler, Lukas; Ulmer, Hanno; Kofler, Heinz

2011-01-01

Background. Histamine intolerance results from an imbalance between histamine intake and degradation. In healthy persons, dietary histamine can be sufficiently metabolized by amine oxidases, whereas persons with low amine oxidase activity are at risk of histamine toxicity. Diamine oxidase (DAO) is the key enzyme in degradation. Histamine elicits a wide range of effects. Histamine intolerance displays symptoms, such as rhinitis, headache, gastrointestinal symptoms, palpitations, urticaria and pruritus. Objective. Diagnosis of histamine intolerance until now is based on case history; neither a validated questionnaire nor a routine test is available. It was the aim of this trial to evaluate the usefullness of a prick-test for the diagnosis of histamine intolerance. Methods. Prick-testing with 1% histamine solution and wheal size-measurement to assess the relation between the wheal in prick-test, read after 20 to 50 minutes, as sign of slowed histamine degradation as well as history and symptoms of histamine intolerance. Results. Besides a pretest with 17 patients with HIT we investigated 156 persons (81 with HIT, 75 controls): 64 out of 81 with histamine intolerance(HIT), but only 14 out of 75 persons from the control-group presented with a histamine wheal ≥3 mm after 50 minutes (P < .0001). Conclusion and Clinical Relevance. Histamine-50 skin-prickt-test offers a simple tool with relevance. PMID:23724226

Renalase prevents AKI independent of amine oxidase activity.

PubMed

Wang, Ling; Velazquez, Heino; Moeckel, Gilbert; Chang, John; Ham, Ahrom; Lee, H Thomas; Safirstein, Robert; Desir, Gary V

2014-06-01

AKI is characterized by increased catecholamine levels and hypertension. Renalase, a secretory flavoprotein that oxidizes catecholamines, attenuates ischemic injury and the associated increase in catecholamine levels in mice. However, whether the amine oxidase activity of renalase is involved in preventing ischemic injury is debated. In this study, recombinant renalase protected human proximal tubular (HK-2) cells against cisplatin- and hydrogen peroxide-induced necrosis. Similarly, genetic depletion of renalase in mice (renalase knockout) exacerbated kidney injury in animals subjected to cisplatin-induced AKI. Interestingly, compared with the intact renalase protein, a 20-amino acid peptide (RP-220), which is conserved in all known renalase isoforms, but lacks detectable oxidase activity, was equally effective at protecting HK-2 cells against toxic injury and preventing ischemic injury in wild-type mice. Furthermore, in vitro treatment with RP-220 or recombinant renalase rapidly activated Akt, extracellular signal-regulated kinase, and p38 mitogen-activated protein kinases and downregulated c-Jun N-terminal kinase. In summary, renalase promotes cell survival and protects against renal injury in mice through the activation of intracellular signaling cascades, independent of its ability to metabolize catecholamines, and we have identified the region of renalase required for these effects. Renalase and related peptides show potential as therapeutic agents for the prevention and treatment of AKI. Copyright © 2014 by the American Society of Nephrology.

Plasma diamine oxidase levels in pregnancy complicated by threatened abortion.

PubMed Central

Legge, M; Duff, G B

1981-01-01

Plasma diamine oxidase levels were assayed in 66 patients who presented with pregnancy complicated by threatened abortion. Levels within the normal range were associated with continuing pregnancies, whereas levels below the normal range were associated with subsequent abortion. Among those patients in whom gestation was greater than eight weeks, 66.6% of diamine oxidase levels correctly predicted the pregnancy outcome. Assay of the diamine oxidase levels at eight weeks of gestation or less gave little useful information. PMID:6785320

Metabolism of 2-phenylethylamine to phenylacetic acid, via the intermediate phenylacetaldehyde, by freshly prepared and cryopreserved guinea pig liver slices.

PubMed

Panoutsopoulos, Georgios I

2004-01-01

2-Phenylethylamine is an endogenous amine, which acts as a neuromodulator of dopaminergic responses. Exogenous 2-phenylethylamine is found in certain foodstuffs and may cause toxic side-effects in susceptible individuals. The present investigation examined the metabolism of 2-phenylethylamine to phenylacetic acid, via phenylacetaldehyde, in freshly prepared and cryopreserved liver slices. Additionally, it compared the relative contribution of aldehyde oxidase, xanthine oxidase and aldehyde dehydrogenase by using specific inhibitors for each oxidizing enzyme. In freshly prepared and cryopreserved liver slices, phenylacetic acid was the main metabolite of 2-phenylethalamine. In freshly prepared liver slices, phenylacetic acid was completely inhibited by disulfiram (inhibitor of aldehyde dehydrogenase), whereas isovanillin (inhibitor of aldehyde oxidase) inhibited acid formation to a lesser extent and allopurinol (inhibitor of xanthine oxidase) had no effect. In cryopreserved liver slices, isovanillin inhibited phenylacetic acid by 85%, whereas disulfiram inhibited acid formation to a lesser extent and allopurinol had no effect. In liver slices, 2-phenylethylamine is rapidly oxidized to phenylacetic acid, via phenylacetaldehyde, by aldehyde dehydrogenase and aldehyde oxidase with no contribution from xanthine oxidase.

Quantum mechanical hydrogen tunneling in bacterial copper amine oxidase reaction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Murakawa, Takeshi; Okajima, Toshihide; Kuroda, Shun'ichi

A key step decisively affecting the catalytic efficiency of copper amine oxidase is stereospecific abstraction of substrate {alpha}-proton by a conserved Asp residue. We analyzed this step by pre-steady-state kinetics using a bacterial enzyme and stereospecifically deuterium-labeled substrates, 2-phenylethylamine and tyramine. A small and temperature-dependent kinetic isotope effect (KIE) was observed with 2-phenylethylamine, whereas a large and temperature-independent KIE was observed with tyramine in the {alpha}-proton abstraction step, showing that this step is driven by quantum mechanical hydrogen tunneling rather than the classical transition-state mechanism. Furthermore, an Arrhenius-type preexponential factor ratio approaching a transition-state value was obtained in the reactionmore » of a mutant enzyme lacking the critical Asp. These results provide strong evidence for enzyme-enhanced hydrogen tunneling. X-ray crystallographic structures of the reaction intermediates revealed a small difference in the binding mode of distal parts of substrates, which would modulate hydrogen tunneling proceeding through either active or passive dynamics.« less

Design and synthesis of a tetradentate '3-amine-1-carboxylate' ligand to mimic the metal binding environment at the non-heme iron(II) oxidase active site.

PubMed

Dungan, Victoria J; Ortin, Yannick; Mueller-Bunz, Helge; Rutledge, Peter J

2010-04-07

Non-heme iron(II) oxidases (NHIOs) catalyse a diverse array of oxidative chemistry in Nature. As part of ongoing efforts to realize biomimetic, iron-mediated C-H activation, we report the synthesis of a new 'three-amine-one-carboxylate' ligand designed to complex with iron(II) and mimic the NHIO active site. The tetradentate ligand has been prepared as a single enantiomer in nine synthetic steps from N-Cbz-L-alanine, pyridine-2,6-dimethanol and diphenylamine, using Seebach oxazolidinone chemistry to control the stereochemistry. X-Ray crystal structures are reported for two important intermediates, along with variable temperature NMR experiments to probe the hindered interconversion of conformational isomers of several key intermediates, 2,6-disubstituted pyridine derivatives. The target ligand and an N-Cbz-protected precursor were each then complexed with iron(II) and tested for their ability to promote alkene dihydroxylation, using hydrogen peroxide as the oxidant.

Biotransformation and bioactivation reactions of alicyclic amines in drug molecules.

PubMed

Bolleddula, Jayaprakasam; DeMent, Kevin; Driscoll, James P; Worboys, Philip; Brassil, Patrick J; Bourdet, David L

2014-08-01

Aliphatic nitrogen heterocycles such as piperazine, piperidine, pyrrolidine, morpholine, aziridine, azetidine, and azepane are well known building blocks in drug design and important core structures in approved drug therapies. These core units have been targets for metabolic attack by P450s and other drug metabolizing enzymes such as aldehyde oxidase and monoamine oxidase (MAOs). The electron rich nitrogen and/or α-carbons are often major sites of metabolism of alicyclic amines. The most common biotransformations include N-oxidation, N-conjugation, oxidative N-dealkylation, ring oxidation, and ring opening. In some instances, the metabolic pathways generate electrophilic reactive intermediates and cause bioactivation. However, potential bioactivation related adverse events can be attenuated by structural modifications. Hence it is important to understand the biotransformation pathways to design stable drug candidates that are devoid of metabolic liabilities early in the discovery stage. The current review provides a comprehensive summary of biotransformation and bioactivation pathways of aliphatic nitrogen containing heterocycles and strategies to mitigate metabolic liabilities.

Quantum chemical modeling of the inhibition mechanism of monoamine oxidase by oxazolidinone and analogous heterocyclic compounds.

PubMed

Erdem, Safiye Sağ; Özpınar, Gül Altınbaş; Boz, Ümüt

2014-02-01

Monoamine oxidase (MAO, EC 1.4.3.4) is responsible from the oxidation of a variety of amine neurotransmitters. MAO inhibitors are used for the treatment of depression or Parkinson's disease. They also inhibit the catabolism of dietary amines. According to one hypothesis, inactivation results from the formation of a covalent adduct to a cysteine residue in the enzyme. If the adduct is stable enough, the enzyme is inhibited for a long time. After a while, enzyme can turn to its active form as a result of adduct breakdown by β-elimination. In this study, the proposed inactivation mechanism was modeled and tested by quantum chemical calculations. Eight heterocyclic methylthioamine derivatives were selected to represent the proposed covalent adducts. Activation energies related to their β-elimination reactions were calculated using ab initio and density functional theory methods. Calculated activation energies were in good agreement with the relative stabilities of the hypothetical adducts predicted in the literature by enzyme inactivation measurements.

Kinetics of intraventricularly injected trace amines and their deuterated isotopomers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Durden, D.A.; Nguyen, T.V.; Boulton, A.A.

Intraventricular injection into the rat brain of four trace amines and a catecholamine resulted in rapid exponential loss of the amines in the first 30 minutes after injection. The half-lives were: phenylethylamine 3.8 min, para-tyramine 5.1 min, meta-tyramine 7.4 min and dopamine 8.0 min. Tryptamine showed a biphasic loss with half-lives of 4.7 min (over the 5 to 10 min period) and 14.1 min (10 to 30 min). The half-lives were substantially increased by deuterium labeling at the alpha carbon position: phenylethylamine 4.8 min, para-tyramine 8.8 min, meta-tyramine 14.1 min, dopamine 13.0 min and tryptamine 6.0 min (5 to 10more » min period) and 28.7 min (10 to 20 min). The loss of the amines was reduced by monoamine oxidase inhibition by pargyline hydrochloride and the deuterium isotope effect was abolished. It is noteworthy that the half-life of dopamine was similar to those of the trace amines in this time period and that the trace amine half-lives after i.v. injection was longer than those obtained from measurements of increases of concentrations of endogenous amines after MAOI in vivo and that of dopamine shorter than values calculated from turnover measurements.« less

Deletion of MAOA and MAOB in a male patient causes severe developmental delay, intermittent hypotonia and stereotypical hand movements

PubMed Central

Whibley, Annabel; Urquhart, Jill; Dore, Jonathan; Willatt, Lionel; Parkin, Georgina; Gaunt, Lorraine; Black, Graeme; Donnai, Dian; Raymond, F Lucy

2010-01-01

Monoamine oxidases (MAO-A and MAO-B) have a key role in the degradation of amine neurotransmitters, such as dopamine, norepinephrine and serotonin. We identified an inherited 240 kb deletion on Xp11.3–p11.4, which encompasses both monoamine oxidase genes but, unlike other published reports, does not affect the adjacent Norrie disease gene (NDP). The brothers who inherited the deletion, and thus have no monoamine oxidase function, presented with severe developmental delay, intermittent hypotonia and stereotypical hand movements. The clinical features accord with published reports of larger microdeletions and selective MAO-A and MAO-B deficiencies in humans and mouse models and suggest considerable functional compensation between MAO-A and MAO-B under normal conditions. PMID:20485326

Path Integral Simulation of the H/D Kinetic Isotope Effect in Monoamine Oxidase B Catalyzed Decomposition of Dopamine.

PubMed

Mavri, Janez; Matute, Ricardo A; Chu, Zhen T; Vianello, Robert

2016-04-14

Brain monoamines regulate many centrally mediated body functions, and can cause adverse symptoms when they are out of balance. A starting point to address challenges raised by the increasing burden of brain diseases is to understand, at atomistic level, the catalytic mechanism of an essential amine metabolic enzyme-monoamine oxidase B (MAO B). Recently, we demonstrated that the rate-limiting step of MAO B catalyzed conversion of amines into imines represents the hydride anion transfer from the substrate α-CH2 group to the N5 atom of the flavin cofactor moiety. In this article we simulated for MAO B catalyzed dopamine decomposition the effects of nuclear tunneling by the calculation of the H/D kinetic isotope effect. We applied path integral quantization of the nuclear motion for the methylene group and the N5 atom of the flavin moiety in conjunction with the QM/MM treatment on the empirical valence bond (EVB) level for the rest of the enzyme. The calculated H/D kinetic isotope effect of 12.8 ± 0.3 is in a reasonable agreement with the available experimental data for closely related biogenic amines, which gives strong support for the proposed hydride mechanism. The results are discussed in the context of tunneling in enzyme centers and advent of deuterated drugs into clinical practice.

Design, synthesis, pharmacological evaluation, QSAR analysis, molecular modeling and ADMET of novel donepezil-indolyl hybrids as multipotent cholinesterase/monoamine oxidase inhibitors for the potential treatment of Alzheimer's disease.

PubMed

Bautista-Aguilera, Oscar M; Esteban, Gerard; Bolea, Irene; Nikolic, Katarina; Agbaba, Danica; Moraleda, Ignacio; Iriepa, Isabel; Samadi, Abdelouahid; Soriano, Elena; Unzeta, Mercedes; Marco-Contelles, José

2014-03-21

The design, synthesis, and pharmacological evaluation of donepezil-indolyl based amines 7-10, amides 12-16, and carboxylic acid derivatives 5 and 11, as multipotent ASS234 analogs, able to inhibit simultaneously cholinesterase (ChE) and monoamine oxidase (MAO) enzymes for the potential treatment of Alzheimer's disease (AD), is reported. Theoretical studies using 3D-Quantitative Structure-Activity Relationship (3D-QSAR) was used to define 3D-pharmacophores for inhibition of MAO A/B, AChE, and BuChE enzymes. We found that, in general, and for the same substituent, amines are more potent ChE inhibitors (see compounds 12, 13 versus 7 and 8) or equipotent (see compounds 14, 15 versus 9 and 10) than the corresponding amides, showing a clear EeAChE inhibition selectivity. For the MAO inhibition, amides were not active, and among the amines, compound 14 was totally MAO A selective, while amines 15 and 16 were quite MAO A selective. Carboxylic acid derivatives 5 and 11 showed a multipotent moderate selective profile as EeACE and MAO A inhibitors. Propargylamine 15 [N-((5-(3-(1-benzylpiperidin-4-yl)propoxy)-1-methyl-1H-indol-2-yl)methyl)prop-2-yn-1-amine] resulted in the most potent hMAO A (IC50 = 5.5 ± 1.4 nM) and moderately potent hMAO B (IC50 = 150 ± 31 nM), EeAChE (IC50 = 190 ± 10 nM), and eqBuChE (IC50 = 830 ± 160 nM) inhibitor. However, the analogous N-allyl and the N-morpholine derivatives 16 and 14 deserve also attention as they show an attractive multipotent profile. To sum up, donepezil-indolyl hybrid 15 is a promising drug for further development for the potential prevention and treatment of AD. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Functionalized boron nitride nanotubes

DOEpatents

Sainsbury, Toby; Ikuno, Takashi; Zettl, Alexander K

2014-04-22

A plasma treatment has been used to modify the surface of BNNTs. In one example, the surface of the BNNT has been modified using ammonia plasma to include amine functional groups. Amine functionalization allows BNNTs to be soluble in chloroform, which had not been possible previously. Further functionalization of amine-functionalized BNNTs with thiol-terminated organic molecules has also been demonstrated. Gold nanoparticles have been self-assembled at the surface of both amine- and thiol-functionalized boron nitride Nanotubes (BNNTs) in solution. This approach constitutes a basis for the preparation of highly functionalized BNNTs and for their utilization as nanoscale templates for assembly and integration with other nanoscale materials.

Inhibition and oxygen activation in copper amine oxidases.

PubMed

Shepard, Eric M; Dooley, David M

2015-05-19

Copper-containing amine oxidases (CuAOs) use both copper and 2,4,5-trihydroxyphenylalanine quinone (TPQ) to catalyze the oxidative deamination of primary amines. The CuAO active site is highly conserved and comprised of TPQ and a mononuclear type II copper center that exhibits five-coordinate, distorted square pyramidal coordination geometry with histidine ligands and equatorially and axially bound water in the oxidized, resting state. The active site is buried within the protein, and CuAOs from various sources display remarkable diversity with respect to the composition of the active site channel and cofactor accessibility. Structural and mechanistic factors that influence substrate preference and inhibitor sensitivity and selectivity have been defined. This Account summarizes the strategies used to design selective CuAO inhibitors based on active site channel characteristics, leading to either enhanced steric fits or the trapping of reactive electrophilic products. These findings provide a framework to support the future development of candidate molecules aimed at minimizing the negative side effects associated with drugs containing amine functionalities. This is vital given the existence of human diamine oxidase and vascular adhesion protein-1, which have distinct amine substrate preferences and are associated with different metabolic processes. Inhibition of these enzymes by antifungal or antiprotozoal agents, as well as classic monoamine oxidase (MAO) inhibitors, may contribute to the adverse side effects associated with drug treatment. These observations provide a rationale for the limited clinical value associated with certain amine-containing pharmaceuticals and emphasize the need for more selective AO inhibitors. This Account also discusses the novel roles of copper and TPQ in the chemistry of O2 activation and substrate oxidation. Reduced CuAOs exist in a redox equilibrium between the Cu(II)-TPQAMQ (aminoquinol) and Cu(I)-TPQSQ (semiquinone). Elucidating the roles of Cu(I), TPQSQ, and TPQAMQ in O2 activation, for example, distinguishing inner-sphere versus outer-sphere electron transfer mechanisms, has been actively investigated since the discovery of TPQSQ in 1991 and has only recently been clarified. Kinetics and spectroscopic studies encompassing metal substitution, stopped-flow and temperature-jump relaxation methods, and oxygen kinetic isotope experiments have provided strong support for an inner-sphere electron transfer step from Cu(I) to O2. Data for two enzymes support a mechanism wherein O2 prebinds to a three-coordinate Cu(I) site, yielding a [Cu(II)(η(1)-O2(-1))](+) intermediate, with H2O2 generated from ensuing rate-determining proton coupled electron transfer from TPQSQ. While kinetics data from the cobalt-substituted yeast enzyme indicated that O2 is reduced through an outer-sphere process involving TPQAMQ, new findings with a bacterial CuAO demonstrate that both the Cu(II) and Co(II) forms of the enzyme operate via parallel mechanisms involving metal-superoxide intermediates. Structural observations of a coordinated TPQSQ-Cu(I) complex in two CuAOs supports previous indications that Cu(II)/(I) ligand substitution chemistry may be mechanistically relevant. Substantial evidence indicates that rapid and reversible inner-sphere reduction of O2 at a three-coordinate Cu(I) site occurs, but the existence of a coordinated semiquinone in some AOs suggests that, in these enzymes, an outer-sphere reaction between O2 and TPQSQ may also be possible, since this is expected to be energetically favorable compared with outer-sphere electron transfer from TPQAMQ to O2.

A metalloenzyme-like catalytic system for the chemoselective oxidative cross-coupling of primary amines to imines under ambient conditions.

PubMed

Largeron, Martine; Fleury, Maurice-Bernard

2015-02-23

The direct oxidative cross-coupling of primary amines is a challenging transformation as homocoupling is usually preferred. We report herein the chemoselective preparation of cross-coupled imines through the synergistic combination of low loadings of Cu(II) metal-catalyst and o-iminoquinone organocatalyst under ambient conditions. This homogeneous cooperative catalytic system has been inspired by the reaction of copper amine oxidases, a family of metalloenzymes with quinone organic cofactors that mediate the selective oxidation of primary amines to aldehydes. After optimization, the desired cross-coupled imines are obtained in high yields with broad substrate scope through a transamination process that leads to the homocoupled imine intermediate, followed by dynamic transimination. The ability to carry out the reactions at room temperature and with ambient air, rather than molecular oxygen as the oxidant, and equimolar amounts of each coupling partner is particularly attractive from an environmentally viewpoint. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

New pyrrole inhibitors of monoamine oxidase: synthesis, biological evaluation, and structural determinants of MAO-A and MAO-B selectivity.

PubMed

La Regina, Giuseppe; Silvestri, Romano; Artico, Marino; Lavecchia, Antonio; Novellino, Ettore; Befani, Olivia; Turini, Paola; Agostinelli, Enzo

2007-03-08

A series of new pyrrole derivatives have been synthesized and evaluated for their monoamine oxidase (MAO) A and B inhibitory activity and selectivity. N-Methyl,N-(benzyl),N-(pyrrol-2-ylmethyl)amine (7) and N-(2-benzyl),N-(1-methylpyrrol-2-ylmethyl)amine (18) were the most selective MAO-B (7, SI = 0.0057) and MAO-A (18, SI = 12500) inhibitors, respectively. Docking and molecular dynamics simulations gave structural insights into the MAO-A and MAO-B selectivity. Compound 18 forms an H-bond with Gln215 through its protonated amino group into the MAO-A binding site. This H-bond is absent in the 7/MAO-A complex. In contrast, compound 7 places its phenyl ring into an aromatic cage of the MAO-B binding pocket, where it forms charge-transfer interactions. The slightly different binding pose of 18 into the MAO-B active site seems to be forced by a bulkier Tyr residue, which replaces a smaller Ile residue present in MAO-A.

Amine Enrichment of Thin-Film Composite Membranes via Low Pressure Plasma Polymerization for Antimicrobial Adhesion.

PubMed

Reis, Rackel; Dumée, Ludovic F; He, Li; She, Fenghua; Orbell, John D; Winther-Jensen, Bjorn; Duke, Mikel C

2015-07-15

Thin-film composite membranes, primarily based on poly(amide) (PA) semipermeable materials, are nowadays the dominant technology used in pressure driven water desalination systems. Despite offering superior water permeation and salt selectivity, their surface properties, such as their charge and roughness, cannot be extensively tuned due to the intrinsic fabrication process of the membranes by interfacial polymerization. The alteration of these properties would lead to a better control of the materials surface zeta potential, which is critical to finely tune selectivity and enhance the membrane materials stability when exposed to complex industrial waste streams. Low pressure plasma was employed to introduce amine functionalities onto the PA surface of commercially available thin-film composite (TFC) membranes. Morphological changes after plasma polymerization were analyzed by SEM and AFM, and average surface roughness decreased by 29%. Amine enrichment provided isoelectric point changes from pH 3.7 to 5.2 for 5 to 15 min of plasma polymerization time. Synchrotron FTIR mappings of the amine-modified surface indicated the addition of a discrete 60 nm film to the PA layer. Furthermore, metal affinity was confirmed by the enhanced binding of silver to the modified surface, supported by an increased antimicrobial functionality with demonstrable elimination of E. coli growth. Essential salt rejection was shown minimally compromised for faster polymerization processes. Plasma polymerization is therefore a viable route to producing functional amine enriched thin-film composite PA membrane surfaces.

Characterization and application of a diamine oxidase from Lathyrus sativus as component of an electrochemical biosensor for the determination of biogenic amines in wine and beer.

PubMed

Di Fusco, Massimo; Federico, Rodolfo; Boffi, Alberto; Macone, Alberto; Favero, Gabriele; Mazzei, Franco

2011-08-01

In this work, we have characterized a diamine oxidase (DAO) from Lathyrus sativus and evaluated its use, for the first time, as biocatalytic component of an electrochemical biosensor for the determination of biogenic amines index in wine and beer samples. Firstly, DAO was electrokinetically characterized free in solution by means of a platinum electrode and then immobilized by using polyazetidine prepolimer on the surface of screen-printed electrodes constituted of two gold working electrodes. The amperometric measurements were carried out by using a flow system at a fixed potential of +600 mV vs the internal silver pseudo reference in phosphate buffer solution (0.1 mol l(-1), pH = 7.4). The analysis of wine and beer samples were performed in flow injection system using the dual channel transducer providing simultaneous detection of sample and blank signal, and the resulting signal (after subtraction of the blank signal) was referred to that of putrescine. The results were compared with those obtained using a modified reference method based on gas chromatography-mass spectrometry analysis on the same samples. The results obtained in the analysis of Italian wines shows the better suitability of DAO-based biosensor in the determination of the biogenic amines (BAs) index expressed as putrescine equivalent in both red and white wines, being less efficient in beer samples where it underestimates by about 50% the BAs content.

The Relationship between Academic Self-Efficacy and Academic-Related Boredom: "MAOA" Gene as a Moderator

ERIC Educational Resources Information Center

Liu, Yangyang; Lu, Zuhong

2017-01-01

The present study sought to examine the relationship between Chinese high school students' academic self-efficacy and their academic-related boredom. Another objective was to explore the moderating effects of mono-amine-oxidase type A ("MAOA") gene polymorphism on this relationship. In a sample of 514 Chinese high school students, we…

Localization of cholesterol in sphingomyelinase-treated fibroblasts.

PubMed Central

Pörn, M I; Slotte, J P

1995-01-01

The distribution of cellular unesterified cholesterol was studied in fibroblasts, which had been depleted of plasma membrane sphingomyelin by exposure to exogenous sphingomyelinase. This treatment has previously been shown to induce an increase in cholesterol esterification, a decrease in the biosynthesis of cholesterol, and a decreased susceptibility of cell cholesterol to oxidation with cholesterol oxidase. When the cellular localization of cholesterol was studied with fluorescent filipin staining, sphingomyelin depletion did not cause any visible changes in the filipin-cholesterol staining pattern, suggesting that the major part of cellular cholesterol was retained in the plasma membrane after sphingomyelinase treatment. After the oxidation of cell-surface cholesterol with cholesterol oxidase, the plasma membrane was no longer stained by filipin, but the plasma membrane cholesterol of sphingomyelin-depleted cells appeared to be resistant to oxidation with cholesterol oxidase when sphingomyelinase was used as an oxidation-promoting agent. However, the use of hypotonic buffer or phosphatidylcholine-specific phospholipase C together with cholesterol oxidase resulted in a complete oxidation of the cell-surface cholesterol in sphingomyelin-depleted cells, as evidenced by the filipin-cholesterol staining pattern. Similar results were obtained when [3H]cholesterol-labelled fibroblasts were used for determination of the susceptibility to cholesterol oxidation. The kinetics of [3H]cholesterol oxidation in sphingomyelin-depleted cells with cholesterol oxidase in hypotonic buffer indicated that approximately 85% of the cellular cholesterol still resided in the plasma membrane after sphingomyelin depletion. These results are contradictory to earlier reports on sphingomyelinase-induced changes in cellular cholesterol distribution and suggest that minor changes in the kinetics of cholesterol transport from the plasma membrane to the endoplasmic reticulum may be responsible for the sphingomyelinase-induced changes in the rates of cholesterol metabolism. Whereas the use of phospholipases to promote the oxidation of cholesterol in some instances might lead to misinterpretations, the use of hypotonic buffer together with cholesterol oxidase proved to be a more reliable method for the determination of cellular cholesterol distribution. Images Figure 1 Figure 2 PMID:7755574

Pre-clinical evaluation of small molecule LOXL2 inhibitors in breast cancer

PubMed Central

Chang, Joan; Lucas, Morghan C.; Leonte, Lidia E.; Garcia-Montolio, Marc; Singh, Lukram Babloo; Findlay, Alison D.; Deodhar, Mandar; Foot, Jonathan S.; Jarolimek, Wolfgang; Timpson, Paul; Erler, Janine T.; Cox, Thomas R.

2017-01-01

Lysyl Oxidase-like 2 (LOXL2), a member of the lysyl oxidase family of amine oxidases is known to be important in normal tissue development and homeostasis, as well as the onset and progression of solid tumors. Here we tested the anti-tumor properties of two generations of novel small molecule LOXL2 inhibitor in the MDA-MB-231 human model of breast cancer. We confirmed a functional role for LOXL2 activity in the progression of primary breast cancer. Inhibition of LOXL2 activity inhibited the growth of primary tumors and reduced primary tumor angiogenesis. Dual inhibition of LOXL2 and LOX showed a greater effect and also led to a lower overall metastatic burden in the lung and liver. Our data provides the first evidence to support a role for LOXL2 specific small molecule inhibitors as a potential therapy in breast cancer. PMID:28199967

Oxygen activation in flavoprotein oxidases: the importance of being positive.

PubMed

Gadda, Giovanni

2012-04-03

The oxidation of flavin hydroquinones by O(2) in solution is slow, with second-order rate constants of ~250 M(-1) s(-1). This is due to the obligatory, single-electron transfer that initiates the reaction being thermodynamically unfavored and poorly catalyzed. Notwithstanding considerations of O(2) accessibility to the reaction site, its desolvation and geometry and other factors that can also contribute to further rate acceleration, flavoprotein oxidases must activate O(2) for reaction with flavin hydroquinones to be able to achieve the 100-1000-fold rate enhancements typically observed. Protein positive charges have been identified in glucose oxidase, monomeric sarcosine oxidase, N-methyltryptophan oxidase and fructosamine oxidase that electrostatically stabilize the transition state for the initial single electron transfer that generates the O(2)(-•)/flavin semiquinone radical pair. In choline oxidase despite the presence of three histidines in the active site, the trimethylammonium group of the reaction product provides such an electrostatic stabilization. A nonpolar site proximal to the flavin C(4a) atom in choline oxidase has also been identified, which contributes to the geometry and desolvation of the O(2) reaction site. The relevance of O(2) activation by product charges to other flavoprotein oxidases, such as for example those catalyzing amine oxidations, is discussed in this review. A nonpolar site close to the flavin C(4a) atom and a positive charge is identified through structural analysis in several flavoprotein oxidases. Mutagenesis has disclosed nonpolar sites in O(2)-reducing enzymes that utilize copper/TPQ or iron. It is predicted that classes of O(2)-reducing enzymes utilizing other cofactors also contain a similar catalytic motif.

N-propargylbenzylamine, a major metabolite of pargyline, is a potent inhibitor of monoamine oxidase type B in rats in vivo: a comparison with deprenyl.

PubMed Central

Karoum, F.

1987-01-01

In an effort to explore the contribution of the metabolites of pargyline towards the in vivo inhibition of monoamine oxidase (MAO), the effects of pargyline and its major metabolites on the production and metabolism of a number of biogenic amines were studied in rats. The administration of pargyline gave rise to three major ethyl acetate extractable metabolites: benzylamine, N-methylbenzylamine and N-propargylbenzylamine (NPB). Only NPB demonstrated in vivo monoamine oxidase inhibitory properties at an acute dose of 30 mg kg-1. The acute effects of pargyline, NPB, and deprenyl on urine and brain concentrations of a number of biogenic amines (phenylethylamine (PEA), m- and p-tyramine, noradrenaline (NA), dopamine, and 5-hydroxytryptamine (5-HT) and their metabolites were evaluated. Increased urine and brain concentrations of PEA were considered to represent in vivo inhibition of type B MAO while decreased concentrations of NA and 5-HT metabolites were regarded as indicators of an in vivo inhibition of MAO type A. NPB, like deprenyl and pargyline, significantly increased urine and brain PEA while only pargyline reduced 5-HT metabolism, suggesting that the metabolism of pargyline to NPB may contribute towards the MAO type B inhibitory effects of pargyline in vivo. Since the therapeutic benefits of MAO inhibitors in clinical practice usually require some period of chronic treatment, the chronic effects of repeated 14 daily doses of the above MAO inhibitors on central and peripheral biogenic amines were evaluated at the following times: during treatment, one day and five days after termination of treatment. The biochemical changes observed during the course of chronic NPB, pargyline and deprenyl treatments generally follow the expected in vitro characteristics of these drugs, but the detailed changes observed suggest clear differences. For example, the in vivo effect of pargyline on urine 5-hydroxyindoleacetic acid excretion was considerably weaker than its effect on the excretion of NA and dopamine metabolites. These changes are opposite to the in vitro effects of pargyline on 5-HT, dopamine and NA oxidative deamination. Inhibitions of the metabolism of all the amines studied were clearly observed during chronic MAOI treatments, but these effects were less evident five days after the end of treatment, suggesting an almost normal metabolism of biogenic amines. It is concluded that while MAO inhibitors may be the primary compound responsible for MAO inhibition, the effects of their metabolites in some cases may also play equally important roles in the regulation of monoamines both in the periphery and the brain.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:3103805

Monoamine oxidase: Radiotracer chemistry and human studies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fowler, Joanna S.; Logan, Jean; Shumay, Elena

Monoamine oxidase (MAO) oxidizes amines from both endogenous and exogenous sources thereby regulating the concentration of neurotransmitter amines such as serot onin, norepinephrine and dopamine as well as many xenobiotics. MAO inhibitor drugs are used in the treatment of Parkinson’s disease and in depression stimulating the development of radiotracer tools to probe the role of MAO in normal human biology and in disease. Over the past 30 since the first radiotracers were developed and the first PET images of MAO in humans were carried out, PET studies of brain MAO in healthy volunteers and in patients have identified different variablesmore » which have contributed to different MAO levels in brain and in peripheral organs. MAO radiotracers and PET have also been used to study the current and developing MAO inhibitor drugs including the selection of doses for clinical trials. In this article, we describe (1) the development of MAO radiotracers; (2) human studies including the relationship of brain MAO levels to genotype, personality, neurological and psychiatric disorders; (3) examples of the use of MAO radiotracers in drug research and development. We will conclude with outstanding needs to improve the radiotracers which are currently used and possible new applications.« less

Semicarbazide-sensitive amine oxidase and extracellular matrix deposition by smooth-muscle cells

NASA Technical Reports Server (NTRS)

Langford, Shannon D.; Trent, Margaret B.; Boor, Paul J.

2002-01-01

We have recently reported in vivo disruption of collagen and elastin architecture within blood vessel walls resulting from the selective inhibition of the enzyme semicarbazide-sensitive amine oxidase (SSAO). This study further investigates the effects of SSAO inhibition on extracellular matrix deposition by smooth-muscle cells (SMCs) cultured from neonatal rat hearts. SMCs were characterized, SSAO activity was measured, and soluble and insoluble collagen and elastin in the extracellular matrix (ECM) were quantified. Cultured neonatal rat heart SMC exhibited a monotypic synthetic phenotype that likely represents a myofibroblast. Detectable levels of SSAO activity present throughout 30-d culture peaked at 7-14 d, coinciding with the production of ECM. The addition of enzyme inhibitors and alternate SSAO substrates (benzylamine) produced varied and, in some cases, marked changes in SSAO activity as well as in the composition of mature and soluble matrix components. Similar to our previous in vivo findings, in vitro SSAO inhibition produced aberrations in collagen and elastin deposition by heart SMC. Because changes in SSAO activity are associated with cardiovascular pathologic states, this enzyme may play a protective or modulating role by regulating ECM production during pathologic insult.

Monoamine oxidase: Radiotracer chemistry and human studies

DOE PAGES

Fowler, Joanna S.; Logan, Jean; Shumay, Elena; ...

2015-03-01

Monoamine oxidase (MAO) oxidizes amines from both endogenous and exogenous sources thereby regulating the concentration of neurotransmitter amines such as serot onin, norepinephrine and dopamine as well as many xenobiotics. MAO inhibitor drugs are used in the treatment of Parkinson’s disease and in depression stimulating the development of radiotracer tools to probe the role of MAO in normal human biology and in disease. Over the past 30 since the first radiotracers were developed and the first PET images of MAO in humans were carried out, PET studies of brain MAO in healthy volunteers and in patients have identified different variablesmore » which have contributed to different MAO levels in brain and in peripheral organs. MAO radiotracers and PET have also been used to study the current and developing MAO inhibitor drugs including the selection of doses for clinical trials. In this article, we describe (1) the development of MAO radiotracers; (2) human studies including the relationship of brain MAO levels to genotype, personality, neurological and psychiatric disorders; (3) examples of the use of MAO radiotracers in drug research and development. We will conclude with outstanding needs to improve the radiotracers which are currently used and possible new applications.« less

Determination of NH2 concentration on 3-aminopropyl tri-ethoxy silane layers and cyclopropylamine plasma polymers by liquid-phase derivatization with 5-iodo 2-furaldehyde

NASA Astrophysics Data System (ADS)

Manakhov, Anton; Čechal, Jan; Michlíček, Miroslav; Shtansky, Dmitry V.

2017-08-01

The quantification of concentration of primary amines, e.g. in plasma polymerized layers is a very important task for surface analysis. However, the commonly used procedure, such as gas phase derivatization with benzaldehydes, shows several drawbacks, the most important of which are the side reaction effects. In the present study we propose and validate a liquid phase derivatization using 5-iodo 2-furaldehyde (IFA). It was demonstrated that the content of NH2 groups can be determined from the atomic concentrations measured by X-ray photoelectron spectroscopy (XPS), in particular from the ratio of I 3d and N 1s peak intensities. First, we demonstrate the method on a prototypical system such as 3-aminopropyl tri-ethoxy silane (APTES) layer. Here the XPS analysis carried out after reaction of APTES layer with IFA gives the fraction of primary amines (NH2/N) of 38.3 ± 7.9%. Comparing this value with that obtained by N 1s curve fitting of APTES layer giving 40.9 ± 9.5% of amine groups, it can be concluded that all primary amines were derivatized by reaction with IFA. The second system to demonstrate the method comprises cyclopropylamine (CPA) plasma polymers that were free from conjugated imines. In this case the method gives the NH2 fraction ∼8.5%. This value is closely matching the NH2/N ratio estimated by 4-trifluoromethyl benzaldehyde (TFBA) derivatization. The reaction of IFA with CPA plasma polymer exhibiting high density of conjugated imines revealed the NH2/N fraction of ∼10.8%. This value was significantly lower compared to 17.3% estimated by TFBA derivatization. As the overestimated density of primary amines measured by TFBA derivatization is probably related to the side reaction of benzaldehydes with conjugated imines, the proposed IFA derivatization of primary amines can be an alternative procedure for the quantification of surface amine groups.

The effect of boron on plasma membrane electron transport and associated proton secretion by cultured carrot cells.

PubMed

Barr, R; Böttger, M; Crane, F L

1993-09-01

Plasma membrane electron transport reactions and associated proton secretion were studied in boron-deficient carrot cells. It was found that the hormone-sensitive plasma membrane NADH oxidase was inhibited by boron deficiency and that under such conditions activity could be restored by exogenous boric acid with or without 2,4-dichlorophenoxy acetic acid. Gramicidin, a channel-forming protonophore, further stimulated NADH oxidase by carrot cells. Proton secretion, associated with plasma membrane H(+)-ATPase, was also affected by boron deficiency, but not as severely as ferricyanide-generated proton secretion, reflecting plasma membrane electron transport. The addition of 1 mM boric acid and 1 microM 2,4-dichlorophenoxy acetic acid to carrot cells fully restored the H+ secretion in presence of ferricyanide. The effect of boron deficiency in cultured carrot cells can, therefore, be directly associated with cell growth through its effect on the plasma membrane NADH oxidase and H+ secretion. Ferricyanide provides a probe which activates transmembrane electron transport that is only coupled to proton release when boron is present.

Method of preparing water purification membranes. [polymerization of allyl amine as thin films in plasma discharge

NASA Technical Reports Server (NTRS)

Hollahan, J. R.; Wydeven, T. J., Jr. (Inventor)

1974-01-01

Allyl amine and chemically related compounds are polymerized as thin films in the presence of a plasma discharge. The monomer compound can be polymerized by itself or in the presence of an additive gas to promote polymerization and act as a carrier. The polymerized films thus produced show outstanding advantages when used as reverse osmosis membranes.

Highly sensitive quantification for human plasma-targeted metabolomics using an amine derivatization reagent.

PubMed

Arashida, Naoko; Nishimoto, Rumi; Harada, Masashi; Shimbo, Kazutaka; Yamada, Naoyuki

2017-02-15

Amino acids and their related metabolites play important roles in various physiological processes and have consequently become biomarkers for diseases. However, accurate quantification methods have only been established for major compounds, such as amino acids and a limited number of target metabolites. We previously reported a highly sensitive high-throughput method for the simultaneous quantification of amines using 3-aminopyridyl-N-succinimidyl carbamate as a derivatization reagent combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Herein, we report the successful development of a practical and accurate LC-MS/MS method to analyze low concentrations of 40 physiological amines in 19 min. Thirty-five of these amines showed good linearity, limits of quantification, accuracy, precision, and recovery characteristics in plasma, with scheduled selected reaction monitoring acquisitions. Plasma samples from 10 healthy volunteers were evaluated using our newly developed method. The results revealed that 27 amines were detected in one of the samples, and that 24 of these compounds could be quantified. Notably, this new method successfully quantified metabolites with high accuracy across three orders of magnitude, with lowest and highest averaged concentrations of 31.7 nM (for spermine) and 18.3 μM (for α-aminobutyric acid), respectively. Copyright © 2016 Elsevier B.V. All rights reserved.

Amine oxidase-based biosensors for spermine and spermidine determination.

PubMed

Boffi, Alberto; Favero, Gabriele; Federico, Rodolfo; Macone, Alberto; Antiochia, Riccarda; Tortolini, Cristina; Sanzó, Gabriella; Mazzei, Franco

2015-02-01

The present work describes the development and optimization of electrochemical biosensors for specific determination of the biogenic polyamine spermine (Spm) and spermidine (Spmd) whose assessment represents a novel important analytical tool in food analysis and human diagnostics. These biosensors have been prepared using novel engineered enzymes: polyamine oxidase (PAO) endowed with selectivity towards Spm and Spmd and spermine oxidase (SMO) characterized by strict specificity towards Spm. The current design entails biosensors in which the enzymes were entrapped in poly(vinyl alcohol) bearing styrylpyridinium groups (PVA-SbQ), a photocrosslinkable gel, onto an electrode surface. Screen-printed electrodes (SPEs) were used as electrochemical transducers for enzymatically produced hydrogen peroxide, operating at different potential vs Ag/AgCl according to the material of the working electrode (WE): +700 mV for graphite (GP) or -100 mV for Prussian blue (PB)-modified SPE, respectively. Biosensor performances were evaluated by means of flow injection amperometric (FIA) measurements. The modified electrodes showed good sensitivity, long-term stability and reproducibility. Under optimal conditions, the PAO biosensor showed a linear range 0.003-0.3 mM for Spm and 0.01-0.4 mM for Spmd, while with the SMO biosensor, a linear range of 0.004-0.5 mM for Spm has been obtained. The main kinetic parameters apparent Michaelis constant (K M), turnover number (K cat) and steady-state current (I max) were determined. The proposed device was then applied to the determination of biogenic amines in blood samples. The results obtained were in good agreement with those obtained with the GC-MS reference method.

pH Dependence of a Mammalian Polyamine Oxidase: Insights into Substrate Specificity and the Role of Lysine 315†

PubMed Central

Pozzi, Michelle Henderson; Gawandi, Vijay; Fitzpatrick, Paul F.

2009-01-01

Mammalian polyamine oxidases (PAO) catalyze the oxidation of N1-acetylspermine and N1-acetylspermidine to produce N-acetyl-3-aminopropanaldehyde and spermidine or putrescine. Structurally, PAO is a member of the monoamine oxidase family of flavoproteins. The effects of pH on kinetic parameters of mouse PAO have been determined to provide insight into the protonation state of the polyamine required for catalysis and the roles of ionizable residues in the active site in amine oxidation. For N1-acetylspermine, N1-acetylspermidine, and spermine, the kcat/Kamine-pH profiles are bell-shaped. In each case the profile agrees with that expected if the productive form of the substrate has a single positively charged nitrogen. The pKi-pH profiles for a series of polyamine analogs are most consistent with the nitrogen at the site of oxidation being neutral and one other nitrogen being positively charged in the reactive form of the substrate. With N1-acetylspermine as substrate, the value of kred, the limiting rate constant for flavin reduction, is pH dependent, decreasing below a pKa value of 7.3, again consistent with the requirement for an uncharged nitrogen for substrate oxidation. Lys315 in PAO corresponds to a conserved active site residue found throughout the monoamine oxidase family. Mutation of Lys315 to methionine has no effect on the kcat/Kamine profile for spermine, the kred value with N1-acetylspermine is only 1.8-fold lower in the mutant protein, and the pKa in the kred-pH profile with N1-acetylspermine shifts to 7.8. These results rule out Lys315 as a source of a pKa in the kcat/Kamine or kcat/kred profiles. They also establish that this residue does not play a critical role in amine oxidation by PAO. PMID:19199575

Piceatannol and resveratrol share inhibitory effects on hydrogen peroxide release, monoamine oxidase and lipogenic activities in adipose tissue, but differ in their antilipolytic properties.

PubMed

Les, Francisco; Deleruyelle, Simon; Cassagnes, Laure-Estelle; Boutin, Jean A; Balogh, Balázs; Arbones-Mainar, José M; Biron, Simon; Marceau, Picard; Richard, Denis; Nepveu, Françoise; Mauriège, Pascale; Carpéné, Christian

2016-10-25

Piceatannol is a hydroxylated derivative of resveratrol. While both dietary polyphenols coexist in edible plants and fruits, and share equivalent concentrations in several wines, the influence of piceatannol on adiposity has been less studied than that of resveratrol. Though resveratrol is now recognized to limit fat deposition in various obesity models, the benefit of its dietary supplementation remains under debate regarding human obesity treatment or prevention. The research for more potent resveratrol analogs is therefore still undergoing. This prompted us to compare various effects of piceatannol and resveratrol directly on human adipose tissue (hAT). Hydrogen peroxide release was measured by Amplex Red-based fluorescence in subcutaneous hAT samples from obese patients. Interactions of stilbenes with human amine oxidases and quinone reductase were assessed by radiometric methods, computational docking and electron paramagnetic resonance. Influences on lipogenic and lipolytic activities were compared in mouse adipocytes. Resveratrol and piceatannol inhibited monoamine oxidase (MAO) with respective IC50 of 18.5 and 133.7 μM, but not semicarbazide-sensitive amine oxidase (SSAO) in hAT. For both stilbenes, the docking scores were better for MAO than for SSAO. Piceatannol and resveratrol similarly hampered hydrogen peroxide detection in assays with and without hAT, while they shared pro-oxidant activities when incubated with purified quinone reductase. They exhibited similar dose-dependent inhibition of adipocyte lipogenic activity. Only piceatannol inhibited basal and stimulated lipolysis when incubated at a dose ≥100 μM. Thus, piceatannol exerted on fat cells dose-dependent effects similar to those of resveratrol, except for a stronger antilipolytic action. In this regard, piceatannol should be useful in limiting the lipotoxicity related to obesity when ingested or administered alone - or might hamper the fat mobilization induced by resveratrol when simultaneously administered with it. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

De novo microdeletion of Xp11.3 exclusively encompassing the monoamine oxidase A and B genes in a male infant with episodic hypotonia: A genomics approach to personalized medicine

PubMed Central

O’Leary, Ryan E.; Shih, Jean C.; Hyland, Keith; Kramer, Nancy; Asher, Y. Jane Tavyev; Graham, John M.

2012-01-01

Monoamine oxidase A and B (MAOA and MAOB) play key roles in deaminating neurotransmitters and various other biogenic amines. Patients deficient in one or both enzymes have distinct metabolic and neurologic profiles. MAOB deficient patients exhibit normal clinical characteristics and behavior, while MAOA deficient patients have borderline intellectual deficiency and impaired impulse control. Patients who lack both MAOA and MAOB have the most extreme laboratory values (urine, blood, and CSF serotonin 4–6 times normal, with elevated O-methylated amine metabolites and reduced deaminated metabolites) in addition to severe intellectual deficiency and behavioral problems. Mice lacking maoa and moab exhibit decreased proliferation of neural stem cells beginning in late gestation and persisting into adulthood These mice show significantly increased monoamine levels, particularly serotonin, as well as anxiety-like behaviors as adults, suggesting that brain maturation in late embryonic development is adversely affected by elevated serotonin levels. We report the case of a male infant with a de novo Xp11.3 microdeletion exclusively encompassing the MAOA and MAOB genes. This newly recognized X-linked disorder is characterized by severe intellectual disability and unusual episodes of hypotonia, which resemble atonic seizures, but have no EEG correlate. A customized low dietary amine diet was implemented in an attempt to prevent the cardiovascular complications that can result from the excessive intake of these compounds. This is the second report of this deletion and the first attempt to maintain the patient’s cardiovascular health through dietary manipulation. Even though a diet low in tyramine, phenylethylamine, and dopa/dopamine is necessary for long-term management, it will not rescue the abnormal monoamine profile seen in combined MAOA and MAOB deficiency. Our patient displays markedly elevated levels of serotonin in blood, serum, urine, and CSF while on this diet. Serotonin biosynthesis inhibitors like para-chlorophenylalanine and p-ethynylphenylalanine may be needed to lower serotonin levels in patients with absent monoamine oxidase enzymes. PMID:22365943

De novo microdeletion of Xp11.3 exclusively encompassing the monoamine oxidase A and B genes in a male infant with episodic hypotonia: a genomics approach to personalized medicine.

PubMed

O'Leary, Ryan E; Shih, Jean C; Hyland, Keith; Kramer, Nancy; Asher, Y Jane Tavyev; Graham, John M

2012-05-01

Monoamine oxidase A and B (MAOA and MAOB) play key roles in deaminating neurotransmitters and various other biogenic amines. Patients deficient in one or both enzymes have distinct metabolic and neurologic profiles. MAOB deficient patients exhibit normal clinical characteristics and behavior, while MAOA deficient patients have borderline intellectual deficiency and impaired impulse control. Patients who lack both MAOA and MAOB have the most extreme laboratory values (urine, blood, and CSF serotonin 4-6 times normal, with elevated O-methylated amine metabolites and reduced deaminated metabolites) in addition to severe intellectual deficiency and behavioral problems. Mice lacking maoa and moab exhibit decreased proliferation of neural stem cells beginning in late gestation and persisting into adulthood. These mice show significantly increased monoamine levels, particularly serotonin, as well as anxiety-like behaviors as adults, suggesting that brain maturation in late embryonic development is adversely affected by elevated serotonin levels. We report the case of a male infant with a de novo Xp11.3 microdeletion exclusively encompassing the MAOA and MAOB genes. This newly recognized X-linked disorder is characterized by severe intellectual disability and unusual episodes of hypotonia, which resemble atonic seizures, but have no EEG correlate. A customized low dietary amine diet was implemented in an attempt to prevent the cardiovascular complications that can result from the excessive intake of these compounds. This is the second report of this deletion and the first attempt to maintain the patient's cardiovascular health through dietary manipulation. Even though a diet low in tyramine, phenylethylamine, and dopa/dopamine is necessary for long-term management, it will not rescue the abnormal monoamine profile seen in combined MAOA and MAOB deficiency. Our patient displays markedly elevated levels of serotonin in blood, serum, urine, and CSF while on this diet. Serotonin biosynthesis inhibitors like para-chlorophenylalanine and p-ethynylphenylalanine may be needed to lower serotonin levels in patients with absent monoamine oxidase enzymes. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Surface composition XPS analysis of a plasma treated polystyrene: Evolution over long storage periods.

PubMed

Ba, Ousmane M; Marmey, Pascal; Anselme, Karine; Duncan, Anthony C; Ponche, Arnaud

2016-09-01

A polystyrene surface (PS) was initially treated by cold nitrogen and oxygen plasma in order to incorporate in particular amine and hydroxyl functions, respectively. The evolution of the chemical nature of the surface was further monitored over a long time period (580 days) by chemical assay, XPS and contact angle measurements. Surface density quantification of primary amine groups was performed using three chemical amine assays: 4-nitrobenzaldehyde (4-NBZ), Sulfo succinimidyl 6-[3'(2 pyridyldithio)-pionamido] hexanoate (Sulfo-LC-SPDP) and iminothiolane (ITL). The results showed amine densities were in the range of 2 per square nanometer (comparable to the results described in the literature) after 5min of nitrogen plasma treatment. Over the time period investigated, chemical assays, XPS and contact angles suggest a drastic significant evolution of the chemical nature of the surface within the first two weeks. Beyond that time period and up to almost two years, nitrogen plasma modified substrates exhibits a slow and continuous oxidation whereas oxygen plasma modifed polystyrene surface is chemically stable after two weeks of storage. The latter appeared to "ease of" showing relatively mild changes within the one year period. Our results suggest that it may be preferable to wait for a chemical "stabilization" period of two weeks before subsequent covalent immobilization of proteins onto the surface. The originality of this work resides in the study of the plasma treated surface chemistry evolution over long periods of storage time (580 days) considerably exceeding those described in the literature. Copyright © 2016 Elsevier B.V. All rights reserved.

Preferential inhibition of the plasma membrane NADH oxidase (NOX) activity by diphenyleneiodonium chloride with NADPH as donor

NASA Technical Reports Server (NTRS)

Morre, D. James

2002-01-01

The cell-surface NADH oxidase (NOX) protein of plant and animal cells will utilize both NADH and NADPH as reduced electron donors for activity. The two activities are distinguished by a differential inhibition by the redox inhibitor diphenyleneiodonium chloride (DPI). Using both plasma membranes and cells, activity with NADPH as donor was markedly inhibited by DPI at submicromolar concentrations, whereas with NADH as donor, DPI was much less effective or had no effect on the activity. The possibility of the inhibition being the result of two different enzymes was eliminated by the use of a recombinant NOX protein. The findings support the concept that NOX proteins serve as terminal oxidases for plasma membrane electron transport involving cytosolic reduced pyridine nucleotides as the natural electron donors and with molecular oxygen as the electron acceptor.

Amine functionalized TiO2-carbon nanotube composite: synthesis, characterization and application to glucose biosensing

NASA Astrophysics Data System (ADS)

Tasviri, Mahboubeh; Rafiee-Pour, Hossain-Ali; Ghourchian, Hedayatollah; Gholami, Mohammad Reza

2011-12-01

The synthesis of amine functionalized TiO2-coated multiwalled carbon nanotubes (NH2-TiO2-CNTs) using sol-gel method was investigated. The synthesized nanocomposite was characterized with XRD, FTIR spectroscopy, BET test and SEM imaging. The results demonstrated a unique nanostructure with no destruction of the CNTs' shape. In addition, the presence of amine groups on the composite surface was confirmed by FTIR. This nanocomposite was used for one-step immobilization of glucose oxidase (GOx) to sense glucose. The result of cyclic voltammetry showed a pair of well-defined and quasi-reversible peaks for direct electron transfer of GOx in the absence of glucose. Also, the result of electrochemical impedance spectroscopy indicated that GOx was successfully immobilized on the surface of NH2-TiO2-CNTs. Furthermore, good amperometric response showed that immobilized GOx on the NH2-TiO2-CNTs exhibits exceptional bioelectrocatalytic activity toward glucose oxidation.

[Monoamine oxidase activity in rat pineal gland: comparison with brain areas, alteration during aging].

PubMed

Razygraev, A V; Taborskaya, K I; Volovik, K Yu; Bunina, A A; Petrosyan, M A

Using benzylamine as a substrate, the amine oxidase activity was determined in the pineal gland of adult rats and compared with the same activity in brain areas and pituitary. Two groups of rats aged 6-8 and 14-15 months were also compared on the basis of this activity. Benzylamine deaminating activity in the pineal gland was significantly higher than in the area preoptica medialis, the corpus mamillare, the tuberculum olfactorium, and the hypophysis, and lower than in the eminentia mediana. The significant increase of the activity in the pineal gland in animals of age from 6-8 to 14-15-months was revealed. Benzylamine deaminating activity in the pineal gland was totally inhibited by 0,002 mM R deprenyl, indicating the B type monoamine oxidase (MAO B) activity. Age-associated increase of MAO B activity in the pineal gland accompanied by decrease of glutathione peroxidase activity, reported earlier, can promote the oxidative damage in the pineal gland during aging.

A Peroxidase-linked Spectrophotometric Assay for the Detection of Monoamine Oxidase Inhibitors

PubMed Central

Zhi, Kangkang; Yang, Zhongduo; Sheng, Jie; Shu, Zongmei; Shi, Yin

2016-01-01

To develop a new more accurate spectrophotometric method for detecting monoamine oxidase inhibitors from plant extracts, a series of amine substrates were selected and their ability to be oxidized by monoamine oxidase was evaluated by the HPLC method and a new substrate was used to develop a peroxidase-linked spectrophotometric assay. 4-(Trifluoromethyl) benzylamine (11) was proved to be an excellent substrate for peroxidase-linked spectrophotometric assay. Therefore, a new peroxidase-linked spectrophotometric assay was set up. The principle of the method is that the MAO converts 11 into aldehyde, ammonia and hydrogen peroxide. In the presence of peroxidase, the hydrogen peroxide will oxidize 4-aminoantipyrine into oxidised 4-aminoantipyrine which can condense with vanillic acid to give a red quinoneimine dye. The production of the quinoneimine dye was detected at 490 nm by a microplate reader. The ⊿OD value between the blank group and blank negative control group in this new method is twice as much as that in Holt’s method, which enables the procedure to be more accurate and avoids the produce of false positive results. The new method will be helpful for researchers to screening monoamine oxidase inhibitors from deep-color plant extracts. PMID:27610153

A Peroxidase-linked Spectrophotometric Assay for the Detection of Monoamine Oxidase Inhibitors.

PubMed

Zhi, Kangkang; Yang, Zhongduo; Sheng, Jie; Shu, Zongmei; Shi, Yin

2016-01-01

To develop a new more accurate spectrophotometric method for detecting monoamine oxidase inhibitors from plant extracts, a series of amine substrates were selected and their ability to be oxidized by monoamine oxidase was evaluated by the HPLC method and a new substrate was used to develop a peroxidase-linked spectrophotometric assay. 4-(Trifluoromethyl) benzylamine (11) was proved to be an excellent substrate for peroxidase-linked spectrophotometric assay. Therefore, a new peroxidase-linked spectrophotometric assay was set up. The principle of the method is that the MAO converts 11 into aldehyde, ammonia and hydrogen peroxide. In the presence of peroxidase, the hydrogen peroxide will oxidize 4-aminoantipyrine into oxidised 4-aminoantipyrine which can condense with vanillic acid to give a red quinoneimine dye. The production of the quinoneimine dye was detected at 490 nm by a microplate reader. The ⊿OD value between the blank group and blank negative control group in this new method is twice as much as that in Holt's method, which enables the procedure to be more accurate and avoids the produce of false positive results. The new method will be helpful for researchers to screening monoamine oxidase inhibitors from deep-color plant extracts.

Spectroscopic studies of the reaction between bovine serum amine oxidase (copper-containing) and some hydrazides and hydrazines.

PubMed Central

Morpurgo, L; Befani, O; Sabatini, S; Mondovì, B; Artico, M; Corelli, F; Massa, S; Stefancich, G; Avigliano, L

1988-01-01

The carbonyl cofactor of bovine serum amine oxidase, recently identified as pyrroloquinoline quinone [Ameyama, Hayashi, Matsushita, Shinagawa & Adachi (1984) Agric. Biol. Chem. 48, 561-565; Lobenstein-Verbeek, Jongejan, Frank & Duine (1984) FEBS Lett. 170, 305-309], reacts stoichiometrically and irreversibly with hydrazides of phenylacetic acid and of benzoic acid. With the phenylacetic hydrazides a reversible intermediate step was detected by competition with substrate, carbonylic reagents or phenylhydrazine, a typical inhibitor of the enzyme. All hydrazides form an intense broad band with maximum absorbance in a narrow wavelength range (350-360 nm), irrespective of the acyl group, suggesting that the transition is located on the organic cofactor. A different situation is found with some phenylhydrazines, where extended conjugation can occur between the cofactor and the phenyl pi-electron system via the azo group, as shown by the lower energy and higher intensity of the transition. In this case the transition is sensitive to substituents in the phenyl ring. The c.d. spectrum of the adducts is influenced by the type of hydrazide (derived from phenylacetic acid or benzoic acid), by pH and by NN-diethyldithiocarbamate binding to copper, probably as a result of shifts of equilibria between hydrazone-azo tautomers. PMID:3146976

High-resolution crystal structure of copper amine oxidase from Arthrobacter globiformis: assignment of bound diatomic molecules as O2.

PubMed

Murakawa, Takeshi; Hayashi, Hideyuki; Sunami, Tomoko; Kurihara, Kazuo; Tamada, Taro; Kuroki, Ryota; Suzuki, Mamoru; Tanizawa, Katsuyuki; Okajima, Toshihide

2013-12-01

The crystal structure of a copper amine oxidase from Arthrobacter globiformis was determined at 1.08 Å resolution with the use of low-molecular-weight polyethylene glycol (LMW PEG; average molecular weight ∼200) as a cryoprotectant. The final crystallographic R factor and Rfree were 13.0 and 15.0%, respectively. Several molecules of LMW PEG were found to occupy cavities in the protein interior, including the active site, which resulted in a marked reduction in the overall B factor and consequently led to a subatomic resolution structure for a relatively large protein with a monomer molecular weight of ∼70,000. About 40% of the presumed H atoms were observed as clear electron densities in the Fo - Fc difference map. Multiple minor conformers were also identified for many residues. Anisotropic displacement fluctuations were evaluated in the active site, which contains a post-translationally derived quinone cofactor and a Cu atom. Furthermore, diatomic molecules, most likely to be molecular oxygen, are bound to the protein, one of which is located in a region that had previously been proposed as an entry route for the dioxygen substrate from the central cavity of the dimer interface to the active site.

Monoamine oxidase inactivation: from pathophysiology to therapeutics.

PubMed

Bortolato, Marco; Chen, Kevin; Shih, Jean C

2008-01-01

Monoamine oxidases (MAOs) A and B are mitochondrial bound isoenzymes which catalyze the oxidative deamination of dietary amines and monoamine neurotransmitters, such as serotonin, norepinephrine, dopamine, beta-phenylethylamine and other trace amines. The rapid degradation of these molecules ensures the proper functioning of synaptic neurotransmission and is critically important for the regulation of emotional behaviors and other brain functions. The byproducts of MAO-mediated reactions include several chemical species with neurotoxic potential, such as hydrogen peroxide, ammonia and aldehydes. As a consequence, it is widely speculated that prolonged excessive activity of these enzymes may be conducive to mitochondrial damages and neurodegenerative disturbances. In keeping with these premises, the development of MAO inhibitors has led to important breakthroughs in the therapy of several neuropsychiatric disorders, ranging from mood disorders to Parkinson's disease. Furthermore, the characterization of MAO knockout (KO) mice has revealed that the inactivation of this enzyme produces a number of functional and behavioral alterations, some of which may be harnessed for therapeutic aims. In this article, we discuss the intriguing hypothesis that the attenuation of the oxidative stress induced by the inactivation of either MAO isoform may contribute to both antidepressant and antiparkinsonian actions of MAO inhibitors. This possibility further highlights MAO inactivation as a rich source of novel avenues in the treatment of mental disorders.

Toxicokinetic, toxicodynamic, and toxicoproteomic aspects of short-term exposure to trenbolone in female fish.

PubMed

Schultz, Irvin R; Nagler, James J; Swanson, Penny; Wunschel, Dave; Skillman, Ann D; Burnett, Vicki; Smith, Derek; Barry, Richard

2013-12-01

The toxicokinetics of trenbolone was characterized during 500 ng/l water exposures in female rainbow trout (Oncorhynchus mykiss) and fathead minnows (Pimephales promelas). Related experiments measured various toxicodynamic effects of exposure. In both species, trenbolone was rapidly absorbed from the water and reached peak plasma levels within 8h of exposure. Afterwards, trenbolone concentrations in trout (66-95 ng/ml) were 2-6 times higher compared with minnows (15-29 ng/ml), which was attributable to greater plasma binding in trout. During water exposures, circulating levels of estradiol (E2) rapidly decreased in both species to a concentration that was 25%-40% of control values by 8-24h of exposure and then remained relatively unchanged for the subsequent 6 days of exposure. In trout, changes in circulating levels of follicle-stimulating hormone were also significantly greater after trenbolone exposure, relative to controls. In both species, the pharmacokinetics of injected E2-d3 was altered by trenbolone exposure with an increase in total body clearance and a corresponding decrease in elimination half-life. The unbound percentage of E2 in trout plasma was 0.25%, which was similar in pre- or postvitellogenic female trout. Subsequent incubation with trenbolone caused the unbound percentage to significantly increase to 2.4% in the previtellogenic trout plasma. iTRAQ-based toxicoproteomic studies in minnows exposed to 5, 50, and 500 ng/l trenbolone identified a total of 148 proteins with 19 downregulated including vitellogenin and 18 upregulated. Other downregulated proteins were fibrinogens, α-2-macroglobulin, and transferrin. Upregulated proteins included amine oxidase, apolipoproteins, parvalbumin, complement system proteins, and several uncharacterized proteins. The results indicate trenbolone exposure is a highly dynamic process in female fish with uptake and tissue equilibrium quickly established, leading to both rapid and delayed toxicodynamic effects.

Redox inactive metal ion triggered N-dealkylation by an iron catalyst with dioxygen activation: a lesson from lipoxygenases.

PubMed

Zhang, Jisheng; Wang, Yujuan; Luo, Nengchao; Chen, Zhuqi; Wu, Kangbing; Yin, Guochuan

2015-06-07

Utilization of dioxygen as the terminal oxidant at ambient temperature is always a challenge in redox chemistry, because it is hard to oxidize a stable redox metal ion like iron(III) to its high oxidation state to initialize the catalytic cycle. Inspired by the dioxygenation and co-oxidase activity of lipoxygenases, herein, we introduce an alternative protocol to activate the sluggish iron(III) species with non-redox metal ions, which can promote its oxidizing power to facilitate substrate oxidation with dioxygen, thus initializing the catalytic cycle. In oxidations of N,N-dimethylaniline and its analogues, adding Zn(OTf)2 to the [Fe(TPA)Cl2]Cl catalyst can trigger the amine oxidation with dioxygen, whereas [Fe(TPA)Cl2]Cl alone is very sluggish. In stoichiometric oxidations, it has also been confirmed that the presence of Zn(OTf)2 can apparently improve the electron transfer capability of the [Fe(TPA)Cl2]Cl complex. Experiments using different types of substrates as trapping reagents disclosed that the iron(IV) species does not occur in the catalytic cycle, suggesting that oxidation of amines is initialized by electron transfer rather than hydrogen abstraction. Combined experiments from UV-Vis, high resolution mass spectrometry, electrochemistry, EPR and oxidation kinetics support that the improved electron transfer ability of iron(III) species originates from its interaction with added Lewis acids like Zn(2+) through a plausible chloride or OTf(-) bridge, which has promoted the redox potential of iron(III) species. The amine oxidation mechanism was also discussed based on the available data, which resembles the co-oxidase activity of lipoxygenases in oxidative dealkylation of xenobiotic metabolisms where an external electron donor is not essential for dioxygen activation.

Benzylamine antihyperglycemic effect is abolished by AOC3 gene invalidation in mice but not rescued by semicarbazide-sensitive amine oxidase expression under the control of aP2 promoter.

PubMed

Grès, Sandra; Bour, Sandy; Valet, Philippe; Carpéné, Christian

2012-12-01

Semicarbazide-sensitive amine oxidase (SSAO) is a transmembrane enzyme that metabolizes primary amines from endogenous or dietary origin. SSAO is highly expressed in adipose, smooth muscle and endothelial cells. In each of these cell types, SSAO is implicated in different biological functions, such as glucose transport activation, extracellular matrix maturation and leucocyte extravasation, respectively. However, the physiological functions of SSAO and their involvement in pathogenesis remain uncompletely characterized. To better understand the role of adipose tissue SSAO, we investigated whether it was necessary and/or sufficient to produce the antihyperglycemic effect of the SSAO-substrate benzylamine, already reported in mice. Therefore, we crossed SSAO-deficient mice invalidated for AOC3 gene and transgenic mice expected to express human SSAO in an adipocyte-specific manner, under the control of aP2 promoter. The aP2-human AOC3 construct (aP2-hAOC3) was equally expressed in the adipose tissue of mice expressing or not the native murine form and almost absent in other tissues. However, the corresponding SSAO activity found in adipose tissue represented only 20 % that of control mice. As a consequence, the benzylamine antihyperglycemic effect observed during glucose tolerance test in control was abolished in AOC3-KO mice but not rescued in mice expressing aP2-hAOC3. The capacity of benzylamine or methylamine to activate glucose uptake in adipocytes exhibited parallel variations in the corresponding genotypes. Although the aP2-hAOC3 construct did not allow a total rescue of SSAO activity in adipose tissue, it could be assessed from our observations that adipocyte SSAO plays a pivotal role in the increased glucose tolerance promoted by pharmacological doses of benzylamine.

Mechanism of Flavoprotein l-6-Hydroxynicotine Oxidase: pH and Solvent Isotope Effects and Identification of Key Active Site Residues.

PubMed

Fitzpatrick, Paul F; Chadegani, Fatemeh; Zhang, Shengnan; Dougherty, Vi

2017-02-14

The flavoenzyme l-6-hydroxynicotine oxidase is a member of the monoamine oxidase family that catalyzes the oxidation of (S)-6-hydroxynicotine to 6-hydroxypseudooxynicotine during microbial catabolism of nicotine. While the enzyme has long been understood to catalyze oxidation of the carbon-carbon bond, it has recently been shown to catalyze oxidation of a carbon-nitrogen bond [Fitzpatrick, P. F., et al. (2016) Biochemistry 55, 697-703]. The effects of pH and mutagenesis of active site residues have now been utilized to study the mechanism and roles of active site residues. Asn166 and Tyr311 bind the substrate, while Lys287 forms a water-mediated hydrogen bond with flavin N5. The N166A and Y311F mutations result in ∼30- and ∼4-fold decreases in k cat /K m and k red for (S)-6-hydroxynicotine, respectively, with larger effects on the k cat /K m value for (S)-6-hydroxynornicotine. The K287M mutation results in ∼10-fold decreases in these parameters and a 6000-fold decrease in the k cat /K m value for oxygen. The shapes of the pH profiles are not altered by the N166A and Y311F mutations. There is no solvent isotope effect on the k cat /K m value for amines. The results are consistent with a model in which both the charged and neutral forms of the amine can bind, with the former rapidly losing a proton to a hydrogen bond network of water and amino acids in the active site prior to the transfer of hydride to the flavin.

Reactive oxygen species spermine metabolites generated from amine oxidases and radiation represent a therapeutic gain in cancer treatments.

PubMed

Amendola, Roberto; Cervelli, Manuela; Fratini, Emiliano; Sallustio, Davide E; Tempera, Giampiero; Ueshima, Taichi; Mariottini, Paolo; Agostinelli, Enzo

2013-09-01

The most frequent interventions in cancer therapy are currently the destruction of cells by irradiation or administration of drugs both able to induce radical formation and toxic metabolites by enzyme-catalyzed reactions. The aim of this study was to determine the cell viability of cells undergoing a DNA damage threshold accomplished by ROS overproduction via both ectopic expression of murine spermine oxidase (mSMOX) and bovine serum amine oxidase (BSAO) enzymes. Low dose of X-irradiation delivers a challenging dose of damage as evaluated in proficient Chinese hamster AA8 cell line and both deficient transcription-coupled nucleotide excision repair (NER) UV61 cells and deficient base excision repair (BER) EM9 cells, at 6 and 24 h after exposure. The priming dose of ROS overexposure by mSMOX provokes an adaptive response in N18TG2, AA8 and EM9 cell lines at 24 h. Interestingly, in the UV61 cells, ROS overexposure by mSMOX delivers an earlier adaptive response to radiation. The enzymatic formation of toxic metabolites has mainly been investigated on wild-type (WT) and multidrug-resistant (MDR) cancer cell lines, using and spermine as substrate of the BSAO enzyme. MDR cells are more sensitive to the toxic polyamine metabolites than WT cells, thus indicating a new therapeutic strategy to overcome MDR tumors. Since SMOX in mammals is differentially activated in a tissue-specific manner and cancer cells can differ in terms of DNA repair and MDR capabilities, it could be of interest to simultaneously treat with very low dose of X-rays and/or to alter SMOX metabolism to generate a differential response in healthy and cancer tissues.

5-hydroxytryptamine actions in adipocytes: involvement of monoamine oxidase-dependent oxidation and subsequent PPARγ activation.

PubMed

Grès, Sandra; Gomez-Zorita, Saioa; Gomez-Ruiz, Ana; Carpéné, Christian

2013-06-01

Serotonin (5-HT) is a brain neurotransmitter instrumental for the antidepressant action of selective inhibitors of serotonin reuptake (SSRIs) while it also plays important roles in peripheral organs. Recently, the 5-HT oxidation products, 5-hydroxyindoleacetate and 5-methoxy-indoleacetate, have been shown to bind to peroxisome proliferator-activated receptor γ (PPARγ) and to enhance lipid accumulation in preadipocytes. Since we already reported that adipocytes exhibit elevated monoamine oxidase (MAO) and primary amine oxidase activities, we verified how adipocytes readily oxidize 5-HT, with the objective to determine whether such oxidation promotes PPARγ activation and lipid storage. To this aim, serotonin was tested on cultured 3T3 F442A preadipocytes and on human adipocytes. Results showed that 5-HT was oxidized by MAO in both models. Daily treatment of 3T3 F442A preadipocytes for 8 days with 100-500 μM 5-HT promoted triglyceride accumulation and emergence of adipogenesis markers. At 250 μM, 5-HT alone reproduced half of 50 nM insulin-induced adipogenesis, and exhibited an additive differentiating effect when combined with insulin. Moreover, the 5-HT-induced expression of PPARγ-responsive genes (PEPCK, aP2/FABP4) was blocked by GW 9662, a PPARγ-inhibitor, or by pargyline, a MAO-inhibitor. In human fat cells, 6-h exposure to 100 μM 5-HT increased PEPCK expression as did the PPARγ-agonist rosiglitazone. Since hydrogen peroxide, another amine oxidation product, did not reproduce such enhancement, we propose that serotonin can promote PPARγ activation in fat cells, via the indoleacetate produced during MAO-dependent oxidation. Such pathway could be involved in the adverse effects of several antidepressant SSRIs on body weight gain.

Cholesterol transport from plasma membranes to intracellular membranes is inhibited by 3 beta-[2-(diethylamino)ethoxy]androst-5-en-17-one.

PubMed

Härmälä, A S; Pörn, M I; Mattjus, P; Slotte, J P

1994-03-24

The compound U1866A (3 beta-[2-(diethylamino)ethoxy]androst-5-en-17-one) has been shown to inhibit the cellular transfer of low-density lipoprotein-derived cholesterol from lysosomes to plasma membranes (Liscum and Faust (1989) J. Biol. Chem. 264, 11796-806). We have in this study examined the effects of U18666A on cholesterol translocation from plasma membranes to intracellular membranes. Translocation of plasma membrane cholesterol was induced by degradation of plasma membrane sphingomyelin. The sphingomyelinase-induced activation of the acyl-CoA cholesterol acyl transferase (ACAT) reaction was completely inhibited in a dose-dependent manner by U18666A, both in cultured human skin fibroblasts and baby hamster kidney cells. Half-maximal inhibition (within 60 min) was obtained with 0.5-1 microgram/ml of U18666A. A time-course study indicated that the onset of inhibition was rapid (within 10-15 min), and reversible if U18666A was removed from the incubation mixture. Using a cholesterol oxidase assay, we observed that the extent of plasma membrane cholesterol translocation in sphingomyelinase-treated HSF cells was significantly lowered in the presence of U18666A (at 3 micrograms/ml). The effect of U18666A on cholesterol translocation was also fully reversible when the drug was withdrawn. In mouse Leydig tumor cells, labeled to constant specific activity with [3H]cholesterol, the compound U18666A inhibited in a dose-dependent manner the cyclic AMP-stimulated secretion of [3H]steroid hormones. The effects seen with compound U18666A appeared to be specific for this molecule, since another hydrophobic amine, imipramine, did not in our experiments affect cholesterol translocation or ACAT activation. Since different cell types display sensitivity to U18666A in various intracellular cholesterol transfer processes, they appear to have a common U18666A-sensitive regulatory mechanism.

Exploiting algal NADPH oxidase for biophotovoltaic energy

DOE PAGES

Anderson, Alexander; Laohavisit, Anuphon; Blaby, Ian K.; ...

2015-01-29

Photosynthetic microbes exhibit light-dependent electron export across the cell membrane, which can generate electricity in biological photovoltaic (BPV) devices. How electrons are exported remains to be determined; the identification of mechanisms would help selection or generation of photosynthetic microbes capable of enhanced electrical output. We show that plasma membrane NADPH oxidase activity is a significant component of light-dependent generation of electricity by the unicellular green alga Chlamydomonas reinhardtii. NADPH oxidases export electrons across the plasma membrane to form superoxide anion from oxygen. The C. reinhardtii mutant lacking the NADPH oxidase encoded by RBO1 is impaired in both extracellular superoxide anionmore » production and current generation in a BPV device. Complementation with the wild-type gene restores both capacities, demonstrating the role of the enzyme in electron export. Monitoring light-dependent extracellular superoxide production with a colorimetric assay is shown to be an effective way of screening for electrogenic potential of candidate algal strains. Furthermore, the results show that algal NADPH oxidases are important for superoxide anion production and open avenues for optimizing the biological component of these devices.« less

Auxin-activated NADH oxidase activity of soybean plasma membranes is distinct from the constitutive plasma membrane NADH oxidase and exhibits prion-like properties

NASA Technical Reports Server (NTRS)

Morre, D. James; Morre, Dorothy M.; Ternes, Philipp

2003-01-01

The hormone-stimulated and growth-related cell surface hydroquinone (NADH) oxidase activity of etiolated hypocotyls of soybeans oscillates with a period of about 24 min or 60 times per 24-h day. Plasma membranes of soybean hypocotyls contain two such NADH oxidase activities that have been resolved by purification on concanavalin A columns. One in the apparent molecular weight range of 14-17 kDa is stimulated by the auxin herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The other is larger and unaffected by 2,4-D. The 2,4-D-stimulated activity absolutely requires 2,4-D for activity and exhibits a period length of about 24 min. Also exhibiting 24-min oscillations is the rate of cell enlargement induced by the addition of 2,4-D or the natural auxin indole-3-acetic acid (IAA). Immediately following 2,4-D or IAA addition, a very complex pattern of oscillations is frequently observed. However, after several hours a dominant 24-min period emerges at the expense of the constitutive activity. A recruitment process analogous to that exhibited by prions is postulated to explain this behavior.

Bacterial Toxicity and Metabolism of Three Hydrazine Fuels.

DTIC Science & Technology

1980-09-01

4- LO 0z V) 8 LL Di: 0 0J E - N z 4 0~ 0 0 44 4’a 44~ C -0 0 0 0~ N 0 coQ NoIi ~ (ADP-SSi 6w/N- HN bw ’ap, uoipOzijInr apihssqns) 3SNOaS3 i %/ 77...irreversible bond with an amine oxidase enzyme (Reference 49). To test the significance of this binding, an experiment was conducted with the

Surface functionalization of microwave plasma-synthesized silica nanoparticles for enhancing the stability of dispersions

NASA Astrophysics Data System (ADS)

Sehlleier, Yee Hwa; Abdali, Ali; Schnurre, Sophie Marie; Wiggers, Hartmut; Schulz, Christof

2014-08-01

Gas phase-synthesized silica nanoparticles were functionalized with three different silane coupling agents (SCAs) including amine, amine/phosphonate and octyltriethoxy functional groups and the stability of dispersions in polar and non-polar dispersing media such as water, ethanol, methanol, chloroform, benzene, and toluene was studied. Fourier transform infrared spectroscopy showed that all three SCAs are chemically attached to the surface of silica nanoparticles. Amine-functionalized particles using steric dispersion stabilization alone showed limited stability. Thus, an additional SCA with sufficiently long hydrocarbon chains and strong positively charged phosphonate groups was introduced in order to achieve electrosteric stabilization. Steric stabilization was successful with hydrophobic octyltriethoxy-functionalized silica nanoparticles in non-polar solvents. The results from dynamic light scattering measurements showed that in dispersions of amine/phosphonate- and octyltriethoxy-functionalized silica particles are dispersed on a primary particle level. Stable dispersions were successfully prepared from initially agglomerated nanoparticles synthesized in a microwave plasma reactor by designing the surface functionalization.

High performance liquid chromatography with UV detection for the simultaneous determination of sympathomimetic amines using 4-(4,5-diphenyl-1H-imidazole-2-yl)benzoyl chloride as a label.

PubMed

Kaddoumi, A; Kubota, A; Nakashima, M N; Takahashi, M; Nakashima, K

2001-10-01

A high performance liquid chromatographic method has been developed for the simultaneous determination of (+/-) fenfluramine (Fen) and phentermine (Phen) in addition to three other sympathomimetic amines-ephedrine (E), norephedrine (NE) and 2-phenylethylamine (2-PEA), using cyclohexylamine (CX) as an internal standard in plasma. The compounds were derivatized with 4-(4,5-diphenyl-1H-imidazole-2-yl)benzoyl chloride (DIB-Cl) to give the DIB-derivatives. The derivatives were then separated using an isocratic HPLC system with UV detection. The limits of detection for Fen, Phen, E, NE and 2-PEA in plasma ranged from 0.32 to 22.9 pmol on column at a signal-to-noise ratio of 3. The recoveries following alkaline extraction from plasma samples of known concentrations were found to be more than 94% for the studied compounds. This method might be useful for the screening of the studied sympathomimetic amines in human plasma samples in forensic as well as toxicological studies. Furthermore, the developed method was modified for the simultaneous determination of Fen and Phen in human and rat plasma using fluoxetine as an internal standard. The methods are reproducible and precise. Finally, the two drugs were administered intraperitoneally to rats in combination, and their plasma levels over the investigated time course were successfully determined. Copyright 2001 John Wiley & Sons, Ltd.

Multivalent Interactions of Human Primary Amine Oxidase with the V and C22 Domains of Sialic Acid-Binding Immunoglobulin-Like Lectin-9 Regulate Its Binding and Amine Oxidase Activity

PubMed Central

Fair-Mäkelä, Ruth; Salo-Ahen, Outi M. H.; Guédez, Gabriela; Bligt-Lindén, Eva; Grönholm, Janne; Jalkanen, Sirpa; Salminen, Tiina A.

2016-01-01

Sialic acid-binding immunoglobulin-like lectin-9 (Siglec-9) on leukocyte surface is a counter-receptor for endothelial cell surface adhesin, human primary amine oxidase (hAOC3), a target protein for anti-inflammatory agents. This interaction can be used to detect inflammation and cancer in vivo, since the labeled peptides derived from the second C2 domain (C22) of Siglec-9 specifically bind to the inflammation-inducible hAOC3. As limited knowledge on the interaction between Siglec-9 and hAOC3 has hampered both hAOC3-targeted drug design and in vivo imaging applications, we have now produced and purified the extracellular region of Siglec-9 (Siglec-9-EC) consisting of the V, C21 and C22 domains, modeled its 3D structure and characterized the hAOC3–Siglec-9 interactions using biophysical methods and activity/inhibition assays. Our results assign individual, previously unknown roles for the V and C22 domains. The V domain is responsible for the unusually tight Siglec-9–hAOC3 interactions whereas the intact C22 domain of Siglec-9 is required for modulating the enzymatic activity of hAOC3, crucial for the hAOC3-mediated leukocyte trafficking. By characterizing the Siglec-9-EC mutants, we could conclude that R120 in the V domain likely interacts with the terminal sialic acids of hAOC3 attached glycans whereas residues R284 and R290 in C22 are involved in the interactions with the active site channel of hAOC3. Furthermore, the C22 domain binding enhances the enzymatic activity of hAOC3 although the sialic acid-binding capacity of the V domain of Siglec-9 is abolished by the R120S mutation. To conclude, our results prove that the V and C22 domains of Siglec-9-EC interact with hAOC3 in a multifaceted and unique way, forming both glycan-mediated and direct protein-protein interactions, respectively. The reported results on the mechanism of the Siglec-9–hAOC3 interaction are valuable for the development of hAOC3-targeted therapeutics and diagnostic tools. PMID:27893774

Phenethylamines in brain and liver of rats with experimentally induced phenylketonuria-like characteristics

PubMed Central

Edwards, David J.; Blau, Karl

1973-01-01

1. Phenethylamines were extracted from brain and liver of rats with phenylketonuria-like characteristics produced in vivo by inhibition of phenylalanine hydroxylase (EC 1.14.3.1) with p-chlorophenylalanine, with or without phenylalanine administration. To protect amines against oxidation by monoamine oxidase, pargyline was also administered. 2. β-Phenethylamine was the major compound found in brain and liver. β-Phenethanolamine and octopamine were also present, in lesser amounts, and the concentrations of these three amines paralleled blood phenylalanine concentrations. By comparison, tissues from control animals had only very low concentrations of these amines. 3. Small amounts of normetadrenaline, m-tyramine and 3-methoxytyramine were also found. 4. The inhibitors used, p-chlorophenylalanine and pargyline, gave rise to p-chlorophenethylamine and benzylamine respectively, the first via decarboxylation, the second probably by breakdown during extraction. 5. Distribution of phenethylamines in different brain regions and in subcellular fractions of rat brain cells was also investigated. The content of phenethylamine was highest in the striatum. 6. These findings are discussed in the light of changes occurring in human patients with uncontrolled phenylketonuria. PMID:4269184

Crystal structure of human lysyl oxidase-like 2 (hLOXL2) in a precursor state.

PubMed

Zhang, Xi; Wang, Qifan; Wu, Jianping; Wang, Jiawei; Shi, Yigong; Liu, Minhao

2018-04-10

Lysyl oxidases (LOXs), a type of copper- and lysyl tyrosylquinone (LTQ) -dependent amine oxidase, catalyze the oxidative deamination of lysine residues of extracellular matrix (ECM) proteins such as elastins and collagens and generate aldehyde groups. The oxidative deamination of lysine represents the foundational step for the cross-linking of elastin and collagen and thus is crucial for ECM modeling. Despite their physiological significance, the structure of this important family of enzymes remains elusive. Here we report the crystal structure of human lysyl oxidase-like 2 (hLOXL2) at 2.4-Å resolution. Unexpectedly, the copper-binding site of hLOXL2 is occupied by zinc, which blocks LTQ generation and the enzymatic activity of hLOXL2 in our in vitro assay. Biochemical analysis confirms that copper loading robustly activates hLOXL2 and supports LTQ formation. Furthermore, the LTQ precursor residues in the structure are distanced by 16.6 Å, corroborating the notion that the present structure may represent a precursor state and that pronounced conformational rearrangements would be required for protein activation. The structure presented here establishes an important foundation for understanding the structure-function relationship of LOX proteins and will facilitate LOX-targeting drug discovery. Copyright © 2018 the Author(s). Published by PNAS.

Reverse-osmosis membranes by plasma polymerization

NASA Technical Reports Server (NTRS)

Hollahan, J. R.; Wydeven, T.

1972-01-01

Thin allyl amine polymer films were developed using plasma polymerization. Resulting dry composite membranes effectively reject sodium chloride during reverse osmosis. Films are 98% sodium chloride rejective, and 46% urea rejective.

Phosphatidylinositol 3-Kinase Plays a Vital Role in Regulation of Rice Seed Vigor via Altering NADPH Oxidase Activity

PubMed Central

Liu, Jian; Zhou, Jun; Xing, Da

2012-01-01

Phosphatidylinositol 3-kinase (PI3K) has been reported to be important in normal plant growth and stress responses. In this study, it was verified that PI3K played a vital role in rice seed germination through regulating NADPH oxidase activity. Suppression of PI3K activity by inhibitors wortmannin or LY294002 could abate the reactive oxygen species (ROS) formation, which resulted in disturbance to the seed germination. And then, the signal cascades that PI3K promoted the ROS liberation was also evaluated. Diphenylene iodonium (DPI), an NADPH oxidase inhibitor, suppressed most of ROS generation in rice seed germination, which suggested that NADPH oxidase was the main source of ROS in this process. Pharmacological experiment and RT-PCR demonstrated that PI3K promoted the expression of Os rboh9. Moreover, functional analysis by native PAGE and the measurement of the 2, 3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazo-lium-5- carboxanilide (XTT) formazan concentration both showed that PI3K promoted the activity of NADPH oxidase. Furthermore, the western blot analysis of OsRac-1 demonstrated that the translocation of Rac-1 from cytoplasm to plasma membrane, which was known as a key factor in the assembly of NADPH oxidase, was suppressed by treatment with PI3K inhibitors, resulting in the decreased activity of NADPH oxidase. Taken together, these data favored the novel conclusion that PI3K regulated NADPH oxidase activity through modulating the recruitment of Rac-1 to plasma membrane and accelerated the process of rice seed germination. PMID:22448275

Role of adrenal hormones in the synthesis of noradrenaline in cardiac sympathetic neurones

PubMed Central

Bhagat, B.

1969-01-01

1. Adrenalectomy or adrenal demedullation affected neither the levels of endogenous catecholamines in the rat heart nor the accumulation of 3H-noradrenaline 1 hr after its intravenous administration. 2. Twenty-four hours after intravenous administration of labelled amine, however, its retention was markedly reduced in the heart of adrenalectomized or demedullated rats. Ganglionic blockade prevented this reduction. 3. Rate calculations from the decline of catecholamine levels after blockade of synthesis with α-methyl-tyrosine showed that cardiac synthesis of noradrenaline increased about four-fold after demedullation and about three-fold after adrenalectomy. This increase in synthesis may compensate for the loss of circulating catecholamines. 4. There was no change in catechol-o-methyl-transferase activity, but monoamine oxidase activity was increased in the homogenates of the heart of adrenalectomized and demedullated rats. The increase in the cardiac monoamine oxidase activity was markedly greater in the adrenalectomized rats than in the demedullated rats. 5. It is suggested that adrenal cortex insufficiency may modulate the rate of synthesis of noradrenaline and monoamine oxidase activity in cardiac sympathetic neurones. PMID:5360339

Dithiocarbamates are teratogenic to developing zebrafish through inhibition of lysyl oxidase activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boxtel, Antonius L. van, E-mail: thijs.van.boxtel@ivm.vu.n; Kamstra, Jorke H.; Fluitsma, Donna M.

2010-04-15

Dithiocarbamates (DTCs) are a class of compounds that are extensively used in agriculture as pesticides. As such, humans and wildlife are undoubtedly exposed to these chemicals. Although DTCs are thought to be relatively safe due to their short half lives, it is well established that they are teratogenic to vertebrates, especially to fish. In zebrafish, these teratogenic effects are characterized by distorted notochord development and shortened anterior to posterior axis. DTCs are known copper (Cu) chelators but this does not fully explain the observed teratogenic effects. We show here that DTCs cause malformations in zebrafish that highly resemble teratogenic effectsmore » observed by direct inhibition of a group of cuproenzymes termed lysyl oxidases (LOX). Additionally, we demonstrate that partial knockdown of three LOX genes, lox, loxl1 and loxl5b, sensitizes the developing embryo to DTC exposure. Finally, we show that DTCs directly inhibit zebrafish LOX activity in an ex vivo amine oxidase assay. Taken together, these results provide the first evidence that DTC induced teratogenic effects are, at least in part, caused by direct inhibition of LOX activity.« less

Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of variants of monoamine oxidase from Aspergillus niger

DOE Office of Scientific and Technical Information (OSTI.GOV)

Atkin, Kate E.; Reiss, Renate; Turner, Nicholas J.

2008-03-01

Crystals of A. niger monoamine oxidase variants display P2{sub 1} or P4{sub 1}2{sub 1}2/P4{sub 3}2{sub 1}2 symmetry, with eight or two molecules in the asymmetric unit, respectively. Monoamine oxidase from Aspergillus niger (MAO-N) is an FAD-dependent enzyme that catalyses the conversion of terminal amines to their corresponding aldehydes. Variants of MAO-N produced by directed evolution have been shown to possess altered substrate specificity. Crystals of two of these variants (MAO-N-3 and MAO-N-5) have been obtained; the former displays P2{sub 1} symmetry with eight molecules per asymmetric unit and the latter has P4{sub 1}2{sub 1}2 or P4{sub 3}2{sub 1}2 symmetry andmore » two molecules per asymmetric unit. Solution of these structures will help shed light on the molecular determinants of improved activity and high enantioselectivity towards a broad range of substrates.« less

A Polyamine Oxidizing Enzyme as a Drug to Treat Breast Cancer

DTIC Science & Technology

2007-07-01

is a viable anticancer therapy. BODY TASK 1. Prepare enough of two polyethylene glycol(PEG)-derivatives of bovine serum amine oxidase (SAO...interfere with its PEGylation and/or obscure the outcome of experiments to test the treatment as an anticancer therapy. Hence, another few weeks were...attached polysaccharide , thus, minimize interference with or mimicking of the mouse’s own SAO. A small amount of bovine SAO was PEGylated at pH 7.4 with

Co-segregation of Norrie disease and idiopathic pulmonary hypertension in a family with a microdeletion of the NDP region at Xp11.3-p11.4.

PubMed

Staropoli, John F; Xin, Winnie; Sims, Katherine B

2010-11-01

Norrie disease is a rare X-linked congenital retinal vasculopathy that may be accompanied by sensorineural deafness, mental retardation, and other neurological deficits. Here we present a family in which Norrie disease co-segregated with either early-onset idiopathic pulmonary hypertension or sudden death preceded by a period of progressive dyspnea. Neither Norrie disease, nor its atypical variants described to date, have been associated with this extended clinical phenotype. Molecular analysis of the Norrie disease gene (NDP) and adjacent loci was performed by multiplex ligation-dependent probe amplification and comparative genomic hybridisation. Affected males in this family showed an inherited hemizygous deletion restricted to NDP and two immediately telomeric genes, monoamine oxidase-B (MAO-B) and monoamine oxidase-A (MAO-A), which encode closely related enzymes that metabolize biogenic amines including serotonin, dopamine, and norepinephrine. Sequencing of the deletion junction showed an unusual pattern in which a region of microhomology flanked intervening genomic sequence. Because abnormalities of biogenic amines, particularly serotonin, have been implicated in the pathophysiology of pulmonary hypertension, we propose that presumed MAO deficiency in these patients may represent a novel risk factor for pulmonary hypertension, particularly forms with very early onset. Fine-mapping of other microdeletions at this locus may provide insights into additional mechanisms for nonrecurrent genomic rearrangements at this and other chromosomal loci.

Expression and functional analysis of the lysine decarboxylase and copper amine oxidase genes from the endophytic fungus Colletotrichum gloeosporioides ES026.

PubMed

Zhang, Xiangmei; Wang, Zhangqian; Jan, Saad; Yang, Qian; Wang, Mo

2017-06-05

Huperzine A (HupA) isolated from Huperzia serrata is an important compound used to treat Alzheimer's disease (AD). Recently, HupA was reported in various endophytic fungi, with Colletotrichum gloeosporioides ES026 previously isolated from H. serrata shown to produce HupA. In this study, we performed next-generation sequencing and de novo RNA sequencing of C. gloeosporioides ES026 to elucidate the molecular functions, biological processes, and biochemical pathways of these unique sequences. Gene ontology and Kyoto Encyclopedia of Genes and Genomes assignments allowed annotation of lysine decarboxylase (LDC) and copper amine oxidase (CAO) for their conversion of L-lysine to 5-aminopentanal during HupA biosynthesis. Additionally, we constructed a stable, high-yielding HupA-expression system resulting from the overexpression of CgLDC and CgCAO from the HupA-producing endophytic fungus C. gloeosporioides ES026 in Escherichia coli. Quantitative reverse transcription polymerase chain reaction analysis confirmed CgLDC and CgCAO expression, and quantitative determination of HupA levels was assessed by liquid chromatography high-resolution mass spectrometry, which revealed that elevated expression of CgLDC and CgCAO produced higher yields of HupA than those derived from C. gloeosporioides ES026. These results revealed CgLDC and CgCAO involvement in HupA biosynthesis and their key role in regulating HupA content in C. gloeosporioides ES026.

Levels of plasma ceruloplasmin protein are markedly lower following dietary copper deficiency in rodents

PubMed Central

Broderius, Margaret; Mostad, Elise; Wendroth, Krista; Prohaska, Joseph R.

2010-01-01

Ceruloplasmin (Cp) is a multicopper oxidase and the most abundant copper binding protein in vertebrate plasma. Loss of function mutations in humans or experimental deletion in mice result in iron overload consistent with a putative ferroxidase function. Prior work suggested plasma may contain multiple ferroxidases. Studies were conducted in Holtzman rats (Rattus novegicus), albino mice (Mus musculus), Cp -/- mice, and adult humans (Homo sapiens) to investigate the copper-iron interaction. Dietary copper-deficient (CuD) rats and mice were produced using a modified AIN-76A diet. Results confirmed that o-dianisidine is a better substrate than paraphenylene diamine (PPD) for assessing diamine oxidase activity of Cp. Plasma from CuD rat dams and pups, and CuD and Cp -/- mice contained no detectable Cp diamine oxidase activity. Importantly, no ferroxidase activity was detectable for CuD rats, mice, or Cp -/- mice compared to robust activity for copper-adequate (CuA) rodent controls using western membrane assay. Immunoblot protocols detected major reductions (60-90%) in Cp protein in plasma of CuD rodents but no alteration in liver mRNA levels by qRT-PCR. Data are consistent with apo-Cp being less stable than holo-Cp. Further research is needed to explain normal plasma iron in CuD mice. Reduction in Cp is a sensitive biomarker for copper deficiency. PMID:20170749

Quinone-Catalyzed Selective Oxidation of Organic Molecules

PubMed Central

Wendlandt, Alison E.

2016-01-01

Lead In Quinones are common stoichiometric reagents in organic chemistry. High potential para-quinones, such as DDQ and chloranil, are widely used and typically promote hydride abstraction. In recent years, many catalytic applications of these methods have been achieved by using transition metals, electrochemistry or O2 to regenerate the oxidized quinone in situ. Complementary studies have led to the development of a different class of quinones that resemble the ortho-quinone cofactors in Copper Amine Oxidases and mediate efficient and selective aerobic and/or electrochemical dehydrogenation of amines. The latter reactions typically proceed via electrophilic transamination and/or addition-elimination reaction mechanisms, rather than hydride abstraction pathways. The collective observations show that the quinone structure has a significant influence on the reaction mechanism and have important implications for the development of new quinone reagents and quinone-catalyzed transformations. PMID:26530485

Crystallization and preliminary X-ray analysis of copper amine oxidase from Escherichia coli K-12.

PubMed

Roh, J H; Suzuki, H; Kumagai, H; Yamashita, M; Azakami, H; Murooka, Y; Mikami, B

1994-05-13

Copper-containing monoamine oxidase (MAO) from Escherichia coli was overproduced in the periplasmic space by expression of the cloned gene. The purified MAO has been crystallized by means of the hanging drop technique using sodium citrate as a precipitant. The crystals belong to the orthorhombic system, space group P2(1)2(1)2(1), with unit cell dimensions of a = 136.1 A, b = 168.4 A and c = 81.6 A. The asymmetric unit contains one molecule of MAO, with a crystal volume per protein mass (Vm) of 2.88 A3/Da and a solvent content of 58% by volume. The crystals diffract X-rays to a resolution limit of at least 2.7 A and are resistant to X-ray radiation damage. They appear to be suitable for X-ray structure analysis.

Identification, expression and tissue distribution of a renalase homologue from mouse.

PubMed

Wang, Jian; Qi, Shaoling; Cheng, Wei; Li, Li; Wang, Fu; Li, Ying-Zi; Zhang, Shu-Ping

2008-12-01

FAD (flavin adenine dinucleotide)-dependent monoamine oxidases play very important roles in many biological processes. A novel monoamine oxidase, named renalase, has been identified in human kidney recently and is found to be markedly reduced in patients with end-stage renal disease (ESRD). Here, we reported the identification of a renalase homologue from mouse, termed mMAO-C (mouse monoamine oxidase-C) after the monoamine oxidase-A and -B (MAO-A and -B). This gene locates on the mouse chromosome 19C1 and its coding region spans 7 exons. The deuced amino acid sequences were predicted to contain a typical secretive signal peptide and a conserved amine oxidase domain. Phylogenetic analysis and multiple sequences alignment indicated that mMAO-C-like sequences exist in all examined species and share significant similarities. This gene has been submitted to the NCBI GenBank database (Accession number: DQ788834). With expression vectors generated from the cloned mMAO-C gene, exogenous protein was effectively expressed in both prokaryotic and eukaryotic cells. Recombinant mMAO-C protein was secreted out of human cell lines, indicating the biological function of its signal peptide. Moreover, tissue expression pattern analysis revealed that mMAO-C gene is predominantly expressed in the mouse kidney and testicle, which implies that kidney and testicle are the main sources of renalase secretion. Shortly, this study provides an insight into understanding the physiological and biological functions of mMAO-C and its homologues in endocrine.

The role of brassinosteroids in the regulation of the plasma membrane H+-ATPase and NADPH oxidase under cadmium stress.

PubMed

Jakubowska, Dagmara; Janicka, Małgorzata

2017-11-01

The present research aim was to define the role of brassinosteroids (BRs) in plant adaptation to cadmium stress. We observed a stimulating effect of exogenous BR on the activity of two plasma membrane enzymes which play a key role in plants adaptation to cadmium stress, H + -ATPase (EC 3.6.3.14) and NADPH oxidase (EC 1.6.3.1). Using anti-phosphothreonine antibody we showed that modification of PM H + -ATPase activity under BR action could result from phosphorylation of the enzyme protein. Also the relative expression of genes encoding both PM H + -ATPase and NADPH oxidase was affected by BR. To confirm the role of BR in the cadmium stimulating effect on activity of both studied plasma membrane enzymes, an assay in the presence of a BR biosynthesis inhibitor (propiconazole) was performed. Moreover, as a tool in our work we used commercially available plant mutants unable to BR biosynthesis or with dysfunctional BR signaling pathway, to further confirm participation of BR in plant adaptation to heavy metal stress. Presented results demonstrate some elements of the brassinosteroid-induced pathway activated under cadmium stress, wherein H + -ATPase and NADPH oxidase are key factors. Copyright © 2017 Elsevier B.V. All rights reserved.

Cross-talk between Aβ and endothelial SSAO/VAP-1 accelerates vascular damage and Aβ aggregation related to CAA-AD.

PubMed

Solé, Montse; Miñano-Molina, Alfredo J; Unzeta, Mercedes

2015-02-01

An association between semicarbazide-sensitive amine oxidase (SSAO) and cerebral amyloid angiopathy (CAA) related to Alzheimer's disease (AD) has been largely postulated. Increased SSAO activity and expression have been detected in cerebrovascular tissue and plasma of AD patients, colocalizing with cerebrovascular amyloid-beta (Aβ) deposits. As an enzyme, SSAO metabolizes primary amines generating hydrogen peroxide, ammonia, and aldehydes. The ability of these products to generate oxidative stress, to enhance the advanced glycation end-product generation, to promote the Aβ aggregation in vitro, and to induce apoptosis supports its role in CAA-related vascular pathology. However, whether the SSAO increase constitutes a cause or it is a consequence of the pathologic process has not been elucidated so far. To set up the nature of this relationship, vascular cell models expressing SSAO were treated with different Aβ forms, simulating the CAA conditions in vitro. It was found that the presence of the vasculotropic Dutch-mutated Aβ1-40 increases (Aβ1-40 D) the SSAO-dependent toxicity, which is accompanied by an increase of SSAO protein availability in endothelial cell membranes. In addition, SSAO enhances Aβ1-40 D and Aβ1-42 deposition on vascular cells by both activity-dependent and -independent mechanisms. Thus, we provide evidences indicating that Aβ itself could be one of the factors inducing SSAO increase in AD, enhancing its toxic effect, and inducing the vascular dysfunction and, in turn, that SSAO stimulates Aβ deposition on the vascular walls, thereby contributing to the CAA-AD progression. Therefore, molecules inhibiting SSAO could provide an alternative treatment for preventing/delaying the progress of CAA-AD-associated vasculopathy. Copyright © 2015 Elsevier Inc. All rights reserved.

Urate oxidase knockdown decreases oxidative stress in a murine hepatic cell line

USDA-ARS?s Scientific Manuscript database

Humans, birds, and some primates do not express the uric acid degrading enzyme urate oxidase (UOX) and, as a result, have plasma uric acid concentrations higher than UOX expressing animals. Although high uric acid concentrations are suggested to increase the antioxidant defense system and provide a...

Surface modification of polyvinyl alcohol/malonic acid nanofibers by gaseous dielectric barrier discharge plasma for glucose oxidase immobilization

NASA Astrophysics Data System (ADS)

Afshari, Esmail; Mazinani, Saeedeh; Ranaei-Siadat, Seyed-Omid; Ghomi, Hamid

2016-11-01

Polymeric nanofiber prepares a suitable situation for enzyme immobilization for variety of applications. In this research, we have fabricated polyvinyl alcohol (PVA)/malonic acid nanofibers using electrospinning. After fabrication of nanofibers, the effect of air, nitrogen, CO2, and argon DBD (dielectric barrier discharge) plasmas on PVA/malonic acid nanofibers were analysed. Among them, air plasma had the most significant effect on glucose oxidase (GOx) immobilization. Attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectrum analysis and X-ray photoelectron spectroscopy (XPS) results revealed that in case of air plasma modified nanofibers, the carboxyl groups on the surface are increased. The scanning electron microscopy (SEM) images showed that, after GOx immobilization, the modified nanofibers with plasma has retained its nanofiber structure. Finally, we analysed reusability and storage stability of GOx immobilized on plasma modified and unmodified nanofibers. The results were more satisfactory for modified nanofibers with respect to unmodified ones.

Direct and Efficient Dehydrogenation of Tetrahydroquinolines and Primary Amines Using Corona Discharge Generated on Ambient Hydrophobic Paper Substrate.

PubMed

Davis, Kathryn M; Badu-Tawiah, Abraham K

2017-04-01

The exposure of an aqueous-based liquid drop containing amines and graphite particles to plasma generated by a corona discharge results in heterogeneous aerobic dehydrogenation reactions. This green oxidation reaction occurring in ambient air afforded the corresponding quinolines and nitriles from tetrahydroquinolines and primary amines, respectively, at >96% yields in less than 2 min of reaction time. The accelerated dehydrogenation reactions occurred on the surface of a low energy hydrophobic paper, which served both as container for holding the reacting liquid drop and as a medium for achieving paper spray ionization of reaction products for subsequent characterization by ambient mass spectrometry. Control experiments indicate superoxide anions (O 2 •- ) are the main reactive species; the presence of graphite particles introduced heterogeneous surface effects, and enabled the efficient sampling of the plasma into the grounded analyte droplet solution. Graphical Abstract ᅟ.

Direct and Efficient Dehydrogenation of Tetrahydroquinolines and Primary Amines Using Corona Discharge Generated on Ambient Hydrophobic Paper Substrate

NASA Astrophysics Data System (ADS)

Davis, Kathryn M.; Badu-Tawiah, Abraham K.

2017-04-01

The exposure of an aqueous-based liquid drop containing amines and graphite particles to plasma generated by a corona discharge results in heterogeneous aerobic dehydrogenation reactions. This green oxidation reaction occurring in ambient air afforded the corresponding quinolines and nitriles from tetrahydroquinolines and primary amines, respectively, at >96% yields in less than 2 min of reaction time. The accelerated dehydrogenation reactions occurred on the surface of a low energy hydrophobic paper, which served both as container for holding the reacting liquid drop and as a medium for achieving paper spray ionization of reaction products for subsequent characterization by ambient mass spectrometry. Control experiments indicate superoxide anions (O2 •-) are the main reactive species; the presence of graphite particles introduced heterogeneous surface effects, and enabled the efficient sampling of the plasma into the grounded analyte droplet solution.

A rapid and sensitive assay method for measuring amine oxidase based on hydrogen peroxide-titanium complex formation.

PubMed

Nag; Saha; Choudhuri

2000-08-22

Hydrogenperoxide (H(2)O(2)) is an end product of diamine and polyamine oxidation by their respective oxidase enzymes. A new sensitive assay method is based on a H(2)O(2)-titanium (Ti) complex formation as an indicator of H(2)O(2) production due to polyamine oxidation. The orange-yellow coloured H(2)O(2)-Ti complex was measured at 410 nm in a Shimadzu spectrophotometer. The assay conditions for maximum diamine oxidase (DAO) and polyamine oxidase (PAO) as standardized here using the hypocotyl tissues of Vigna catjang Endl. cv Pusa Barsati consisted of pH 7.4 (40 mM potassium phosphate buffer), 3 mM substrate (putrescine or spermine), 37 degrees C incubation temperature and 30 min incubation time in the presence of catechol (10(-2) M) used as an inhibitor of both peroxidase and catalase activity. The method described here was significantly more sensitive than the starch-iodide method [T.A. Smith, Biochem. Biophys. Res. Commun. 41 (1970) 1452-1456], which could be improved further if measured under the same assay conditions as described for the H(2)O(2)-Ti method. Sensitivity of the present method was tested by assaying DAO/PAO activity in auxin treated hypocotyls of Vigna and comparing it with the starch-iodide method in two other plant samples.

Levels and interactions of plasma xanthine oxidase, catalase and liver function parameters in Nigerian children with Plasmodium falciparum infection.

PubMed

Iwalokun, B A; Bamiro, S B; Ogunledun, A

2006-12-01

Elevated plasma levels of xanthine oxidase and liver function parameters have been associated with inflammatory events in several human diseases. While xanthine oxidase provides in vitro protection against malaria, its pathophysiological functions in vivo and interactions with liver function parameters remain unclear. This study examined the interactions and plasma levels of xanthine oxidase (XO) and uric acid (UA), catalase (CAT) and liver function parameters GOT, GPT and bilirubin in asymptomatic (n=20), uncomplicated (n=32), and severe (n=18) falciparum malaria children aged 3-13 years. Compared to age-matched control (n=16), significant (p<0.05) elevation in xanthine oxidase by 100-550%, uric acid by 15.4-153.8%, GOT and GPT by 22.1-102.2%, and total bilirubin by 2.3-86% according to parasitaemia (geometric mean parasite density (GMPD)=850-87100 parasites/microL) was observed in the malarial children. Further comparison with control revealed higher CAT level (16.2+/-0.5 vs 14.6+/-0.4 U/L; p<0.05) lacking significant (p>0.05) correlation with XO, but lower CAT level (13.4-5.4 U/L) with improved correlations (r=-0.53 to -0.91; p<0.05) with XO among the asymptomatic and symptomatic malaria children studied. 75% of control, 45% of asymptomatic, 21.9% of uncomplicated, and none of severe malaria children had Hb level>11.0 g/dL. Multivariate analyses further revealed significant (p<0.05) correlations between liver function parameters and xanthine oxidase (r=0.57-0.64) only in the severe malaria group. We conclude that elevated levels of XO and liver enzymes are biochemical features of Plasmodium falciparum parasitaemia in Nigerian children, with both parameters interacting differently to modulate the catalase response in asymptomatic and symptomatic falciparum malaria.

Improving Glyphosate Oxidation Activity of Glycine Oxidase from Bacillus cereus by Directed Evolution

PubMed Central

Zhan, Tao; Zhang, Kai; Chen, Yangyan; Lin, Yongjun; Wu, Gaobing; Zhang, Lili; Yao, Pei; Shao, Zongze; Liu, Ziduo

2013-01-01

Glyphosate, a broad spectrum herbicide widely used in agriculture all over the world, inhibits 5-enolpyruvylshikimate-3-phosphate synthase in the shikimate pathway, and glycine oxidase (GO) has been reported to be able to catalyze the oxidative deamination of various amines and cleave the C-N bond in glyphosate. Here, in an effort to improve the catalytic activity of the glycine oxidase that was cloned from a glyphosate-degrading marine strain of Bacillus cereus (BceGO), we used a bacteriophage T7 lysis-based method for high-throughput screening of oxidase activity and engineered the gene encoding BceGO by directed evolution. Six mutants exhibiting enhanced activity toward glyphosate were screened from two rounds of error-prone PCR combined with site directed mutagenesis, and the beneficial mutations of the six evolved variants were recombined by DNA shuffling. Four recombinants were generated and, when compared with the wild-type BceGO, the most active mutant B3S1 showed the highest activity, exhibiting a 160-fold increase in substrate affinity, a 326-fold enhancement in catalytic efficiency against glyphosate, with little difference between their pH and temperature stabilities. The role of these mutations was explored through structure modeling and molecular docking, revealing that the Arg51 mutation is near the active site and could be an important residue contributing to the stabilization of glyphosate binding, while the role of the remaining mutations is unclear. These results provide insight into the application of directed evolution in optimizing glycine oxidase function and have laid a foundation for the development of glyphosate-tolerant crops. PMID:24223901

Fatigue and stress studies : an improved semi-automated procedure for flurometric determination of plasma catecholamines.

DOT National Transportation Integrated Search

1966-04-01

A semiautomated technique is described for the estimation of total catecholamines in plasma by the trihydroxyindole procedure. The method utilizes conventional alumina-column chromatography for isolation of the amines. Catecholamine oxidation, tautom...

BEHAVIORAL OUTCOMES OF MONOAMINE OXIDASE DEFICIENCY: PRECLINICAL AND CLINICAL EVIDENCE

PubMed Central

Bortolato, Marco; Shih, Jean C.

2012-01-01

Monoamine oxidase (MAO) isoenzymes A and B are mitochondrial-bound proteins, catalyzing the oxidative deamination of monoamine neurotransmitters as well as xenobiotic amines. Although they derive from a common ancestral progenitor gene, are located at X-chromosome and display 70% structural identity, their substrate preference, regional distribution, and physiological role are divergent. In fact, while MAO-A has high affinity for serotonin and norepinephrine, MAO-B primarily serves the catabolism of 2-phenylethylamine (PEA) and contributes to the degradation of other trace amines and dopamine. Convergent lines of preclinical and clinical evidence indicate that variations in MAO enzymatic activity—due to either genetic or environmental factors—can exert a profound influence on behavioral regulation and play a role in the pathophysiology of a large spectrum of mental and neurodegenerative disorders, ranging from antisocial personality disorder to Parkinson’s disease. Over the past few years, numerous advances have been made in our understanding of the phenotypical variations associated with genetic polymorphisms and mutations of the genes encoding for both isoenzymes. In particular, novel findings on the phenotypes of MAO-deficient mice are highlighting novel potential implications of both isoenzymes in a broad spectrum of mental disorders, ranging from autism and anxiety to impulse-control disorders and ADHD. These studies will lay the foundation for future research on the neurobiological and neurochemical bases of these pathological conditions, as well as the role of gene × environment interactions in the vulnerability to several mental disorders. PMID:21971001

Behavioral outcomes of monoamine oxidase deficiency: preclinical and clinical evidence.

PubMed

Bortolato, Marco; Shih, Jean C

2011-01-01

Monoamine oxidase (MAO) isoenzymes A and B are mitochondrial-bound proteins, catalyzing the oxidative deamination of monoamine neurotransmitters as well as xenobiotic amines. Although they derive from a common ancestral progenitor gene, are located at X-chromosome and display 70% structural identity, their substrate preference, regional distribution, and physiological role are divergent. In fact, while MAO-A has high affinity for serotonin and norepinephrine, MAO-B primarily serves the catabolism of 2-phenylethylamine (PEA) and contributes to the degradation of other trace amines and dopamine. Convergent lines of preclinical and clinical evidence indicate that variations in MAO enzymatic activity--due to either genetic or environmental factors--can exert a profound influence on behavioral regulation and play a role in the pathophysiology of a large spectrum of mental and neurodegenerative disorders, ranging from antisocial personality disorder to Parkinson's disease. Over the past few years, numerous advances have been made in our understanding of the phenotypical variations associated with genetic polymorphisms and mutations of the genes encoding for both isoenzymes. In particular, novel findings on the phenotypes of MAO-deficient mice are highlighting novel potential implications of both isoenzymes in a broad spectrum of mental disorders, ranging from autism and anxiety to impulse-control disorders and ADHD. These studies will lay the foundation for future research on the neurobiological and neurochemical bases of these pathological conditions, as well as the role of gene × environment interactions in the vulnerability to several mental disorders. Copyright © 2011 Elsevier Inc. All rights reserved.

Diamond surface functionalization with biomimicry - Amine surface tether and thiol moiety for electrochemical sensors

NASA Astrophysics Data System (ADS)

Sund, James B.; Causey, Corey P.; Wolter, Scott D.; Parker, Charles B.; Stoner, Brian R.; Toone, Eric J.; Glass, Jeffrey T.

2014-05-01

The surface of conducting diamond was functionalized with a terminal thiol group that is capable of binding and detecting nitrogen-oxygen species. The functionalization process employed multiple steps starting with doped diamond films grown by plasma enhanced chemical vapor deposition followed by hydrogen termination and photochemical attachment of a chemically protected amine alkene. The surface tether was deprotected to reveal the amine functionality, which enabled the tether to be extended with surface chemistry to add a terminal thiol moiety for electrochemical sensing applications. Each step of the process was validated using X-ray photoelectron spectroscopy analysis.

The robust bio-immobilization based on pulsed plasma polymerization of cyclopropylamine and glutaraldehyde coupling chemistry

NASA Astrophysics Data System (ADS)

Manakhov, Anton; Makhneva, Ekaterina; Skládal, Petr; Nečas, David; Čechal, Jan; Kalina, Lukáš; Eliáš, Marek; Zajíčková, Lenka

2016-01-01

The performance of immunosensing devices crucially depends on the methodology of antibody or antigen immobilization on the sensor surface. Hence, the stable intermediate layers capable of specific and reproducible binding of antibodies are required. Herein, we introduce the amine rich (NHx concentration of 6 at.%) layers prepared by pulsed plasma polymerization of cyclopropylamine (CPA) for functionalization of the quartz crystal microbalance (QCM) surface by the antibody specific to human serum albumin. In these layers the amine groups serve as anchor for the antibody binding. The sensitivity of QCM sensors prepared in this way surpasses the one for the previously reported sensors functionalized by the thiol-based self-assembled monolayers by the factor of 2. Our results thus show that CPA plasma polymers have a significant potential for further development of the active layers for biosensing applications.

Reverse effects of DPI administration combined with glutamine supplementation on function of rat neutrophils induced by overtraining.

PubMed

Dong, Jingmei; Chen, Peijie; Liu, Qing; Wang, Ru; Xiao, Weihua; Zhang, Yajun

2013-04-01

To examine the excessive reactive oxygen species (ROS) mediated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and the combined effect of glutamine supplementation and diphenyleneiodonium (DPI) on the function of neutrophils induced by overtraining. Fifty male Wistar rats were randomly divided into 5 groups: control group (C), overtraining group (E), DPI-administration group (D), glutamine-supplementation group (G), and combined DPI and glutamine group (DG). Blood was sampled from the orbital vein after rats were trained on treadmill for 11 wk. Cytokine and lipid peroxidation in blood plasma were measured by enzyme-linked immunosorbent assay. The colocalization between gp91phox and p47phox of the NADPH oxidase was detected using immunocytochemistry and confocal microscopy. The activity of NADPH oxidase was assessed by chemiluminescence. Neutrophils' respiratory burst and phagocytosis function were measured by flow cytometry. NADPH oxidase was activated by overtraining. Cytokine and lipid peroxidation in blood plasma and the activity of NADPH oxidase were markedly increased in Group E compared with group C. Neutrophil function was lower in group E than group C. Both lower neutrophils function and higher ROS production were reversed in Group DG. The glutamine and DPI interference alone in group D and group G was less effective than DPI and glutamine combined in group DG. Activation of NADPH oxidase is responsible for the production of superoxide anions, which leads to excessive ROS and is related to the decrease in neutrophil function induced by overtraining. The combined DPI administration and glutamine supplementation reversed the decreased neutrophil function after overtraining.

Controlling of N-alkylpolyamine analogue metabolism by selective deuteration.

PubMed

Ucal, Sebahat; Häkkinen, Merja R; Alanne, Aino-Liisa; Alhonen, Leena; Vepsäläinen, Jouko; Keinänen, Tuomo A; Hyvönen, Mervi T

2018-02-14

Replacing protium with deuterium is an efficient method to modulate drug metabolism. N -alkylated polyamine analogues are polyamine antimetabolites with proven anticancer efficacy. We have characterized earlier the preferred metabolic routes of N 1 , N 12 -diethylspermine (DESpm), N 1 -benzyl- N 12 -ethylspermine (BnEtSpm) and N 1 , N 12 -dibenzylspermine (DBSpm) by human recombinant spermine oxidase (SMOX) and acetylpolyamine oxidase (APAO). Here, we studied the above analogues, their variably deuterated counterparts and their metabolites as substrates and inhibitors of APAO, SMOX, semicarbazide-sensitive amine oxidase (SSAO), diamine oxidase (DAO) and monoamine oxidases. We found that targeted deuteration efficiently redirected the preferable cleavage site and suppressed reaction rate by APAO and SMOX in vitro We found a three- to six-fold decline in V max with moderate variable effect on K m when deuterium was located at the preferred hydrogen abstraction site of the analogue. We also found some of the metabolites to be potent inhibitors of DAO and SSAO. Surprisingly, analogue deuteration did not markedly alter the anti-proliferative efficacy of the drugs in DU145 prostate cancer cells, while in mouse embryonic fibroblasts, which had higher basal APAO and SMOX activities, moderate effect was observed. Interestingly, the anti-proliferative efficacy of the analogues did not correlate with their ability to suppress polyamine biosynthetic enzymes, induce spermidine/spermine- N 1 -acetyltransferase or deplete intracellular polyamine levels, but correlated with their ability to induce SMOX. Our data show that selective deuteration of N -alkyl polyamine analogues enables metabolic switching, offering the means for selective generation of bioactive metabolites inhibiting, e.g. SSAO and DAO, thus setting a novel basis for in vivo studies of this class of analogues. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Oligoamine analogues in combination with 2-difluoromethylornithine synergistically induce re-expression of aberrantly silenced tumour-suppressor genes

PubMed Central

Wu, Yu; Steinbergs, Nora; Murray-Stewart, Tracy; Marton, Laurence J.; Casero, Robert A.

2011-01-01

Epigenetic gene silencing is an important mechanism in the initiation and progression of cancer. Abnormal DNA CpG island hypermethylation and histone modifications are involved in aberrant silencing of tumour-suppressor genes. LSD1 (lysine-specific demethylase 1) was the first enzyme identified to specifically demethylate H3K4 (Lys4 of histone H3). Methylated H3K4 is an important mark associated with transcriptional activation. The flavin adenine dinucleotide-binding amine oxidase domain of LSD1 is homologous with two polyamine oxidases, SMO (spermine oxidase) and APAO (N1-acetylpolyamine oxidase). We have demonstrated previously that long-chain polyamine analogues, the oligoamines, are inhibitors of LSD1. In the present paper we report the synergistic effects of specific oligoamines in combination with DFMO (2-difluoromethylornithine), an inhibitor of ornithine decarboxylase, in human colorectal cancer cells. DFMO treatment depletes natural polyamines and increases the uptake of exogenous polyamines. The combination of oligoamines and DFMO results in a synergistic re-expression of aberrantly silenced tumour-suppressor genes, including SFRP2 (secreted frizzled-related protein 2), which encodes a Wnt signalling pathway antagonist and plays an anti-tumorigenic role in colorectal cancer. The treatment-induced re-expression of SFRP2 is associated with increased H3K4me2 (di-methyl H3K4) in the gene promoter. The combination of LSD1-inhibiting oligoamines and DFMO represents a novel approach to epigenetic therapy of cancer. PMID:22132744

Mechanistic Studies of para-Substituted N,N'-Dibenzyl-1,4-diaminobutanes as Substrates for a Mammalian Polyamine Oxidase

PubMed Central

Pozzi, Michelle Henderson; Gawandi, Vijay; Fitzpatrick, Paul F.

2009-01-01

The kinetics of oxidation of a series of para-substituted N, N'-dibenzyl-1,4-diaminobutanes by the flavoprotein polyamine oxidase from mouse have been determined to gain insight into the mechanism of amine oxidation by this member of the monoamine oxidase structural family. The kcat/Km values are maximal at pH 9, consistent with the singly charged substrate being the active form. The rate constant for flavin reduction, kred, by N,N'-dibenzyl-1,4-diaminobutane decreases about 5-fold below a pKa of ~8; this is attributed to the need for a neutral nitrogen at the site of oxidation. The kred and kcat values are comparable for each of the N, N'-dibenzyl-1,4-diaminobutanes, consistent with rate-limiting reduction. The deuterium kinetic isotope effects on kred and kcat are identical for each of the N, N'-dibenzyl-1,4-diaminobutanes, consistent with rate-limiting cleavage of the substrate CH bond. The kred values for seven different para-substituted N, N'-dibenzyl-1,4-diaminobutanes correlate with a combination of the van der Waals volume and σ value of the substrates, with ρ values of −0.59 at pH 8.6 and −0.09 at pH 6.6. These results are consistent with direct transfer of a hydride from the neutral CN bond of the substrate to the flavin as the mechanism of polyamine oxidase. PMID:19911805

Determination of bromhexine in plasma by reversed-phase liquid chromatography. Interference of lipoproteins on extraction.

PubMed

Johansson, M; Lenngren, S

1988-11-18

Extraction of the hydrophobic tertiary amine bromhexine from plasma using cyclohexane-heptafluorobutanol (99.5:0.5, v/v) was studied at different pH values. The extraction yield from buffer solutions was quantitative at pH greater than 4.1, but from plasma the extraction yield decreased with increasing pH. Furthermore, at pH 8.4 the extraction yield varied greatly (56-99%) in different human plasma. The addition of lipoproteins to phosphate buffer, at pH 8.1, decreased the extraction yields considerably. Quantitative extraction from plasma was obtained by using a very long extraction time at pH 8.4 or by decreasing the pH to 5.4. The chromatographic system consisted of a reversed-phase column (Nucleosil C18, 5 microns) with an acidic mobile phase (methanol-phosphate buffer, pH 2) containing an aliphatic tertiary amine. UV detection at 308 or 254 nm was used. The limit of quantitation was 5 ng/ml using 3.00 ml of plasma and detection at 254 nm. The intra-assay precision for bromhexine was better than 3.6% at 5 ng/ml.

Cultured rat vascular smooth muscle cells are resistant to methylamine toxicity: no correlation to semicarbazide-sensitive amine oxidase

NASA Technical Reports Server (NTRS)

Langford, S. D.; Trent, M. B.; Boor, P. J.

2001-01-01

Methylamine (MA), a component of serum and a metabolite of nicotine and certain insecticides and herbicides, is metabolized by semicarbazide-sensitive amine oxidase (SSAO). MA is toxic to cultured human umbilical vein and calf pulmonary artery endothelial cells. Endothelial cells, which do not exhibit endogenous SSAO activity, are exposed to SSAO circulating in serum. In contrast, vascular smooth muscle cells (VSMC) do exhibit innate SSAO activity both in vivo and in vitro. This property, together with the critical localization of VSMC within the arterial wall, led us to investigate the potential toxicity of MA to VSMC. Cultured rat VSMC were treated with MA (10-5 to 1 M). In some cultures, SSAO was selectively inhibited with semicarbazide or MDL-72145 [(E)-2-(3,4-dimethoxyphenyl)-3-fluoroallylamine]. Cytotoxicity was measured via MTT, vital dye exclusion, and clonogenic assays. MA proved to be toxic to VSMC only at relatively high concentrations (LC(50) of 0.1 M). The inhibition of SSAO with semicarbazide or MDL-72145 did not increase MA toxicity, suggesting that the production of formaldehyde via tissue-bound, SSAO-mediated MA metabolism does not play a role in the minimal toxicity observed in isolated rat VSMC. The omission of fetal calf serum (FCS), which contains high SSAO activity, from media similarly showed little effect on cytotoxicity. We conclude that VSMC--in contrast to previous results in endothelial cells--are relatively resistant to MA toxicity, and SSAO does not play a role in VSMC injury by MA.

Molecular cloning, expression, and functional analysis of the copper amine oxidase gene in the endophytic fungus Shiraia sp. Slf14 from Huperzia serrata.

PubMed

Yang, Huilin; Peng, Silu; Zhang, Zhibin; Yan, Riming; Wang, Ya; Zhan, Jixun; Zhu, Du

2016-12-01

Huperzine A (HupA) is a drug used for the treatment of Alzheimer's disease. However, the biosynthesis of this medicinally important compound is not well understood. The HupA biosynthetic pathway is thought to be initiated by the decarboxylation of lysine to form cadaverine, which is then converted to 5-aminopentanal by copper amine oxidase (CAO). In this study, we cloned and expressed an SsCAO gene from a HupA-producing endophytic fungus, Shiraia sp. Slf14. Analysis of the deduced protein amino acid sequence showed that it contained the Asp catalytic base, conserved motif Asn-Tyr-Asp/Glu, and three copper-binding histidines. The cDNA of SsCAO was amplified and expressed in Escherichia coli BL21(DE3), from which a 76 kDa protein was obtained. The activity of this enzyme was tested, which provided more information about the SsCAO gene in the endophytic fungus. Gas Chromatograph-Mass Spectrometry (GC-MS) revealed that this SsCAO could accept cadaverine as a substrate to produce 5-aminopentanal, the precursor of HupA. Phylogenetic tree analysis indicated that the SsCAO from Shiraia sp. Slf14 was closely related to Stemphylium lycopersici CAO. This is the first report on the cloning and expression of a CAO gene from HupA-producing endophytic fungi. Functional characterization of this enzyme provides new insights into the biosynthesis of the HupA an anti-Alzheimer's drug. Copyright © 2016 Elsevier Inc. All rights reserved.

Behavioral disinhibition and reduced anxiety-like behaviors in monoamine oxidase B-deficient mice.

PubMed

Bortolato, Marco; Godar, Sean C; Davarian, Shieva; Chen, Kevin; Shih, Jean C

2009-12-01

Monoamine oxidase (MAO) B catalyzes the degradation of beta-phenylethylamine (PEA), a trace amine neurotransmitter implicated in mood regulation. Although several studies have shown an association between low MAO B activity in platelets and behavioral disinhibition in humans, the nature of this relation remains undefined. To investigate the impact of MAO B deficiency on the emotional responses elicited by environmental cues, we tested MAO B knockout (KO) mice in a set of behavioral assays capturing different aspects of anxiety-related manifestations, such as the elevated plus maze, defensive withdrawal, marble burying, and hole board. Furthermore, MAO B KO mice were evaluated for their exploratory patterns in response to unfamiliar objects and risk-taking behaviors. In comparison with their wild-type (WT) littermates, MAO B KO mice exhibited significantly lower anxiety-like responses and shorter latency to engage in risk-taking behaviors and exploration of unfamiliar objects. To determine the neurobiological bases of the behavioral differences between WT and MAO B KO mice, we measured the brain-regional levels of PEA in both genotypes. Although PEA levels were significantly higher in all brain regions of MAO B KO in comparison with WT mice, the most remarkable increments were observed in the striatum and prefrontal cortex, two key regions for the regulation of behavioral disinhibition. However, no significant differences in transcript levels of PEA's selective receptor, trace amine-associated receptor 1 (TAAR1), were detected in either region. Taken together, these results suggest that MAO B deficiency may lead to behavioral disinhibition and decreased anxiety-like responses partially through regional increases of PEA levels.

Monoamine oxidases: the biochemistry of the proteins as targets in medicinal chemistry and drug discovery.

PubMed

Ramsay, Rona R

2012-01-01

Monoamine oxidase (MAO, EC.1.4.3.4) has been a drug target for 60 years, with the primary rationale of developing drugs to treat neuropsychiatric disorders. The biological importance of MAO is to regulate amine levels is the brain and to metabolize amines and drugs in the periphery. This review of the biochemistry of MAO A and MAO B describes the functional properties of the two enzymes integrated with knowledge of the structures of the many MAOinhibitor complexes published in the last 10 years. The analysis of activity, and the chemical and kinetic mechanisms are discussed. Inhibition studies on human MAO in vitro are now facilitated by assays using readily available materials but the kinetics of MAO involving alternative oxidative pathways and sensitivity to the oxygen concentration mean that careful analysis of the data is required as well as the good practice of determining mechanism of inhibition and kinetic constants. Kinetic constants can then be compared with thermodynamic calculations. Inhibitors bind to both the oxidized and reduced forms of MAO present during turnover so both forms should be considered when using molecular dynamics to facilitate drug design. Interaction of inhibitors with the active site can be detected as changes in the visible spectrum and these changes can provide clues about the flavin adduct formed for irreversible inhibitors or about the proximity to the flavin for reversible inhibitors. Developing areas (knock-out mice for behavior, the imidazoline binding site, and imaging to monitor the activity and the inhibition of MAO in the patient) are mentioned.

High density gold nanoparticles immobilized on surface via plasma deposited APTES film for decomposing organic compounds in microchannels

NASA Astrophysics Data System (ADS)

Rao, Xi; Guyon, Cédric; Ognier, Stephanie; Da Silva, Bradley; Chu, Chenglin; Tatoulian, Michaël; Hassan, Ali Abou

2018-05-01

Immobilization of colloidal particles (e.g. gold nanoparticles (AuNps)) on the inner surface of micro-/nano- channels has received a great interest for catalysis. A novel catalytic ozonation setup using a gold-immobilized microchannel reactor was developed in this work. To anchor AuNps, (3-aminopropyl) triethoxysilane (APTES) with functional amine groups was deposited using plasma enhanced chemical vapor deposition (PECVD) process. The results clearly evidenced that PECVD processing exhibited relatively high efficiency for grafting amine groups and further immobilizing AuNPs. The catalytic activity of gold immobilized microchannel was evaluated by pyruvic acid ozonation. The decomposition rate calculated from High Performance Liquid Chromatography (HPLC) indicated a much better catalytic performance of gold in microchannel than that in batch. The results confirmed immobilizing gold nanoparticles on plasma deposited APTES for preparing catalytic microreactors is promising for the wastewater treatment in the future.

Priming of the neutrophil NADPH oxidase activation: role of p47phox phosphorylation and NOX2 mobilization to the plasma membrane.

PubMed

El-Benna, Jamel; Dang, Pham My-Chan; Gougerot-Pocidalo, Marie-Anne

2008-07-01

Neutrophils play an essential role in host defense against microbial pathogens and in the inflammatory reaction. Upon activation, neutrophils produce superoxide anion (O*2), which generates other reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), hydroxyl radical (OH*) and hypochlorous acid (HOCl), together with microbicidal peptides and proteases. The enzyme responsible for O2* production is called the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase or respiratory burst oxidase. This multicomponent enzyme system is composed of two trans-membrane proteins (p22phox and gp91phox/NOX2, which form the cytochrome b558), three cytosolic proteins (p47phox, p67phox, p40phox) and a GTPase (Rac1 or Rac2), which assemble at membrane sites upon cell activation. NADPH oxidase activation in phagocytes can be induced by a large number of soluble and particulate factors. Three major events accompany NAPDH oxidase activation: (1) protein phosphorylation, (2) GTPase activation, and (3) translocation of cytosolic components to the plasma membrane to form the active enzyme. Actually, the neutrophil NADPH oxidase exists in different states: resting, primed, activated, or inactivated. The resting state is found in circulating blood neutrophils. The primed state can be induced by neutrophil adhesion, pro-inflammatory cytokines, lipopolysaccharide, and other agents and has been characterized as a "ready to go" state, which results in a faster and higher response upon exposure to a second stimulus. The active state is found at the inflammatory or infection site. Activation is induced by the pathogen itself or by pathogen-derived formylated peptides and other agents. Finally, inactivation of NADPH oxidase is induced by anti-inflammatory agents to limit inflammation. Priming is a "double-edged sword" process as it contributes to a rapid and efficient elimination of the pathogens but can also induce the generation of large quantities of toxic ROS by hyperactivation of the NADPH oxidase, which can damage surrounding tissues and participate to inflammation. In order to avoid extensive damage to host tissues, NADPH oxidase priming and activation must be tightly regulated. In this review, we will discuss some of the mechanisms of NADPH oxidase priming in neutrophils and the relevance of this process to physiology and pathology.

A Novel Extracellular Multicopper Oxidase from Phanerochaete chrysosporium with Ferroxidase Activity

PubMed Central

Larrondo, Luis F.; Salas, Loreto; Melo, Francisco; Vicuña, Rafael; Cullen, Daniel

2003-01-01

Lignin degradation by the white rot basidiomycete Phanerochaete chrysosporium involves various extracellular oxidative enzymes, including lignin peroxidase, manganese peroxidase, and a peroxide-generating enzyme, glyoxal oxidase. Recent studies have suggested that laccases also may be produced by this fungus, but these conclusions have been controversial. We identified four sequences related to laccases and ferroxidases (Fet3) in a search of the publicly available P. chrysosporium database. One gene, designated mco1, has a typical eukaryotic secretion signal and is transcribed in defined media and in colonized wood. Structural analysis and multiple alignments identified residues common to laccase and Fet3 sequences. A recombinant MCO1 (rMCO1) protein expressed in Aspergillus nidulans had a molecular mass of 78 kDa, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the copper I-type center was confirmed by the UV-visible spectrum. rMCO1 oxidized various compounds, including 2,2′-azino(bis-3-ethylbenzthiazoline-6-sulfonate) (ABTS) and aromatic amines, although phenolic compounds were poor substrates. The best substrate was Fe2+, with a Km close to 2 μM. Collectively, these results suggest that the P. chrysosporium genome does not encode a typical laccase but rather encodes a unique extracellular multicopper oxidase with strong ferroxidase activity. PMID:14532088

Modulation of resistance artery tone by the trace amine β-phenylethylamine: dual indirect sympathomimetic and α1-adrenoceptor blocking actions.

PubMed

Narang, Deepak; Kerr, Paul M; Lunn, Stephanie E; Beaudry, Rhys; Sigurdson, Julie; Lalies, Margaret D; Hudson, Alan L; Light, Peter E; Holt, Andrew; Plane, Frances

2014-10-01

The trace amine β-phenylethylamine (PEA) is normally present in the body at low nanomolar concentrations but can reach micromolar levels after ingestion of drugs that inhibit monoamine oxidase and primary amine oxidase. In vivo, PEA elicits a robust pressor response, but there is no consensus regarding the underlying mechanism, with both vasodilation and constriction reported in isolated blood vessels. Using functional and biochemical approaches, we found that at low micromolar concentrations PEA (1-30 μM) enhanced nerve-evoked vasoconstriction in the perfused rat mesenteric bed but at a higher concentration (100 μM) significantly inhibited these responses. The α2-adrenoceptor antagonist rauwolscine (1 µM) also enhanced nerve-mediated vasoconstriction, but in the presence of both rauwolscine (1 µM) and PEA (30 µM) together, nerve-evoked responses were initially potentiated and then showed time-dependent rundown. PEA (10 and 100 μM) significantly increased noradrenaline outflow from the mesenteric bed as determined by high-pressure liquid chromatography coupled with electrochemical detection. In isolated endothelium-denuded arterial segments, PEA (1 µM to 1 mM) caused concentration-dependent reversal of tone elicited by the α1-adrenoceptor agonists noradrenaline (EC50 51.69 ± 10.8 μM; n = 5), methoxamine (EC50 68.21 ± 1.70 μM; n = 5), and phenylephrine (EC50 67.74 ± 16.72 μM; n = 5) but was ineffective against tone induced by prostaglandin F2 α or U46619 (9,11-dideoxy-9α,11α-methanoepoxyprostaglandin F2 α). In rat brain homogenates, PEA displaced binding of both [(3)H]prazosin (Ki ≈ 25 μM) and [(3)H]rauwolscine (Ki ≈ 1.2 μM), ligands for α1- and α2-adrenoceptors, respectively. These data provide the first demonstration that dual indirect sympathomimetic and α1-adrenoceptor blocking actions underlie the vascular effects of PEA in resistance arteries. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Erythrocyte NADPH oxidase activity modulated by Rac GTPases, PKC, and plasma cytokines contributes to oxidative stress in sickle cell disease

PubMed Central

Pushkaran, Suvarnamala; Konstantinidis, Diamantis G.; Koochaki, Sebastian; Malik, Punam; Mohandas, Narla; Zheng, Yi; Joiner, Clinton H.; Kalfa, Theodosia A.

2013-01-01

Chronic inflammation has emerged as an important pathogenic mechanism in sickle cell disease (SCD). One component of this inflammatory response is oxidant stress mediated by reactive oxygen species (ROS) generated by leukocytes, endothelial cells, plasma enzymes, and sickle red blood cells (RBC). Sickle RBC ROS generation has been attributed to sickle hemoglobin auto-oxidation and Fenton chemistry reactions catalyzed by denatured heme moieties bound to the RBC membrane. In this study, we demonstrate that a significant part of ROS production in sickle cells is mediated enzymatically by NADPH oxidase, which is regulated by protein kinase C, Rac GTPase, and intracellular Ca2+ signaling within the sickle RBC. Moreover, plasma from patients with SCD and isolated cytokines, such as transforming growth factor β1 and endothelin-1, enhance RBC NADPH oxidase activity and increase ROS generation. ROS-mediated damage to RBC membrane components is known to contribute to erythrocyte rigidity and fragility in SCD. Erythrocyte ROS generation, hemolysis, vaso-occlusion, and the inflammatory response to tissue damage may therefore act in a positive-feedback loop to drive the pathophysiology of sickle cell disease. These findings suggest a novel pathogenic mechanism in SCD and may offer new therapeutic targets to counteract inflammation and RBC rigidity and fragility in SCD. PMID:23349388

A decade of crystallization drops: crystallization of the cbb3 cytochrome c oxidase from Pseudomonas stutzeri.

PubMed

Buschmann, Sabine; Richers, Sebastian; Ermler, Ulrich; Michel, Hartmut

2014-04-01

The cbb3 cytochrome c oxidases are distant members of the superfamily of heme copper oxidases. These terminal oxidases couple O2 reduction with proton transport across the plasma membrane and, as a part of the respiratory chain, contribute to the generation of an electrochemical proton gradient. Compared with other structurally characterized members of the heme copper oxidases, the recently determined cbb3 oxidase structure at 3.2 Å resolution revealed significant differences in the electron supply system, the proton conducting pathways and the coupling of O2 reduction to proton translocation. In this paper, we present a detailed report on the key steps for structure determination. Improvement of the protein quality was achieved by optimization of the number of lipids attached to the protein as well as the separation of two cbb3 oxidase isoenzymes. The exchange of n-dodecyl-β-D-maltoside for a precisely defined mixture of two α-maltosides and decanoylsucrose as well as the choice of the crystallization method had a most profound impact on crystal quality. This report highlights problems frequently encountered in membrane protein crystallization and offers meaningful approaches to improve crystal quality. © 2014 The Protein Society.

A decade of crystallization drops: Crystallization of the cbb3 cytochrome c oxidase from Pseudomonas stutzeri

PubMed Central

Buschmann, Sabine; Richers, Sebastian; Ermler, Ulrich; Michel, Hartmut

2014-01-01

The cbb3 cytochrome c oxidases are distant members of the superfamily of heme copper oxidases. These terminal oxidases couple O2 reduction with proton transport across the plasma membrane and, as a part of the respiratory chain, contribute to the generation of an electrochemical proton gradient. Compared with other structurally characterized members of the heme copper oxidases, the recently determined cbb3 oxidase structure at 3.2 Å resolution revealed significant differences in the electron supply system, the proton conducting pathways and the coupling of O2 reduction to proton translocation. In this paper, we present a detailed report on the key steps for structure determination. Improvement of the protein quality was achieved by optimization of the number of lipids attached to the protein as well as the separation of two cbb3 oxidase isoenzymes. The exchange of n-dodecyl-β-d-maltoside for a precisely defined mixture of two α-maltosides and decanoylsucrose as well as the choice of the crystallization method had a most profound impact on crystal quality. This report highlights problems frequently encountered in membrane protein crystallization and offers meaningful approaches to improve crystal quality. PMID:24488923

Plasma Membrane ATPase Activity following Reversible and Irreversible Freezing Injury 1

PubMed Central

Iswari, S.; Palta, Jiwan P.

1989-01-01

Plasma membrane ATPase has been proposed as a site of functional alteration during early stages of freezing injury. To test this, plasma membrane was purified from Solanum leaflets by a single step partitioning of microsomes in a dextran-polyethylene glycol two phase system. Addition of lysolecithin in the ATPase assay produced up to 10-fold increase in ATPase activity. ATPase activity was specific for ATP with a Km around 0.4 millimolar. Presence of the ATPase enzyme was identified by immunoblotting with oat ATPase antibodies. Using the phase partitioning method, plasma membrane was isolated from Solanum commersonii leaflets which had four different degrees of freezing damage, namely, slight (reversible), partial (partially reversible), substantial and total (irreversible). With slight (reversible) damage the plasma membrane ATPase specific activity increased 1.5- to 2-fold and its Km was decreased by about 3-fold, whereas the specific activity of cytochrome c reductase and cytochrome c oxidase in the microsomes were not different from the control. However, with substantial (lethal, irreversible) damage, there was a loss of membrane protein, decrease in plasma membrane ATPase specific activity and decrease in Km, while cytochrome c oxidase and cytochrome c reductase were unaffected. These results support the hypothesis that plasma membrane ATPase is altered by slight freeze-thaw stress. Images Figure 1 Figure 2 PMID:16666856

Laser Scribed Graphene Biosensor for Detection of Biogenic Amines in Food Samples Using Locally Sourced Materials.

PubMed

Vanegas, Diana C; Patiño, Laksmi; Mendez, Connie; Oliveira, Daniela Alves de; Torres, Alba M; Gomes, Carmen L; McLamore, Eric S

2018-04-24

In foods, high levels of biogenic amines (BA) are the result of microbial metabolism that could be affected by temperatures and storage conditions. Thus, the level of BA is commonly used as an indicator of food safety and quality. This manuscript outlines the development of laser scribed graphene electrodes, with locally sourced materials, for reagent-free food safety biosensing. To fabricate the biosensors, the graphene surface was functionalized with copper microparticles and diamine oxidase, purchased from a local supermarket; and then compared to biosensors fabricated with analytical grade materials. The amperometric biosensor exhibits good electrochemical performance, with an average histamine sensitivity of 23.3 µA/mM, a lower detection limit of 11.6 µM, and a response time of 7.3 s, showing similar performance to biosensors constructed from analytical grade materials. We demonstrated the application of the biosensor by testing total BA concentration in fish paste samples subjected to fermentation with lactic acid bacteria. Biogenic amines concentrations prior to lactic acid fermentation were below the detection limit of the biosensor, while concentration after fermentation was 19.24 ± 8.21 mg histamine/kg, confirming that the sensor was selective in a complex food matrix. The low-cost, rapid, and accurate device is a promising tool for biogenic amine estimation in food samples, particularly in situations where standard laboratory techniques are unavailable, or are cost prohibitive. This biosensor can be used for screening food samples, potentially limiting food waste, while reducing chances of foodborne outbreaks.

Inhibition of monoamine oxidase by moclobemide: effects on monoamine metabolism and secretion of anterior pituitary hormones and cortisol in healthy volunteers.

PubMed Central

Koulu, M; Scheinin, M; Kaarttinen, A; Kallio, J; Pyykkö, K; Vuorinen, J; Zimmer, R H

1989-01-01

1. Single oral doses (100, 200 and 300 mg) of moclobemide, a reversible inhibitor of monoamine oxidase (MAO) with predominant effects on the A-type of the enzyme, were administered to eight young, healthy male volunteers in a double-blind, random-order, placebo-controlled study. The investigation was thereafter continued in an open fashion by administering a single 10 mg dose of the MAO-B inhibitor deprenyl to the same subjects. 2. Deamination of catecholamines was powerfully and dose-dependently inhibited by moclobemide, as evidenced by up to 40% decreases in the urinary excretion of deaminated catecholamine metabolites, corresponding increases in the excretion of non-deaminated, methylated metabolites, and up to 79% average decreases in the plasma concentration of 3,4-dihydroxyphenylglycol (DHPG), a deaminated metabolite of noradrenaline (NA), and up to 75% average decreases in the plasma concentrations of 3,4-dihydroxyphenylacetic acid (DOPAC), a deaminated metabolite of dopamine. The urinary excretion of 5-hydroxyindoleacetic acid (5-HIAA) was only slightly reduced. In contrast, deprenyl, in a dose which almost totally inhibited MAO-B activity in blood platelets, did not appreciably affect the plasma concentrations of DHPG or DOPAC. 3. Due to the rapid, reversible, dose-dependent and MAO-A specific effect of moclobemide on plasma concentrations of DHPG, it is suggested that DHPG in plasma may be a useful indicator of the magnitude and duration of MAO-A inhibition in man. 4. Sympatho-adrenal function at rest was not significantly altered by moclobemide, as judged by unchanged plasma catecholamine concentrations and stable blood pressure and heart rate recordings. 5. Monoamine oxidase type B activity in blood platelets was slightly (less than 30%) and transiently inhibited after moclobemide. 6. The secretion of prolactin was dose-dependently stimulated by moclobemide, whereas the plasma concentrations of growth hormone (hGH) and cortisol remained unchanged. PMID:2469451

Enhanced central serotonin release from slices of rat hypothalamus following repeated nialamide administration: evidence supporting the overactive serotonin receptor theory of depression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Offord, S.J.

1986-01-01

Researchers are suggesting unipolar affective disorders may be related to an abnormality in biogenic amine receptor-sensitivity. This abnormality may be a result of a dysfunction in central serotonin (5-HT) release mechanisms. 5-HT neurotransmission is modulated by presynaptic autoreceptors, which are members of the 5-HT/sub 1/ receptor subtype. The autoreceptor is thought to play an important role in the homeostasis of the central 5-HT synapse and could be a site at which some antidepressants mediate their therapeutic effect. The number of 5-HT/sub 1/ type receptor binding sites are reduced and behavior mediated by this receptor is abolished following repeated injections ofmore » monoamine oxidase inhibitor type antidepressants. These changes did not occur following a single injection. It was hypothesized that repeated treatment with a monoamine oxidase inhibitor would reduce the sensitivity of 5-HT autoreceptors and enhance 5-HT release. Rats were pretreated with single or repeated (twice daily for 7 days) intraperitoneal injections of nialamide (40 mg/kg) or chlorimipramine (10 mg/kg) and the ability of the autoreceptor agonist to inhibit potassium-induced /sup 3/H-5-HT release was evaluated using an in vitro superfusion system. These changes in 5-HT autoreceptor activity are consistent with other reports evaluating monoamine oxidase inhibitors on 5-HT/sub 1/ type receptors. It is hypothesized that the changes in 5-HT neurotransmission are related to the antidepressant mechanism of monoamine oxidase inhibitors.« less

Isolated sulfite oxidase deficiency.

PubMed

Rupar, C A; Gillett, J; Gordon, B A; Ramsay, D A; Johnson, J L; Garrett, R M; Rajagopalan, K V; Jung, J H; Bacheyie, G S; Sellers, A R

1996-12-01

Isolated sulfite oxidase (SO) deficiency is an autosomal recessively inherited inborn error of sulfur metabolism. In this report of a ninth patient the clinical history, laboratory results, neuropathological findings and a mutation in the sulfite oxidase gene are described. The data from this patient and previously published patients with isolated sulfite oxidase deficiency and molybdenum cofactor deficiency are summarized to characterize this rare disorder. The patient presented neonatally with intractable seizures and did not progress developmentally beyond the neonatal stage. Dislocated lenses were apparent at 2 months. There was increased urine excretion of sulfite and S-sulfocysteine and a decreased concentration of plasma cystine. A lactic acidemia was present for 6 months. Liver sulfite oxidase activity was not detectable but xanthine dehydrogenase activity was normal. The boy died of respiratory failure at 32 months. Neuropathological findings of cortical necrosis and extensive cavitating leukoencephalopathy were reminiscent of those seen in severe perinatal asphyxia suggesting an etiology of energy deficiency. A point mutation that resulted in a truncated protein missing the molybdenum-binding site has been identified.

Antioxidant treatment prevents the development of fructose-induced abdominal adipose tissue dysfunction.

PubMed

Fariña, Juan Pablo; García, María Elisa; Alzamendi, Ana; Giovambattista, Andrés; Marra, Carlos Alberto; Spinedi, Eduardo; Gagliardino, Juan José

2013-07-01

In the present study, we tested the effect of OS (oxidative stress) inhibition in rats fed on an FRD [fructose-rich diet; 10% (w/v) in drinking water] for 3 weeks. Normal adult male rats received a standard CD (commercial diet) or an FRD without or with an inhibitor of NADPH oxidase, APO (apocynin; 5 mM in drinking water; CD-APO and FRD-APO). We thereafter measured plasma OS and metabolic-endocrine markers, AAT (abdominal adipose tissue) mass and cell size, FA (fatty acid) composition (content and release), OS status, LEP (leptin) and IRS (insulin receptor substrate)-1/IRS-2 mRNAs, ROS (reactive oxygen species) production, NADPH oxidase activity and LEP release by isolated AAT adipocytes. FRD-fed rats had larger AAT mass without changes in body weight, and higher plasma levels of TAG (triacylglycerol), FAs, TBARS (thiobarbituric acid-reactive substance) and LEP. Although no significant changes in glucose and insulin plasma levels were observed in these animals, their HOMA-IR (homoeostasis model assessment of insulin resistance) values were significantly higher than those of CD. The AAT from FRD-fed rats had larger adipocytes, higher saturated FA content, higher NADPH oxidase activity, greater ROS production, a distorted FA content/release pattern, lower insulin sensitivity together with higher and lower mRNA content of LEP and IRS-1-/2 respectively, and released a larger amount of LEP. The development of all the clinical, OS, metabolic, endocrine and molecular changes induced by the FRD were significantly prevented by APO co-administration. The fact that APO treatment prevented both changes in NADPH oxidase activity and the development of all the FRD-induced AAT dysfunctions in normal rats strongly suggests that OS plays an important role in the FRD-induced MS (metabolic syndrome) phenotype.

The “gating” residues Ile199 and Tyr326 in human monoamine oxidase B function in substrate and inhibitor recognition

PubMed Central

Milczek, Erika M.; Binda, Claudia; Rovida, Stefano; Mattevi, Andrea; Edmondson, Dale E.

2011-01-01

Summary The major structural difference between human monoamine oxidases A (MAO A) and B (MAO B) is that MAO A has a monopartite substrate cavity of ~550 Å3 volume and MAO B contains a dipartite cavity structure with volumes of ~290 Å3 (entrance cavity) and ~400 Å3 (substrate cavity). Ile199 and Tyr326 side chains separate these two cavities in MAO B. To probe the function of these gating residues, Ile199Ala and Ile199Ala Tyr326Ala mutant forms of MAO B were investigated. Structural data on the Ile199Ala MAO B mutant show no alterations in active site geometries compared to WT enzyme while the Ile199Ala-Tyr326Ala MAO B mutant exhibits alterations in residues 100–103 which are part of the loop gating the entrance to the active site. Both mutant enzymes exhibit catalytic properties with increased amine KM but unaltered kcat values. The altered KM values on mutation are attributed to the influence of the cavity structure in the binding and subsequent deprotonation of the amine substrate. Both mutant enzymes exhibit weaker binding affinities relative to WT enzyme for small reversible inhibitors. Ile199Ala MAO B exhibits an increase in binding affinity for reversible MAO B specific inhibitors which bridge both cavities. The Ile199Ala-Tyr326Ala double mutant exhibits inhibitor binding properties more similar to those of MAO A than to MAO B. These results demonstrate the bipartite cavity structure in MAO B plays an important role in substrate and inhibitor recognition to distinguish its specificities from those of MAO A and provides insights into specific reversible inhibitor design for these membrane-bound enzymes. PMID:21978362

Computational tool for immunotoxic assessment of pyrethroids toward adaptive immune cell receptors

PubMed Central

Kumar, Anoop; Behera, Padma Charan; Rangra, Naresh Kumar; Dey, Suddhasattya; Kant, Kamal

2018-01-01

Background: Pyrethroids have prominently known for their insecticidal actions worldwide, but recent reports as anticancer and antiviral applications gained a lot of interest to further understand their safety and immunotoxicity. Objective: This encouraged us to carry out our present study to evaluate the interactions of pyrethroids toward adaptive immune cell receptors. Materials and Methods: Type 1 and Type 2 pyrethroids were tested on T (CD4 and CD8) and B (CD28 and CD45) immune cell receptors using Maestro 9.3 (Schrödinger, LLC, Cambridge, USA). In addition, top-ranked tested ligands were too explored for toxicity prediction in rodents using ProTOX tool. Results: Pyrethroids (specifically type 2) such as fenvalerate (−5.534 kcal/mol: CD8), fluvalinate (−4.644 and − 4.431 kcal/mol: CD4 and CD45), and cypermethrin (−3.535 kcal/mol: CD28) have outcome in less energy or more affinity for B-cell and T-cell immune receptors which may later result in the immunosuppressive and hypersensitivity reactions. Conclusion: The current findings have uncovered that there is a further need to assess the Type 2 pyrethroids with wet laboratory experiments to understand the chemical nature of pyrethroid-induced immunotoxicity. SUMMARY Fenvalerate showed apex glide score toward CD8 immune receptor, while fluvalinate confirmed top-ranked binding with CD4 and CD45 immune proteinsIn addition, cypermethrin outcame in top glide score against CD28 immune receptorTop dock hits (Type 2) pyrethroids have shown probable toxicity targets toward AOFA: Amine oxidase (flavin-containing) A and PGH1: Prostaglandin G/H synthase 1, respectively. Abbreviations used: PDB: Protein Data Bank; AOFA: Amine oxidase (flavin-containing) A; PGH 1: Prostaglandin G/H synthase 1. PMID:29576712

Computational tool for immunotoxic assessment of pyrethroids toward adaptive immune cell receptors.

PubMed

Kumar, Anoop; Behera, Padma Charan; Rangra, Naresh Kumar; Dey, Suddhasattya; Kant, Kamal

2018-01-01

Pyrethroids have prominently known for their insecticidal actions worldwide, but recent reports as anticancer and antiviral applications gained a lot of interest to further understand their safety and immunotoxicity. This encouraged us to carry out our present study to evaluate the interactions of pyrethroids toward adaptive immune cell receptors. Type 1 and Type 2 pyrethroids were tested on T (CD4 and CD8) and B (CD28 and CD45) immune cell receptors using Maestro 9.3 (Schrödinger, LLC, Cambridge, USA). In addition, top-ranked tested ligands were too explored for toxicity prediction in rodents using ProTOX tool. Pyrethroids (specifically type 2) such as fenvalerate (-5.534 kcal/mol: CD8), fluvalinate (-4.644 and - 4.431 kcal/mol: CD4 and CD45), and cypermethrin (-3.535 kcal/mol: CD28) have outcome in less energy or more affinity for B-cell and T-cell immune receptors which may later result in the immunosuppressive and hypersensitivity reactions. The current findings have uncovered that there is a further need to assess the Type 2 pyrethroids with wet laboratory experiments to understand the chemical nature of pyrethroid-induced immunotoxicity. Fenvalerate showed apex glide score toward CD8 immune receptor, while fluvalinate confirmed top-ranked binding with CD4 and CD45 immune proteinsIn addition, cypermethrin outcame in top glide score against CD28 immune receptorTop dock hits (Type 2) pyrethroids have shown probable toxicity targets toward AOFA: Amine oxidase (flavin-containing) A and PGH1: Prostaglandin G/H synthase 1, respectively. Abbreviations used: PDB: Protein Data Bank; AOFA: Amine oxidase (flavin-containing) A; PGH 1: Prostaglandin G/H synthase 1.

Empirical Valence Bond Simulations of the Hydride-Transfer Step in the Monoamine Oxidase A Catalyzed Metabolism of Noradrenaline.

PubMed

Poberžnik, Matic; Purg, Miha; Repič, Matej; Mavri, Janez; Vianello, Robert

2016-11-10

Monoamine oxidases (MAOs) A and B are flavoenzymes responsible for the metabolism of biogenic amines, such as dopamine, serotonin, and noradrenaline (NA), which is why they have been extensively implicated in the etiology and course of various neurodegenerative disorders and, accordingly, used as primary pharmacological targets to treat these debilitating cognitive diseases. The precise chemical mechanism through which MAOs regulate the amine concentration, which is vital for the development of novel inhibitors, is still not unambiguously determined in the literature. In this work, we present atomistic empirical valence bond simulations of the rate-limiting step of the MAO-A-catalyzed NA (norepinephrine) degradation, involving hydride transfer from the substrate α-methylene group to the flavin moiety of the flavin adenine dinucleotide prosthetic group, employing the full dimensionality and thermal fluctuations of the hydrated enzyme, with extensive configurational sampling. We show that MAO-A lowers the free energy of activation by 14.3 kcal mol -1 relative to that of the same reaction in aqueous solution, whereas the calculated activation free energy of ΔG ‡ = 20.3 ± 1.6 kcal mol -1 is found to be in reasonable agreement with the correlated experimental value of 16.5 kcal mol -1 . The results presented here strongly support the fact that both MAO-A and MAO-B isoforms function by the same hydride-transfer mechanism. We also considered a few point mutations of the "aromatic cage" tyrosine residue (Tyr444Phe, Tyr444Leu, Tyr444Trp, Tyr444His, and Tyr444Glu), and the calculated changes in the reaction barriers are in agreement with the experimental values, thus providing further support to the proposed mechanism.

Effect of sulfite treatment on total antioxidant capacity, total oxidant status, lipid hydroperoxide, and total free sulfydryl groups contents in normal and sulfite oxidase-deficient rat plasma.

PubMed

Herken, Emine Nur; Kocamaz, Erdogan; Erel, Ozcan; Celik, Hakim; Kucukatay, Vural

2009-08-01

Sulfites, which are commonly used as preservatives, are continuously formed in the body during the metabolism of sulfur-containing amino acids. Sulfite oxidase (SOX) is an essential enzyme in the pathway of the oxidative degradation of sulfite to sulfate protecting cells from sulfite toxicity. This article investigated the effect of sulfite on total antioxidant capacity (TAC), total oxidant status, lipid hydroperoxide (LOOH), and total free sulfydryl groups (-SH) levels in normal and SOX-deficient male albino rat plasma. For this purpose, rats were divided into four groups: control, sulfite-treated, SOX-deficient, and sulfite-treated SOX-deficient groups. SOX deficiency was established by feeding rats a low molybdenum diet and adding to their drinking water 200 ppm tungsten. Sulfite (70 mg/kg) was administered to the animals via their drinking water. SOX deficiency together with sulfite treatment caused a significant increase in the plasma LOOH and total oxidant status levels. -SH content of rat plasma significantly decreased by both sulfite treatment and SOX deficiency compared to the control. There was also a significant decrease in plasma TAC level by sulfite treatment. In conclusion, sulfite treatment affects the antioxidant/oxidant balance of the plasma cells of the rats toward oxidants in SOX-deficient groups.

Probing the mechanism of proton coupled electron transfer to dioxygen: the oxidative half-reaction of bovine serum amine oxidase.

PubMed

Su, Q; Klinman, J P

1998-09-08

Bovine serum amine oxidase (BSAO) catalyzes the oxidative deamination of primary amines, concomitant with the reduction of molecular oxygen to hydrogen peroxide via a ping-pong mechanism. A protocol has been developed for an analysis of chemical and kinetic mechanisms in the conversion of dioxygen to hydrogen peroxide. Steady-state kinetics show that two groups need to be deprotonated to facilitate the oxidative half-reaction. The pH dependence of Vmax/Km(O2) reveals pKa's of 6.2 +/- 0.3 and 7.0 +/- 0.2, respectively. A pKa of 7.2 +/- 0.1 has been obtained from a titration of anaerobically reduced BSAO using UV-vis spectrophotometry. The near identity of the pKa obtained from the reduced enzyme titration with the second pKa from steady-state kinetics suggests that this second pKa arises from the reduced cofactor. The assignment of pKa is supported by the observed pH dependence for formation of the cofactor semiquinone signal, detected by EPR spectroscopy under anaerobic conditions. To address the nature of rate-limiting steps in the oxidative half-reaction, the solvent isotope effect, viscosity effect, and O-18 isotope effect on Vmax/Km(O2) have been determined. The solvent isotope effect is indistinguishable from unity, ruling out a proton transfer as a rate-determining step. Use of glucose as a solvent viscosogen shows no viscosity effect, indicating that binding of oxygen is not in the rate-determining step. The O-18 kinetic isotope effect is independent of pH with an average value of 18(V/K) = 1.0097 +/- 0. 0010. This has been compared to calculated equilibrium O-18 isotope effects for various dioxygen intermediate species [Tian and Klinman (1993) J. Am. Chem. Soc. 115, 8891], leading to the conclusion that either the first electron transfer to dioxygen or the desorption of product peroxide from a Cu(II)-OOH complex could be the rate-limiting step. The distribution of steady-state enzyme species was, therefore, analyzed through a combination of stopped-flow experiments and analysis of DV and D(V/K) for benzylamine oxidation. We conclude that the major species accumulating in the steady state are the oxidized cofactor-substrate Schiff base complex and the reduced, aminoquinol form of cofactor. These data rule out a slow release of product hydroperoxide from the aminoquinone form of enzyme, leading to the conclusion that the first electron transfer from substrate-reduced cofactor to dioxygen is the rate-determining step in the oxidative half-reaction. This step is also estimated to be 40% rate-limiting in kcat. An important mechanistic conclusion from this study is that dioxygen binding is a separate step from the rate-limiting electron-transfer step to form superoxide. On the basis of a recently determined X-ray structure for the active form of a yeast amine oxidase from Hansenula polymorpha [Li et al. (1998) Structure 6, 293], a hydrophobic space has been identified near the O-2 position of reduced cofactor as the putative dioxygen binding site. Movement of superoxide from this site onto the Cu(II) at the active site may occur prior to further electron transfer from cofactor to superoxide.

The voltage dependence of NADPH oxidase reveals why phagocytes need proton channels

NASA Astrophysics Data System (ADS)

DeCoursey, Thomas E.; Morgan, Deri; Cherny, Vladimir V.

2003-04-01

The enzyme NADPH oxidase in phagocytes is important in the body's defence against microbes: it produces superoxide anions (O2-, precursors to bactericidal reactive oxygen species). Electrons move from intracellular NADPH, across a chain comprising FAD (flavin adenine dinucleotide) and two haems, to reduce extracellular O2 to O2-. NADPH oxidase is electrogenic, generating electron current (Ie) that is measurable under voltage-clamp conditions. Here we report the complete current-voltage relationship of NADPH oxidase, the first such measurement of a plasma membrane electron transporter. We find that Ie is voltage-independent from -100mV to >0mV, but is steeply inhibited by further depolarization, and is abolished at about +190mV. It was proposed that H+ efflux mediated by voltage-gated proton channels compensates Ie, because Zn2+ and Cd2+ inhibit both H+ currents and O2- production. Here we show that COS-7 cells transfected with four NADPH oxidase components, but lacking H+ channels, produce O2- in the presence of Zn2+ concentrations that inhibit O2- production in neutrophils and eosinophils. Zn2+ does not inhibit NADPH oxidase directly, but through effects on H+ channels. H+ channels optimize NADPH oxidase function by preventing membrane depolarization to inhibitory voltages.

A new approach to the immobilisation of poly(ethylene oxide) for the reduction of non-specific protein adsorption on conductive substrates

NASA Astrophysics Data System (ADS)

Cole, Martin A.; Thissen, Helmut; Losic, Dusan; Voelcker, Nicolas H.

2007-04-01

Biomedical and biotechnological devices often require surface modifications to improve their performance. In most cases, uniform coatings are desired which provide a specific property or lead to a specific biological response. In the present work, we have generated pinhole-free coatings providing amine functional groups achieved by electropolymerisation of tyramine on highly doped silicon substrates. Furthermore, amine groups were used for the subsequent grafting of poly(ethylene oxide) aldehyde via reductive amination. All surface modification steps were characterized by X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). The results indicate that the stability and the density of amine functional groups introduced at the surface via electropolymerisation compare favourably with alternative coatings frequently used in biomedical and biotechnological devices such as plasma polymer films. Furthermore, protein adsorption on amine and poly(ethylene oxide) coatings was studied by XPS and a colorimetric assay to test enzymatic activity. The grafting of poly(ethylene oxide) under cloud point conditions on electropolymerised tyramine layers resulted in surfaces with extremely low protein fouling character.

Polyamines and Their Metabolites as Diagnostic Markers of Human Diseases

PubMed Central

Park, Myung Hee; Igarashi, Kazuei

2013-01-01

Polyamines, putrescine, spermidine and spermine, are ubiquitous in living cells and are essential for eukaryotic cell growth. These polycations interact with negatively charged molecules such as DNA, RNA, acidic proteins and phospholipids and modulate various cellular functions including macromolecular synthesis. Dysregulation of the polyamine pathway leads to pathological conditions including cancer, inflammation, stroke, renal failure and diabetes. Increase in polyamines and polyamine synthesis enzymes is often associated with tumor growth, and urinary and plasma contents of polyamines and their metabolites have been investigated as diagnostic markers for cancers. Of these, diacetylated derivatives of spermidine and spermine are elevated in the urine of cancer patients and present potential markers for early detection. Enhanced catabolism of cellular polyamines by polyamine oxidases (PAO), spermine oxidase (SMO) or acetylpolyamine oxidase (AcPAO), increases cellular oxidative stress and generates hydrogen peroxide and a reactive toxic metabolite, acrolein, which covalently incorporates into lysine residues of cellular proteins. Levels of protein-conjuagated acrolein (PC-Acro) and polyamine oxidizing enzymes were increased in the locus of brain infarction and in plasma in a mouse model of stroke and also in the plasma of stroke patients. When the combined measurements of PC-Acro, interleukin 6 (IL-6), and C-reactive protein (CRP) were evaluated, even silent brain infarction (SBI) was detected with high sensitivity and specificity. Considering that there are no reliable biochemical markers for early stage of stroke, PC-Acro and PAOs present promising markers. Thus the polyamine metabolites in plasma or urine provide useful tools in early diagnosis of cancer and stroke. PMID:24009852

The transdermal delivery system of monoamine oxidase inhibitors.

PubMed

VanDenBerg, Chad M

2012-01-01

Monoamine oxidase inhibitors (MAOIs) were once widely used as effective treatments for major depressive disorder, particularly for patients with atypical or treatment-resistant depression. Today, MAOIs have largely been replaced by newer antidepressants because of concerns over potential serious side effects due to their mechanism of action. Monoamine oxidase (MAO) is an enzyme that metabolizes serotonin, norepinephrine, and dopamine, the neurotransmitters that are most associated with depression; inhibiting MAO, therefore, makes more of these neurotransmitters available for synaptic action. However, MAO also metabolizes tyramine, a trace amine found in some foods that acts as a sympathomimetic. Allowing excess tyramine to accumulate via MAO inhibition can result in hypertensive crisis due to the release of norepinephrine; therefore, patients taking an MAOI have had to follow dietary restrictions to avoid tyramine-rich foods. Hypertensive crisis may also be precipitated by using MAOIs in conjunction with other drugs that have vasoconstrictive properties, that act as sympathomimetics, or that inhibit the reuptake of norepinephrine. Serotonin syndrome is another serious adverse effect that can potentially occur when using an MAOI with another drug that inhibits the reuptake of serotonin. In this article, the mechanism of action of MAOIs is reviewed, along with that of a newer MAOI formulation that lessens the need for dietary restrictions and has a greater safety and tolerability profile than the older oral formulations. © Copyright 2012 Physicians Postgraduate Press, Inc.

Hypoxia-Inducible Factor 1-Regulated Lysyl Oxidase Is Involved in Staphylococcus aureus Abscess Formation

PubMed Central

Beerlage, Christiane; Greb, Jessica; Kretschmer, Dorothee; Assaggaf, Mohammad; Trackman, Philip C.; Hansmann, Martin-Leo; Bonin, Michael; Eble, Johannes A.; Peschel, Andreas; Brüne, Bernhard

2013-01-01

Hypoxia-inducible factor 1 (HIF-1) is the key transcription factor involved in the adaptation of mammals to hypoxia and plays a crucial role in cancer angiogenesis. Recent evidence suggests a leading role for HIF-1 in various inflammatory and infectious diseases. Here we describe the role of HIF-1 in Staphylococcus aureus infections by investigating the HIF-1-dependent host cell response. For this purpose, transcriptional profiling of HIF-1α-deficient HepG2 and control cells, both infected with Staphylococcus aureus, was performed. Four hours after infection, the expression of 190 genes, 24 of which were regulated via HIF-1, was influenced. LOX (encoding lysyl oxidase) was one of the upregulated genes with a potential impact on the course of S. aureus infection. LOX is an amine oxidase required for biosynthetic cross-linking of extracellular matrix components. LOX was upregulated in vitro in different cell cultures infected with S. aureus and also in vivo, in kidney abscesses of mice intravenously infected with S. aureus and in clinical skin samples from patients with S. aureus infections. Inhibition of LOX by β-aminopropionitrile (BAPN) did not affect the bacterial load in kidneys or blood but significantly influenced abscess morphology and collagenization. Our data provide evidence for a crucial role of HIF-1-regulated LOX in abscess formation. PMID:23649089

The trimethylammonium headgroup of choline is a major determinant for substrate binding and specificity in choline oxidase.

PubMed

Gadda, Giovanni; Powell, Nichole L N; Menon, Prashanthi

2004-10-15

Choline oxidase catalyzes the oxidation of choline to glycine betaine via two sequential flavin-linked transfers of hydride equivalents to molecular oxygen and formation of a betaine aldehyde intermediate. In the present study, choline and glycine betaine analogs were used as substrates and inhibitors for the enzyme to investigate the structural determinants that are relevant for substrate recognition and specificity. Competitive inhibition patterns with respect to choline were determined for a number of substituted amines at pH 6.5 and 25 degrees C. The Kis values for the carboxylate-containing ligands glycine betaine, N,N-dimethylglycine, and N-methylglycine increased monotonically with decreasing number of methyl groups, consistent with the trimethylammonium portion of the ligand being important for binding. In contrast, the acetate portion of glycine betaine did not contribute to binding, as suggested by lack of changes in the Kis values upon substituting glycine betaine with inhibitors containing methyl, ethyl, allyl, and 2-amino-ethyl side chains. In agreement with the inhibition data, the specificity of the enzyme for the organic substrate (kcat/Km value) decreased when N,N-dimethylethanolamine, N-methylethanolamine, and the isosteric substrate 3,3-dimethyl-1-butanol were used as substrate instead of choline; a contribution of approximately 7 kcal mol(-1) toward substrate discrimination was estimated for the interaction of the trimethylammonium portion of the substrate with the active site of choline oxidase.

Genomic identification and biochemical characterization of the mammalian polyamine oxidase involved in polyamine back-conversion.

PubMed Central

Vujcic, Slavoljub; Liang, Ping; Diegelman, Paula; Kramer, Debora L; Porter, Carl W

2003-01-01

In the polyamine back-conversion pathway, spermine and spermidine are first acetylated by spermidine/spermine N1 -acetyltransferase (SSAT) and then oxidized by polyamine oxidase (PAO) to produce spermidine and putrescine respectively. Although PAO was first purified more than two decades ago, the protein has not yet been linked to genomic sequences. In the present study, we apply a BLAST search strategy to identify novel oxidase sequences located on human chromosome 10 and mouse chromosome 7. Homologous mammalian cDNAs derived from human brain and mouse mammary tumour were deduced to encode proteins of approx. 55 kDa having 82% sequence identity. When either cDNA was transiently transfected into HEK-293 cells, intracellular spermine pools decreased by approx. 30%, whereas spermidine increased 2-4-fold. Lysates of human PAO cDNA-transfected HEK-293 cells, but not vector-transfected cells, rapidly oxidized N1-acetylspermine to spermidine. Substrate specificity determinations with the lysate assay revealed a preference ranking of N1-acetylspermine= N1-acetylspermidine> N1,N12-diacetylspermine>>spermine; spermidine was not acted upon. This ranking is identical to that reported for purified PAO and distinctly different from the recently identified spermine oxidase (SMO), which prefers spermine over N1-acetylspermine. Monoethyl- and diethylspermine analogues also served as substrates for PAO, and were internally cleaved adjacent to a secondary amine. We deduce that the present oxidase sequences are those of the FAD-dependent PAO involved in the polyamine back-conversion pathway. In Northern blot analysis, PAO mRNA was much less abundant in HEK-293 cells than SMO or SSAT mRNA, and all three were differentially induced in a similar manner by selected polyamine analogues. The identification of PAO sequences, together with the recently identified SMO sequences, provides new opportunities for understanding the dynamics of polyamine homoeostasis and for interpreting metabolic and cellular responses to clinically-relevant polyamine analogues and inhibitors. PMID:12477380

Plasma membrane NADH oxidase of maize roots responds to gravity and imposed centrifugal forces

NASA Technical Reports Server (NTRS)

Bacon, E.; Morre, D. J.

2001-01-01

NADH oxidase activities measured with excised roots of dark-grown maize (Zea mays) seedlings and with isolated plasma membrane vesicles from roots of dark-grown maize oscillated with a regular period length of 24 min and were inhibited by the synthetic auxin 2,4-dichlorophenoxyacetic [correction of dichorophenoxyacetic] acid. The activities also responded to orientation with respect to gravity and to imposed centrifugal forces. Turning the roots upside down resulted in stimulation of the activity with a lag of about 10 min. Returning the sections to the normal upright position resulted in a return to initial rates. The activity was stimulated reversibly to a maximum of about 2-fold with isolated plasma membrane vesicles, when subjected to centrifugal forces of 25 to 250 x g for 1 to 4 min duration. These findings are the first report of a gravity-responsive enzymatic activity of plant roots inhibited by auxin and potentially related to the gravity-induced growth response. c2001 Editions scientifiques et medicales Elsevier SAS.

The monoamine oxidase A gene promoter repeat and prostate cancer risk.

PubMed

White, Thomas A; Kwon, Erika M; Fu, Rong; Lucas, Jared M; Ostrander, Elaine A; Stanford, Janet L; Nelson, Peter S

2012-11-01

Amine catabolism by monoamine oxidase A (MAOA) contributes to oxidative stress, which plays a role in prostate cancer (PCa) development and progression. An upstream variable-number tandem repeat (uVNTR) in the MAOA promoter influences gene expression and activity, and may thereby affect PCa susceptibility. Caucasian (n = 2,572) men from two population-based case-control studies of PCa were genotyped for the MAOA-VNTR. Logistic regression was used to assess PCa risk in relation to genotype. Common alleles of the MAOA-VNTR were not associated with the relative risk of PCa, nor did the relationship differ by clinical features of the disease. The rare 5-copy variant (frequency: 0.5% in cases; 1.8% in controls), however, was associated with a reduced PCa risk (odds ratio, OR = 0.30, 95% CI 0.13-0.71). A rare polymorphism of the MAOA promoter previously shown to confer low expression was associated with a reduced risk of developing PCa. This novel finding awaits confirmation in other study populations. Copyright © 2012 Wiley Periodicals, Inc.

Synthesis and evaluation of biaryl derivatives for structural characterization of selective monoamine oxidase B inhibitors toward Parkinson's disease therapy.

PubMed

Yeon, Seul Ki; Choi, Ji Won; Park, Jong-Hyun; Lee, Ye Rim; Kim, Hyeon Jeong; Shin, Su Jeong; Jang, Bo Ko; Kim, Siwon; Bahn, Yong-Sun; Han, Gyoonhee; Lee, Yong Sup; Pae, Ae Nim; Park, Ki Duk

2018-01-01

Benzyloxyphenyl moiety is a common structure of highly potent, selective and reversible inhibitors of monoamine oxidase B (MAO-B), safinamide and sembragiline. We synthesized 4-(benzyloxy)phenyl and biphenyl-4-yl derivatives including halogen substituents on the terminal aryl unit. In addition, we modified the carbon linker between amine group and the biaryl linked unit. Among synthesized compounds, 12c exhibited the most potent and selective MAO-B inhibitory effect (hMAO-B IC 50 : 8.9 nM; >10,000-fold selectivity over MAO-A) as a competitive inhibitor. In addition, 12c showed greater MAO-B inhibitory activity and selectivity compared to well-known MAO-B inhibitors such as selegiline, safinamide and sembragiline. In the MPTP-induced mouse model of Parkinson's disease (PD), 12c significantly protected the tyrosine hydroxylase (TH)-immunopositive DAergic neurons and attenuated the PD-associated behavioral deficits. This study suggests characteristic structures as a MAO-B inhibitor that may provide a good insight for the development of therapeutic agents for PD. Copyright © 2017 Elsevier Ltd. All rights reserved.

Monoamine oxidases as sources of oxidants in the heart

PubMed Central

Kaludercic, Nina; Mialet-Perez, Jeanne; Paolocci, Nazareno; Parini, Angelo; Di Lisa, Fabio

2014-01-01

Oxidative stress can be generated at several sites within the mitochondria. Among these, monoamine oxidases (MAO) have been described as a prominent source. MAO are mitochondrial flavoenzymes responsible for the oxidative deamination of catecholamines, serotonin and biogenic amines, and during this process they generate H2O2 and aldehyde intermediates. The role of MAO in cardiovascular pathophysiology has only recently gathered some attention since it has been demonstrated that both H2O2 and aldehydes may target mitochondrial function and consequently affect function and viability of the myocardium. In the present review, we will discuss the role of MAO in catecholamine and serotonin clearance and cycling in relation to cardiac structure and function. The relevant contribution of each MAO isoform (MAO-A or -B) will be discussed in relation to mitochondrial dysfunction and myocardial injury. Finally, we will examine both beneficial effects of their pharmacological or genetic inhibition along with potential adverse effects observed at baseline in MAO knockout mice, as well as the deleterious effects following their over-expression specifically at cardiomyocyte level. PMID:24412580

The Role of Aldehyde Oxidase and Xanthine Oxidase in the Biotransformation of a Novel Negative Allosteric Modulator of Metabotropic Glutamate Receptor Subtype 5

PubMed Central

Morrison, Ryan D.; Blobaum, Anna L.; Byers, Frank W.; Santomango, Tammy S.; Bridges, Thomas M.; Stec, Donald; Brewer, Katrina A.; Sanchez-Ponce, Raymundo; Corlew, Melany M.; Rush, Roger; Felts, Andrew S.; Manka, Jason; Bates, Brittney S.; Venable, Daryl F.; Rodriguez, Alice L.; Jones, Carrie K.; Niswender, Colleen M.; Conn, P. Jeffrey; Lindsley, Craig W.; Emmitte, Kyle A.

2012-01-01

Negative allosteric modulation (NAM) of metabotropic glutamate receptor subtype 5 (mGlu5) represents a therapeutic strategy for the treatment of childhood developmental disorders, such as fragile X syndrome and autism. VU0409106 emerged as a lead compound within a biaryl ether series, displaying potent and selective inhibition of mGlu5. Despite its high clearance and short half-life, VU0409106 demonstrated efficacy in rodent models of anxiety after extravascular administration. However, lack of a consistent correlation in rat between in vitro hepatic clearance and in vivo plasma clearance for the biaryl ether series prompted an investigation into the biotransformation of VU0409106 using hepatic subcellular fractions. An in vitro appraisal in rat, monkey, and human liver S9 fractions indicated that the principal pathway was NADPH-independent oxidation to metabolite M1 (+16 Da). Both raloxifene (aldehyde oxidase inhibitor) and allopurinol (xanthine oxidase inhibitor) attenuated the formation of M1, thus implicating the contribution of both molybdenum hydroxylases in the biotransformation of VU0409106. The use of 18O-labeled water in the S9 experiments confirmed the hydroxylase mechanism proposed, because 18O was incorporated into M1 (+18 Da) as well as in a secondary metabolite (M2; +36 Da), the formation of which was exclusively xanthine oxidase-mediated. This unusual dual and sequential hydroxylase metabolism was confirmed in liver S9 and hepatocytes of multiple species and correlated with in vivo data because M1 and M2 were the principal metabolites detected in rats administered VU0409106. An in vitro-in vivo correlation of predicted hepatic and plasma clearance was subsequently established for VU0409106 in rats and nonhuman primates. PMID:22711749

Allelic variations in the CYBA gene of NADPH oxidase and risk of kidney complications in patients with type 1 diabetes.

PubMed

Patente, Thiago A; Mohammedi, Kamel; Bellili-Muñoz, Naïma; Driss, Fathi; Sanchez, Manuel; Fumeron, Frédéric; Roussel, Ronan; Hadjadj, Samy; Corrêa-Giannella, Maria Lúcia; Marre, Michel; Velho, Gilberto

2015-09-01

Oxidative stress plays a pivotal role in the pathophysiology of diabetic nephropathy, and the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase system is an important source of reactive oxygen species in hyperglycemic conditions in the kidney. Plasma concentration of advanced oxidation protein products (AOPP), a marker of oxidative stress, is increased in patients with diabetic nephropathy. We investigated associations of variants in the CYBA gene, encoding the regulatory subunit p22(phox) of NADPH oxidase, with diabetic nephropathy and plasma AOPP and myeloperoxidase (MPO) concentrations in type 1 diabetic patients. Seven SNPs in the CYBA region were analyzed in 1357 Caucasian subjects with type 1 diabetes from the SURGENE (n=340), GENEDIAB (n=444), and GENESIS (n=573) cohorts. Duration of follow-up was 10, 9, and 6 years, respectively. Cox proportional hazards and logistic regression analyses were used to estimate hazard ratios (HR) or odds ratios (OR) for incidence and prevalence of diabetic nephropathy. The major G-allele of rs9932581 was associated with the incidence of renal events defined as new cases of microalbuminuria or the progression to a more severe stage of nephropathy during follow-up (HR 1.59, 95% CI 1.17-2.18, P=0.003) in SURGENE. The same allele was associated with established/advanced nephropathy (OR 1.52, 95% CI 1.22-1.92, P=0.0001) and with the incidence of end-stage renal disease (ESRD) (HR 2.01, 95% CI 1.30-3.24, P=0.001) in GENEDIAB/GENESIS pooled studies. The risk allele was also associated with higher plasma AOPP concentration in subsets of SURGENE and GENEDIAB, with higher plasma MPO concentration in a subset of GENEDIAB, and with lower estimated glomerular filtration rate (eGFR) in the three cohorts. In conclusion, a functional variant in the promoter of the CYBA gene was associated with lower eGFR and with prevalence and incidence of diabetic nephropathy and ESRD in type 1 diabetic patients. These results are consistent with a role for NADPH oxidase in the pathophysiology of kidney complications of diabetes. Copyright © 2015 Elsevier Inc. All rights reserved.

Platelet monoamine oxidase B and plasma β-phenylethylamine in Parkinson's disease

PubMed Central

Zhou, G; Miura, Y; Shoji, H; Yamada, S; Matsuishi, T

2001-01-01

OBJECTIVE—To evaluate the correlation between changes in platelet monoamine oxidase type B (MAO-B) activity and plasma β-phenylethylamine (PEA) concentrations in patients with Parkinson's disease and controls.
METHODS—Platelet MAO-B activity and plasma PEA were measured with gas chromatography-mass spectrometry (GC-MS) in patients with Parkinson's disease treated with levodopa (12 men and 12 women) or selegiline (three men and three women), and physically healthy subjects as a control group (10 men and 10women).
RESULTS—Platelet MAO-B activity was significantly higher in the Parkinson's disease group (mean 542 (SD 318) pmol/107 platelets/30 min) than in the control group (mean 349 (SD 307) pmol/107 platelets/30 min) (p<0.05). By contrast, the plasma PEA concentrations in patients with Parkinson's disease were significantly lower than in the control group (mean 532 (SD 243) pg/ml; 931 (SD 560) pg/ml) (p<0.01). The plasma PEA concentrations in patients with Parkinson's disease treated with selegiline were prominently higher than in patients with no selegiline treatment (p<0.001). There was a significantly negative correlation between platelet MAO-B activity and plasma PEA concentrations in patients (n=24, r=−0.466, p<0.001).
CONCLUSIONS—The increase in platelet MAO-B activity and decrease in plasma PEA concentrations in patients with Parkinson's disease may be involved in the pathophysiological processes of the disease, and these changes are reversed by treatment with selegiline.

 PMID:11160474

Plasma concentration of diamine oxidase (DAO) predicts 1-month mortality of acute-on-chronic hepatitis B liver failure.

PubMed

Li, Feng-Cai; Li, Yue-Kai; Fan, Yu-Chen; Wang, Kai

2018-05-26

Acute-on-chronic hepatitis B liver failure (ACHBLF) has high 1-month mortality but it is difficult to predict. This present study was aimed to determine the diagnostic value of plasma diamine oxidase (DAO) in predicting the 1-month mortality of ACHBLF. A total of 106 consecutive newly diagnosed ACHBLF patients were retrospectively collected. The plasma expression of DAO was determined using enzyme-linked immunosorbent assay (ELISA). The plasma DAO level of survivals [14.0 (7.1; 26.5) ng/mL] was significantly lower than the nonsurvivals [58.6 (32.5; 121.3) ng/mL, P < .001]. The plasma DAO level, hepatic encephalopathy, spontaneous bacterial peritonitis and model for end-stage liver disease (MELD) score were independent factors associated with the 1-month mortality for ACHBLF. The cut-off point of 15.2 ng/mL for plasma DAO level with sensitivity of 95.45%, specificity of 62.5%, 22.6 for MELD score with sensitivity of 90.91%, specificity of 67.5%, 0.07 for DAO plus MELD with sensitivity of 87.88%, specificity of 80% were selected to discriminate 1-month morality of ACHBLF. Furthermore, DAO plus MELD score showed high AUROC than MELD score for predicting 1-month (0.916 vs. 0.843, P < .01). The plasma DAO level plus MELD > 0.07 predicts poor 1-month mortality of ACHBLF. Copyright © 2018 Elsevier B.V. All rights reserved.

Mechanistic and Structural Analyses of the Role of His67 in the Yeast Polyamine Oxidase Fms1†

PubMed Central

Adachi, Mariya S.; Taylor, Alexander B.; Hart, P. John; Fitzpatrick, Paul F.

2012-01-01

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N1-acetylspermine to spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. Within the active site of Fms1, His67 is positioned to form hydrogen bonds with the polyamine substrate. This residue is also conserved in other polyamine oxidases. The catalytic properties of H67Q, H67N, and H67A Fms1 have been characterized to evaluate the role of this residue in catalysis. With both spermine and N1-acetylspermine as the amine substrate, the value of the first-order rate constant for flavin reduction decreases 2–3 orders of magnitude, with the H67Q mutation having the smallest effect and H67N the largest. The kcat/KO2 value changes very little upon mutation with N1-acetylspermine as the amine substrate and decreases only an order of magnitude with spermine. The kcat/KM-pH profiles with N1-acetylspermine are bell-shaped for all the mutants; the similarity to the profile of the wild-type enzyme rules out His67 as being responsible for either of the pKa values. The pH profiles for the rate constant for flavin reduction for all the mutant enzymes similarly show the same pKa as wild-type Fms1, about ~7.4; this pKa is assigned to the substrate N4. The kcat/KO2-pH profiles for wild-type Fms1 and the H67A enzyme both show a pKa of about ~6.9; this suggests His67 is not responsible for this pH behaviour. With the H67Q, H67N, and H67A enzymes the kcat value decreases when a single residue is protonated, as is the case with the wild-type enzyme. The structure of H67Q Fms1 has been determined at a resolution of 2.4 Å. The structure shows that the mutation disrupts a hydrogen bond network in the active site, suggesting that His67 is important both for direct interactions with the substrate and to maintain the overall active site structure. PMID:22642831

Genomic and experimental evidence for multiple metabolic functions in the RidA/YjgF/YER057c/UK114 (Rid) protein family

DOE Office of Scientific and Technical Information (OSTI.GOV)

Niehaus, Thomas D.; Gerdes, Svetlana; Hodge-Hanson, Kelsey

It is now recognized that enzymatic or chemical side-reactions can convert normal metabolites to useless or toxic ones and that a suite of enzymes exists to mitigate such metabolite damage. Examples are the reactive imine/enamine intermediates produced by threonine dehydratase, which damage the pyridoxal 5'-phosphate cofactor of various enzymes causing inactivation. This damage is pre-empted by RidA proteins, which hydrolyze the imines before they do harm. RidA proteins belong to the YjgF/YER057c/UK114 family (here renamed the Rid family). Most other members of this diverse and ubiquitous family lack defined functions. Phylogenetic analysis divided the Rid family into a widely distributed,more » apparently archetypal RidA subfamily and seven other subfamilies (Rid1 to Rid7) that are largely confined to bacteria and often co-occur in the same organism with RidA and each other. The Rid1 to Rid3 subfamilies, but not the Rid4 to Rid7 subfamilies, have a conserved arginine residue that, in RidA proteins, is essential for imine-hydrolyzing activity. Analysis of the chromosomal context of bacterial RidA genes revealed clustering with genes for threonine dehydratase and other pyridoxal 5'-phosphate-dependent enzymes, which fits with the known RidA imine hydrolase activity. Clustering was also evident between Rid family genes and genes specifying FAD-dependent amine oxidases or enzymes of carbamoyl phosphate metabolism. Biochemical assays showed that Salmonella enterica RidA and Rid2, but not Rid7, can hydrolyze imines generated by amino acid oxidase. Genetic tests indicated that carbamoyl phosphate overproduction is toxic to S. enterica cells lacking RidA, and metabolomic profiling of Rid knockout strains showed ten-fold accumulation of the carbamoyl phosphate-related metabolite dihydroorotate. Like the archetypal RidA subfamily, the Rid2, and probably the Rid1 and Rid3 subfamilies, have imine-hydrolyzing activity and can pre-empt damage from imines formed by amine oxidases as well as by pyridoxal 5'-phosphate enzymes. The RidA subfamily has an additional damage pre-emption role in carbamoyl phosphate metabolism that has yet to be biochemically defined. As a result, the Rid4 to Rid7 subfamilies appear not to hydrolyze imines and thus remain mysterious.« less

Genomic and experimental evidence for multiple metabolic functions in the RidA/YjgF/YER057c/UK114 (Rid) protein family

DOE PAGES

Niehaus, Thomas D.; Gerdes, Svetlana; Hodge-Hanson, Kelsey; ...

2015-05-15

It is now recognized that enzymatic or chemical side-reactions can convert normal metabolites to useless or toxic ones and that a suite of enzymes exists to mitigate such metabolite damage. Examples are the reactive imine/enamine intermediates produced by threonine dehydratase, which damage the pyridoxal 5'-phosphate cofactor of various enzymes causing inactivation. This damage is pre-empted by RidA proteins, which hydrolyze the imines before they do harm. RidA proteins belong to the YjgF/YER057c/UK114 family (here renamed the Rid family). Most other members of this diverse and ubiquitous family lack defined functions. Phylogenetic analysis divided the Rid family into a widely distributed,more » apparently archetypal RidA subfamily and seven other subfamilies (Rid1 to Rid7) that are largely confined to bacteria and often co-occur in the same organism with RidA and each other. The Rid1 to Rid3 subfamilies, but not the Rid4 to Rid7 subfamilies, have a conserved arginine residue that, in RidA proteins, is essential for imine-hydrolyzing activity. Analysis of the chromosomal context of bacterial RidA genes revealed clustering with genes for threonine dehydratase and other pyridoxal 5'-phosphate-dependent enzymes, which fits with the known RidA imine hydrolase activity. Clustering was also evident between Rid family genes and genes specifying FAD-dependent amine oxidases or enzymes of carbamoyl phosphate metabolism. Biochemical assays showed that Salmonella enterica RidA and Rid2, but not Rid7, can hydrolyze imines generated by amino acid oxidase. Genetic tests indicated that carbamoyl phosphate overproduction is toxic to S. enterica cells lacking RidA, and metabolomic profiling of Rid knockout strains showed ten-fold accumulation of the carbamoyl phosphate-related metabolite dihydroorotate. Like the archetypal RidA subfamily, the Rid2, and probably the Rid1 and Rid3 subfamilies, have imine-hydrolyzing activity and can pre-empt damage from imines formed by amine oxidases as well as by pyridoxal 5'-phosphate enzymes. The RidA subfamily has an additional damage pre-emption role in carbamoyl phosphate metabolism that has yet to be biochemically defined. As a result, the Rid4 to Rid7 subfamilies appear not to hydrolyze imines and thus remain mysterious.« less

Mechanistic and structural analyses of the role of His67 in the yeast polyamine oxidase Fms1.

PubMed

Adachi, Mariya S; Taylor, Alexander B; Hart, P John; Fitzpatrick, Paul F

2012-06-19

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N(1)-acetylspermine to spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. Within the active site of Fms1, His67 is positioned to form hydrogen bonds with the polyamine substrate. This residue is also conserved in other polyamine oxidases. The catalytic properties of H67Q, H67N, and H67A Fms1 have been characterized to evaluate the role of this residue in catalysis. With both spermine and N(1)-acetylspermine as the amine substrate, the value of the first-order rate constant for flavin reduction decreases 2-3 orders of magnitude, with the H67Q mutation having the smallest effect and H67N the largest. The k(cat)/K(O2) value changes very little upon mutation with N(1)-acetylspermine as the amine substrate and decreases only an order of magnitude with spermine. The k(cat)/K(M)-pH profiles with N(1)-acetylspermine are bell-shaped for all the mutants; the similarity to the profile of the wild-type enzyme rules out His67 as being responsible for either of the pK(a) values. The pH profiles for the rate constant for flavin reduction for all the mutant enzymes similarly show the same pK(a) as wild-type Fms1, about ∼7.4; this pK(a) is assigned to the substrate N4. The k(cat)/K(O2)-pH profiles for wild-type Fms1 and the H67A enzyme both show a pK(a) of about ∼6.9; this suggests His67 is not responsible for this pH behavior. With the H67Q, H67N, and H67A enzymes the k(cat) value decreases when a single residue is protonated, as is the case with the wild-type enzyme. The structure of H67Q Fms1 has been determined at a resolution of 2.4 Å. The structure shows that the mutation disrupts a hydrogen bond network in the active site, suggesting that His67 is important both for direct interactions with the substrate and to maintain the overall active site structure.

Upregulation of BMSCs Osteogenesis by Positively-Charged Tertiary Amines on Polymeric Implants via Charge/iNOS Signaling Pathway

PubMed Central

Zhang, Wei; Liu, Na; Shi, Haigang; Liu, Jun; Shi, Lianxin; Zhang, Bo; Wang, Huaiyu; Ji, Junhui; Chu, Paul K.

2015-01-01

Positively-charged surfaces on implants have a similar potential to upregulate osteogenesis of bone marrow-derived mesenchymal stem cells (BMSCs) as electromagnetic therapy approved for bone regeneration. Generally, their osteogenesis functions are generally considered to stem from the charge-induced adhesion of extracellular matrix (ECM) proteins without exploring the underlying surface charge/cell signaling molecule pathways. Herein, a positively-charged surface with controllable tertiary amines is produced on a polymer implant by plasma surface modification. In addition to inhibiting the TNF-α expression, the positively-charged surface with tertiary amines exhibits excellent cytocompatibility as well as remarkably upregulated osteogenesis-related gene/protein expressions and calcification of the contacted BMSCs. Stimulated by the charged surface, these BMSCs display high iNOS expressions among the three NOS isoforms. Meanwhile, downregulation of the iNOS by L-Can or siRNA inhibit osteogenic differentiation in the BMSCs. These findings suggest that a positively-charged surface with tertiary amines induces osteogenesis of BMSCs via the surface charge/iNOS signaling pathway in addition to elevated ECM protein adhesion. Therefore, creating a positively-charged surface with tertiary amines is a promising approach to promote osseointegration with bone tissues. PMID:25791957

Functionalization of polymer surfaces by medium frequency non-thermal plasma

NASA Astrophysics Data System (ADS)

Felix, T.; Trigueiro, J. S.; Bundaleski, N.; Teodoro, O. M. N. D.; Sério, S.; Debacher, N. A.

2018-01-01

This work addresses the surface modification of different polymers by argon dielectric barrier discharge, using bromoform vapours. Atomic Force Microscopy and Scanning Electron Microscopy showed that plasma etching occurs in stages and may be related to the reach of the species generated and obviously the gap between the electrodes. In addition, the stages of flatten surface or homogeneity may be the result of the transient crosslinking promoted by the intense UV radiation generated by the non- thermal plasma. X-ray Photoelectron Spectroscopy analysis showed that bromine was inserted on the polymer surface as Csbnd Br bonds and as adsorbed HBr. The obtained results demonstrate that the highest degree of bromofunctionalization was achieved on polypropylene surface, which contains about 8,5% of Br. After its derivatization in ammonia, Br disappeared and about 6% of nitrogen in the form of amine group was incorporated at the surface. This result can be considered as a clear fingerprint of the Br substitution by the amine group, thus illustrating the efficiency of the proposed method for functionalization of polymer surfaces.

NAD+/NADH and/or CoQ/CoQH2 ratios from plasma membrane electron transport may determine ceramide and sphingosine-1-phosphate levels accompanying G1 arrest and apoptosis.

PubMed

De Luca, Thomas; Morré, Dorothy M; Zhao, Haiyun; Morré, D James

2005-01-01

To elucidate possible biochemical links between growth arrest from antiproliferative chemotherapeutic agents and apoptosis, our work has focused on agents (EGCg, capsaicin, cis platinum, adriamycin, anti-tumor sulfonylureas, phenoxodiol) that target tNOX. tNOX is a cancer-specific cell surface NADH oxidase (ECTO-NOX protein), that functions in cancer cells as the terminal oxidase for plasma membrane electron transport. When tNOX is active, coenzyme Q(10) (ubiquinone) of the plasma membrane is oxidized and NADH is oxidized at the cytosolic surface of the plasma membrane. However, when tNOX is inhibited and plasma membrane electron transport is diminished, both reduced coenzyme Q(10) (ubiquinol) and NADH would be expected to accumulate. To relate inhibition of plasma membrane redox to increased ceramide levels and arrest of cell proliferation in G(1) and apoptosis, we show that neutral sphingomyelinase, a major contributor to plasma membrane ceramide, is inhibited by reduced glutathione and ubiquinone. Ubiquinol is without effect or stimulates. In contrast, sphingosine kinase, which generates anti-apoptotic sphingosine-1-phosphate, is stimulated by ubiquinone but inhibited by ubiquinol and NADH. Thus, the quinone and pyridine nucleotide products of plasma membrane redox, ubiquinone and ubiquinol, as well as NAD(+) and NADH, may directly modulate in a reciprocal manner two key plasma membrane enzymes, sphingomyelinase and sphingosine kinase, potentially leading to G(1) arrest (increase in ceramide) and apoptosis (loss of sphingosine-1-phosphate). As such, the findings provide potential links between coenzyme Q(10)-mediated plasma membrane electron transport and the anticancer action of several clinically-relevant anticancer agents.

Phenylethylamine-induced generation of reactive oxygen species and ascorbate free radicals in tobacco suspension culture: mechanism for oxidative burst mediating Ca2+ influx.

PubMed

Kawano, T; Pinontoan, R; Uozumi, N; Morimitsu, Y; Miyake, C; Asada, K; Muto, S

2000-11-01

In the previous paper [Kawano et al. (2000a) Plant Cell Physiol. 41: 1251], we demonstrated that addition of phenylethylamine (PEA) and benzylamine can induce an immediate and transient burst of active oxygen species (AOS) in tobacco suspension culture. Detected AOS include H2O2, superoxide anion and hydroxyl radicals. Use of several inhibitors suggested the presence of monoamine oxidase-like H2O2-generating activity in the cellular soluble fraction. It was also suggested that peroxidase(s) or copper amine oxidase(s) are involved in the extracellular superoxide production as a consequence of H2O2 production. Since more than 85% of the PEA-dependent AOS generating activity was localized in the extracellular space (extracellular fluid + cell wall), extracellularly secreted enzymes, probably peroxidases, may largely contribute to the oxidative burst induced by PEA. The PEA-induced AOS generation was also observed in the horseradish peroxidase (HRP) reaction mixture, supporting the hypothesis that peroxidases catalyze the oxidation of PEA leading to AOS generation. In addition to AOS production, we observed that PEA induced an increase in monodehydroascorbate radicals (MDA) in the cell suspension culture and in HRP reaction mixture using electron spin resonance spectroscopy and the newly invented MDA reductase-coupled method. Here we report that MDA production is an indicator of peroxidase-mediated generation of PEA radical species in tobacco suspension culture.

Role of L-alanine for redox self-sufficient amination of alcohols.

PubMed

Klatte, Stephanie; Wendisch, Volker F

2015-01-23

In white biotechnology biocatalysis represents a key technology for chemical functionalization of non-natural compounds. The plasmid-born overproduction of an alcohol dehydrogenase, an L-alanine-dependent transaminase and an alanine dehydrogenase allows for redox self-sufficient amination of alcohols in whole cell biotransformation. Here, conditions to optimize the whole cell biocatalyst presented in (Bioorg Med Chem 22:5578-5585, 2014), and the role of L-alanine for efficient amine functionalization of 1,10-decanediol to 1,10-diaminodecane were analyzed. The enzymes of the cascade for amine functionalization of alcohols were characterized in vitro to find optimal conditions for an efficient process. Transaminase from Chromobacterium violaceum, TaCv, showed three-fold higher catalytic efficiency than transaminase from Vibrio fluvialis, TaVf, and improved production at 37°C. At 42°C, TaCv was more active, which matched thermostable alcohol dehydrogenase and alanine dehydrogenase and improved the 1,10-diaminodecane production rate four-fold. To study the role of L-alanine in the whole cell biotransformation, the L-alanine concentration was varied and 1,10.diaminodecane formation tested with constant 10 mM 1,10- decanediol and 100 mM NH4Cl. Only 5.6% diamine product were observed without added L-alanine. L-alanine concentrations equimolar to that of the alcohol enabled for 94% product formation but higher L-alanine concentrations allowed for 100% product formation. L-alanine was consumed by the E. coli biocatalyst, presumably due to pyruvate catabolism since up to 16 mM acetate accumulated. Biotransformation employing E. coli strain YYC202/pTrc99a-ald-adh-ta Cv, which is unable to catabolize pyruvate, resulted in conversion with a selectivity of 42 mol-%. Biotransformation with E. coli strains only lacking pyruvate oxidase PoxB showed similar reduced amination of 1,10-decanediol indicating that oxidative decarboxylation of pyruvate to acetate by PoxB is primarily responsible for pyruvate catabolism during redox self-sufficient amination of alcohols using this whole cell biocatalyst. The replacement of the transaminase TaVf by TaCv, which showed higher activity at 42°C, in the artificial operon ald-adh-ta improved amination of alcohols in whole cell biotransformation. The addition of L-alanine, which was consumed by E. coli via pyruvate catabolism, was required for 100% product formation possibly by providing maintenance energy. Metabolic engineering revealed that pyruvate catabolism occurred primarily via oxidative decarboxylation to acetate by PoxB under the chosen biotranformation conditions.

Comparison of the effects of moclobemide and selegiline on tyramine-evoked mydriasis in man

PubMed Central

Bitsios, P; Langley, R W; Tavernor, S; Pyykkö, K; Scheinin, M; Szabadi, E; Bradshaw, C M

1998-01-01

Aims To examine the feasibility of using the human iris in vivo for the assessment of the interaction between tyramine and monoamine oxidase (MAO) inhibitors. To examine the relative roles of the two forms of MAO in terminating the response to sympathomimetic amines in the iris, by comparing the effects of single oral doses of moclobemide, a selective MAO-A inhibitor, and selegiline, a selective MAO-B inhibitor, on mydriatic responses to tyramine. Methods Twelve healthy male volunteers participated in three monthly sessions, each associated with ingestion of one capsule (moclobemide 450 mg, selegiline 10 mg, or placebo), according to a double-blind, balanced, cross-over design. Tyramine hydrochloride eye-drops (75 mm, 2×10 μl) were instilled three times in the left conjuctival sac at 40 min intervals. Pupil diameter was monitored with a binocular infrared television pupillometer before and for 4.5 h after ingestion of the capsule. The pupillary response to tyramine was expressed as the area under the pupil diameter×time curve (arbitrary units). A blood sample was taken before and 2 h after ingestion of the capsule, for the assay of platelet MAO-B activity, and plasma 3,4-dihydroxyphenylglycol (DHPG) concentration, an index of MAO-A activity. Platelet MAO activity was assayed radiochemically, using [14C]-phenylethylamine as substrate, and plasma DHPG by high performance liquid chromatography (h.p.l.c.). The results were analysed using analysis of variance with repeated measures, followed by Bonferroni's corrected t-test, using a significance criterion of P < 0.05. Results Both moclobemide and selegiline, compared with placebo, caused significant miosis in the right (untreated) eye. The changes in pupil diameter (mm±s.e. mean) from the pretreatment measurement were: placebo −0.09±0.07, moclobemide −0.52±0.09, selegiline −0.26±0.1. The mydriatic response to tyramine was potentiated by moclobemide, compared with the response recorded in the presence of placebo. The responses to tyramine (arbitrary units±s.e. mean) were: placebo 77.08±11.65, moclobemide 140.25±18.9, selegiline 72.75±12.35. Both moclobemide and selegiline significantly reduced platelet MAO activity, compared with placebo. The changes in platelet MAO activity (nmol h−1 mg−1 protein±s.e. mean) from the pretreatment level were: placebo 0.5±0.62, moclobemide −6.7±0.66, selegiline −17.7±0.87. Moclobemide significantly reduced plasma DHPG concentration, compared with placebo. The changes in plasma DHPG concentration (nmol l−1±s.e. mean) from the pretreatment level were: placebo −0.01±0.24, moclobemide −4.98±0.32, selegiline −0.51±0.26. Conclusions The potentiation of tyramine-evoked mydriasis by moclobemide is likely to reflect the inhibition of MAO-A activity in the iris, consistent with the activity of this enzyme in sympathetic nerve terminals. The lack of effect of selegiline on tyramine-evoked mydriasis argues against a role of MAO-B in terminating the effects of sympathomimetic amines in the iris. The effects of the two drugs on platelet MAO activity and plasma DHPG concentration are in agreement with previous reports and consistent with the relative selectivity of moclobemide for MAO-A and of selegiline for MAO-B. The miosis caused by the two MAO inhibitors is likely to be due to a central sympatholytic action of the drugs. PMID:9663810

Multifunctional Triblock Nanocarrier (PAMAM-PEG-PLL) for the Efficient Intracellular siRNA Delivery and Gene Silencing

PubMed Central

2011-01-01

A novel triblock poly(amido amine)-poly(ethylene glycol)-poly-l-lysine (PAMAM-PEG-PLL) nanocarrier was designed, synthesized, and evaluated for the delivery of siRNA. The design of the nanocarrier is unique and provides a solution to most of the common problems associated with the delivery and therapeutic applications of siRNA. Every component in the triblock nanocarrier plays a significant role and performs multiple functions: (1) tertiary amine groups in the PAMAM dendrimer work as a proton sponge and play a vital role in the endosomal escape and cytoplasmic delivery of siRNA; (2) PEG, a linker connecting PLL and PAMAM dendrimers renders nuclease stability and protects siRNA in human plasma; (3) PLL provides primary amines to form polyplexes with siRNA through electrostatic interaction and also acts as penetration enhancer; and (4) conjugation to PEG and PAMAM reduced toxicity of PLL and the entire triblock nanocarrier PAMAM-PEG-PLL. The data obtained show that the polyplexes resulted from the conjugation of siRNA, and the proposed nanocarriers were effectively taken up by cancer cells and induced the knock down of the target BCL2 gene. In addition, triblock nanocarrier/siRNA polyplexes showed excellent stability in human plasma. PMID:21322531

Recent advances in Parkinson's disease therapy: use of monoamine oxidase inhibitors.

PubMed

Henchcliffe, Claire; Schumacher, H Christian; Burgut, F Tuna

2005-11-01

Monoamine oxidase inhibitors inhibit dopamine metabolism and are therefore effective in treating Parkinson's disease, a condition associated with progressive striatal dopamine deficiency secondary to degeneration of dopaminergic neurons in the substantia nigra. Selegiline is currently the most widely used monoamine oxidase-B inhibitor for Parkinson's disease, but has a low and variable bioavailability, and is metabolized to L-methamphetamine and L-amphetamine that carry a risk for potential neurotoxicity. There are two new approaches that circumvent these potential disadvantages. First, selegiline orally disintegrating tablets provide a novel delivery form of selegiline, avoiding first pass metabolism by rapid absorption through the oral mucosa, thus leading to significantly lower plasma concentrations of L-metamphetamine and L-amphetamine. Selegiline orally disintegrating tablets prove to be clinically effective and safe in patients with moderately advanced Parkinson's disease. Second, rasagiline is a new monoamine oxidase inhibitor, without known neurotoxic metabolites. In large clinical trials, rasagiline proves effective as monotherapy in early Parkinson's disease, as well as adjunctive therapy to levodopa in advanced disease. Clinical data suggest, in addition, a disease-modifying effect of rasagiline that may correlate with neuroprotective activity of monoamine oxidase-B inhibitors in animal models of Parkinson's disease.

Purine nucleoside phosphorylase and xanthine oxidase activities in erythrocytes and plasma from marine, semiaquatic and terrestrial mammals.

PubMed

López-Cruz, Roberto I; Pérez-Milicua, Myrna Barjau; Crocker, Daniel E; Gaxiola-Robles, Ramón; Bernal-Vertiz, Jaime A; de la Rosa, Alejandro; Vázquez-Medina, José P; Zenteno-Savín, Tania

2014-05-01

Purine nucleoside phosphorylase (PNP) and xanthine oxidase (XO) are key enzymes involved in the purine salvage pathway. PNP metabolizes purine bases to synthetize purine nucleotides whereas XO catalyzes the oxidation of purines to uric acid. In humans, PNP activity is reported to be high in erythrocytes and XO activity to be low in plasma; however, XO activity increases after ischemic events. XO activity in plasma of northern elephant seals has been reported during prolonged fasting and rest and voluntary associated apneas. The objective of this study was to analyze circulating PNP and XO activities in marine mammals adapted to tolerate repeated cycles of ischemia/reperfusion associated with diving (bottlenose dolphin, northern elephant seal) in comparison with semiaquatic (river otter) and terrestrial mammals (human, pig). PNP activities in plasma and erythrocytes, as well as XO activity in plasma, from all species were quantified by spectrophotometry. No clear relationship in circulating PNP or XO activity could be established between marine, semiaquatic and terrestrial mammals. Erythrocytes from bottlenose dolphins and humans are highly permeable to nucleosides and glucose, intraerythrocyte PNP activity may be related to a release of purine nucleotides from the liver. High-energy costs will probably mean a higher ATP degradation rate in river otters, as compared to northern elephant seals or dolphins. Lower erythrocyte PNP activity and elevated plasma XO activity in northern elephant seal could be associated with fasting and/or sleep- and dive-associated apneas. Copyright © 2014 Elsevier Inc. All rights reserved.

The plasma membrane-associated NADH oxidase (ECTO-NOX) of mouse skin responds to blue light

NASA Technical Reports Server (NTRS)

Morre, D. James; Morre, Dorothy M.

2003-01-01

NADH oxidases of the external plasma membrane surface (ECTO-NOX proteins) are characterized by oscillations in activity with a regular period length of 24 min. Explants of mouse skin exhibit the oscillatory activity as estimated from the decrease in A(340) suggesting that individual ECTO-NOX molecules must somehow be induced to function synchronously. Transfer of explants of mouse skin from darkness to blue light (495 nm, 2 min, 50 micromol m(-1) s(-1)) resulted in initiation of a new activity maximum (entrainment) with a midpoint 36 min after light exposure followed by maxima every 24 min thereafter. Addition of melatonin resulted in a new maximum 24 min after melatonin addition. The findings suggest that the ECTO-NOX proteins play a central role in the entrainment of the biological clock both by light and by melatonin.

Root respiratory burst oxidase homologue-dependent H2O2 production confers salt tolerance on a grafted cucumber by controlling Na+ exclusion and stomatal closure

PubMed Central

Niu, Mengliang; Huang, Yuan; Sun, Shitao; Sun, Jingyu; Cao, Haishun; Shabala, Sergey

2018-01-01

Abstract Plant salt tolerance can be improved by grafting onto salt-tolerant rootstocks. However, the underlying signaling mechanisms behind this phenomenon remain largely unknown. To address this issue, we used a range of physiological and molecular techniques to study responses of self-grafted and pumpkin-grafted cucumber plants exposed to 75 mM NaCl stress. Pumpkin grafting significantly increased the salt tolerance of cucumber plants, as revealed by higher plant dry weight, chlorophyll content and photochemical efficiency (Fv/Fm), and lower leaf Na+ content. Salinity stress resulted in a sharp increase in H2O2 production, reaching a peak 3 h after salt treatment in the pumpkin-grafted cucumber. This enhancement was accompanied by elevated relative expression of respiratory burst oxidase homologue (RBOH) genes RbohD and RbohF and a higher NADPH oxidase activity. However, this increase was much delayed in the self-grafted plants, and the difference between the two grafting combinations disappeared after 24 h. The decreased leaf Na+ content of pumpkin-grafted plants was achieved by higher Na+ exclusion in roots, which was driven by the Na+/H+ antiporter energized by the plasma membrane H+-ATPase, as evidenced by the higher plasma membrane H+-ATPase activity and higher transcript levels for PMA and SOS1. In addition, early stomatal closure was also observed in the pumpkin-grafted cucumber plants, reducing water loss and maintaining the plant’s hydration status. When pumpkin-grafted plants were pretreated with an NADPH oxidase inhibitor, diphenylene iodonium (DPI), the H2O2 level decreased significantly, to the level found in self-grafted plants, resulting in the loss of the salt tolerance. Inhibition of the NADPH oxidase-mediated H2O2 signaling in the root also abolished a rapid stomatal closure in the pumpkin-grafted plants. We concluded that the pumpkin-grafted cucumber plants increase their salt tolerance via a mechanism involving the root-sourced respiratory burst oxidase homologue-dependent H2O2 production, which enhances Na+ exclusion from the root and promotes an early stomatal closure. PMID:29145593

Development and Experimental Verification of Surface Effects in a Fluidic Model

DTIC Science & Technology

2006-01-01

FROM A HE PLASMA INSIDE A POLYSTYRENE MICROCHANNEL. 43 FIGURE 30: THE EMISSION SPECTRA FROM A MIXED HEXAFLUOROETHYLENE/HE PLASMA INSIDE THE...MICROCHANNEL 47 FIGURE 35: THE ADSORPTION OF GLUCOSE OXIDASE TO DIFFERENT POLYMER SURFACES WAS SHOWN TO HAVE A SIGNIFICANT EFFECT ON ELECTROOSMOTIC FLOW...approach involves neglecting non-ideal (convective-diffusive) effects 5 by assuming well- mixed protein in contact with an idealized surface. Coupled

Macrophage NADPH oxidase flavocytochrome B localizes to the plasma membrane and Rab11-positive recycling endosomes.

PubMed

Casbon, Amy-Jo; Allen, Lee-Ann H; Dunn, Kenneth W; Dinauer, Mary C

2009-02-15

Flavocytochrome b(558), the catalytic core of the phagocytic NADPH oxidase, mediates the transfer of electrons from NADPH to molecular oxygen to generate superoxide for host defense. Flavocytochrome b is a membrane heterodimer consisting of a large subunit gp91(phox) (NOX2) and a smaller subunit, p22(phox). Although in neutrophils flavocytochrome b has been shown to localize to the plasma membrane and specific granules, little is known about its distribution in macrophages. Using immunofluorescent staining and live cell imaging of fluorescently tagged gp91(phox) and p22(phox), we demonstrate in a Chinese hamster ovary cell model system and in RAW 264.7 and primary murine bone marrow-derived macrophages that flavocytochrome b is found in the Rab11-positive recycling endocytic compartment, as well as in Rab5-positive early endosomes and plasma membrane. Additionally, we show that unassembled p22(phox) and gp91(phox) subunits localize to the endoplasmic reticulum, which redistribute to the cell surface and endosomal compartments following heterodimer formation. These studies show for the first time that flavocytochrome b localizes to intracellular compartments in macrophages that recycle to the plasma membrane, which may act as a reservoir to deliver flavocytochrome b to the cell surface and phagosome membranes.

Activity of xanthine oxidase in plasma correlates with indices of insulin resistance and liver dysfunction in Japanese patients with type 2 diabetes mellitus and metabolic syndrome: A pilot exploratory study.

PubMed

Sunagawa, Sumito; Shirakura, Takashi; Hokama, Noboru; Kozuka, Chisayo; Yonamine, Masato; Namba, Toyotaka; Morishima, Satoko; Nakachi, Sawako; Nishi, Yukiko; Ikema, Tomomi; Okamoto, Shiki; Matsui, Chieko; Hase, Naoki; Tamura, Mizuho; Shimabukuro, Michio; Masuzaki, Hiroaki

2018-06-03

There is a controversy whether hyperuricemia is an independent risk for cardiometabolic diseases. Serum level of uric acid is affected by a wide variety of factors involved in its production and excretion. On the other hand, evidence has accumulated that locally and systemically activated xanthine oxidase (XO), a rate limiting enzyme for production of uric acid, is linked to metabolic derangement in humans and rodents. We therefore explored the clinical implication of plasma XO activity in patients with type 2 diabetes mellitus (T2DM) and metabolic syndrome (MetS). We enrolled 60 patients with T2DM and MetS. MetS was defined according to the 2005 International Diabetes Federation guidelines. Plasma XO activity was measured by highly sensitive fluorometric assay measuring the conversion of pterin to isoxanthopterin, and explored associations between the value of plasma XO activity and metabolic parameters. Value of plasma XO activity was correlated with indices of insulin resistance and level of circulating liver transaminases. On the other hand, level of serum uric acid was not correlated with indices of insulin resistance. The value of plasma XO activity was not correlated with serum uric acid level. Plasma XO activity correlates with indices of insulin resistance and liver dysfunction in Japanese patients with T2DM and MetS. Through assessing the plasma XO activity, patients demonstrating normal level of serum uric acid with higher activity of XO can be screened, thereby possibly providing a clue to uncover metabolic risks in T2DM and MetS. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Immobilization of glucose oxidase using CoFe2O4/SiO2 nanoparticles as carrier

NASA Astrophysics Data System (ADS)

Wang, Hai; Huang, Jun; Wang, Chao; Li, Dapeng; Ding, Liyun; Han, Yun

2011-04-01

Aminated-CoFe2O4/SiO2 magnetic nanoparticles (NPs) were prepared from primary silica particles using modified StÖber method. Glucose oxidase (GOD) was immobilized on CoFe2O4/SiO2 NPs via cross-linking with glutaraldehyde (GA). The optimal immobilization condition was achieved with 1% (v/v) GA, cross-linking time of 3 h, solution pH of 7.0 and 0.4 mg GOD (in 3.0 mg carrier). The immobilized GOD showed maximal catalytic activity at pH 6.5 and 40 °C. After immobilization, the GOD exhibited improved thermal, storage and operation stability. The immobilized GOD still maintained 80% of its initial activity after the incubation at 50 °C for 25 min, whereas free enzyme had only 20% of initial activity after the same incubation. After kept at 4 °C for 28 days, the immobilized and free enzyme retained 87% and 40% of initial activity, respectively. The immobilized GOD maintained approximately 57% of initial activity after reused 7 times. The KM (Michaelis-Menten constant) values for immobilized GOD and free GOD were 14.6 mM and 27.1 mM, respectively.

Biochemical effects and growth inhibition in MCF-7 cells caused by novel sulphonamido oxa-polyamine derivatives.

PubMed

Pavlov, V; Lin, P Kong Thoo; Rodilla, V

2002-04-01

The novel polyamine derivatives sulphonamido oxa-spermine (oxa-Spm) and sulphonamido oxa-spermidine (oxa-Spd) exhibited rapid cytotoxic action towards MCF-7 human breast cancer cells with IC50 values of 4.35 and 6.47 pM, respectively, after 24-h drug exposure. Neither compound is a substrate of serum amine oxidase. Both oxa-Spm and oxa-Spd caused cell shrinkage, as determined by phase-contrast microscopy. After incubation with 10 microM of either compound for 8 h, the cells underwent chromatin condensation and nuclear fragmentation. However, no clear DNA ladder was obtained by electrophoresis. The sulphonamido oxa-polyamine derivatives and especially oxa-Spd enhanced the activity of polyamine oxidase (PAO), an enzyme capable of oxidising N1-acetylated spermine and spermidine to spermidine and putrescine, respectively, generating cytotoxic H2O2 and 3-acetamidopropanal as by-products. The intracellular polyamine content was only marginally reduced in response to drug treatment. In conclusion, our data show that these novel sulphonamido oxa-polyamine derivatives possess high cytotoxic activity against MCF-7 cells and indicate that induction of PAO may mediate their cytotoxicity via apoptosis.

Synthesis and evaluation of quinazoline amino acid derivatives as mono amine oxidase (MAO) inhibitors.

PubMed

Khattab, Sherine Nabil; Haiba, Nesreen Saied; Asal, Ahmed Mosaad; Bekhit, Adnan A; Amer, Adel; Abdel-Rahman, Hamdy M; El-Faham, Ayman

2015-07-01

A series of quinazolinone amino acid ester and quinazolinone amino acid hydrazides were prepared under microwave irradiation as well as conventional condition. The microwave irradiation afforded the product in less reaction time, higher yield and purity. The structures of the synthesized compounds were confirmed by IR, NMR, and elemental analysis. The new synthesized compounds were studied for their monoamine oxidase inhibitory activity. They showed more selective inhibitory activity toward MAO-A than MAO-B. Compounds 7, 10, and 15 showed MAO-A inhibition activity (IC50=3.6×10(-9), 2.8×10(-9), 2.1×10(-9) M, respectively) comparable to that of the standard clorgyline (IC50=2.9×10(-9)M). 2-(2-(Benzo[d][1,3]dioxol-5-yl)-4-oxo-1,2-dihydroquinazolin-3(4H)-yl)acetohydrazide 15 showed selective MAO-A inhibition activity (SI=39524) superior to that of the standard clorgyline (SI=33793). The acute toxicity of the synthesized compounds was determined. In addition, computer-assisted simulated docking experiments were performed to rationalize the biological activity. Copyright © 2015 Elsevier Ltd. All rights reserved.

Congruence between PM H+-ATPase and NADPH oxidase during root growth: a necessary probability.

PubMed

Majumdar, Arkajo; Kar, Rup Kumar

2018-07-01

Plasma membrane (PM) H + -ATPase and NADPH oxidase (NOX) are two key enzymes responsible for cell wall relaxation during elongation growth through apoplastic acidification and production of ˙OH radical via O 2 ˙ - , respectively. Our experiments revealed a putative feed-forward loop between these enzymes in growing roots of Vigna radiata (L.) Wilczek seedlings. Thus, NOX activity was found to be dependent on proton gradient generated across PM by H + -ATPase as evident from pharmacological experiments using carbonyl cyanide m-chlorophenylhydrazone (CCCP; protonophore) and sodium ortho-vanadate (PM H + -ATPase inhibitor). Conversely, H + -ATPase activity retarded in response to different ROS scavengers [CuCl 2 , N, N' -dimethylthiourea (DMTU) and catalase] and NOX inhibitors [ZnCl 2 and diphenyleneiodonium (DPI)], while H 2 O 2 promoted PM H + -ATPase activity at lower concentrations. Repressing effects of Ca +2 antagonists (La +3 and EGTA) on the activity of both the enzymes indicate its possible mediation. Since, unlike animal NOX, the plant versions do not possess proton channel activity, harmonized functioning of PM H + -ATPase and NOX appears to be justified. Plasma membrane NADPH oxidase and H + -ATPase are functionally synchronized and they work cooperatively to maintain the membrane electrical balance while mediating plant cell growth through wall relaxation.

Mechanistic and structural analyses of the roles of active site residues in yeast polyamine oxidase Fms1: characterization of the N195A and D94N enzymes.

PubMed

Adachi, Mariya S; Taylor, Alexander B; Hart, P John; Fitzpatrick, Paul F

2012-10-30

Flavoprotein Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine in the biosynthetic pathway for pantothenic acid. The same reaction is catalyzed by the mammalian polyamine and spermine oxidases. The active site of Fms1 contains three amino acid residues positioned to interact with the polyamine substrate, His67, Asn195, and Asp94. These three residues form a hydrogen-bonding triad with Asn195 being the central residue. Previous studies of the effects of mutating His67 are consistent with that residue being important both for interacting with the substrate and for maintaining the hydrogen bonds in the triad [Adachi, M. S., Taylor, A. B., Hart, P. J., and Fitzpatrick, P. F. (2012) Biochemistry 51, 4888-4897]. The N195A and D94N enzymes have now been characterized to evaluate their roles in catalysis. Both mutations primarily affect the reductive half-reaction. With N(1)-acetylspermine as the substrate, the rate constant for flavin reduction decreases ~450-fold for both mutations; the effects with spermine as the substrate are smaller, 20-40-fold. The k(cat)/K(amine)- and k(cat)-pH profiles with N(1)-acetylspermine are only slightly changed from the profiles for the wild-type enzyme, consistent with the pK(a) values arising from the amine substrate or product and not from active site residues. The structure of the N195A enzyme was determined at a resolution of 2.0 Å. The structure shows a molecule of tetraethylene glycol in the active site and establishes that the mutation has no effect on the protein structure. Overall, the results are consistent with the role of Asn195 and Asp94 being to properly position the polyamine substrate for oxidation.

Early cannabinoid exposure influences neuroendocrine and reproductive functions in male mice: I. Prenatal exposure.

PubMed

Dalterio, S; Steger, R; Mayfield, D; Bartke, A

1984-01-01

Maternal exposure to delta 9-tetrahydrocannabinol (THC), the major psychoactive constituent in marihuana, or to the non-psychoactive cannabinol (CBN) or cannabidiol (CBD) alters endocrine functions and concentrations of brain biogenic amines in their male offspring. Prenatal CBN exposure on day 18 of gestation resulted in decreased plasma FSH levels, testicular testosterone (T) concentrations, and seminal vesicles weights, but increased plasma levels of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) post-castration in adulthood. Prenatal exposure to THC significantly enhanced the responsiveness of the testes to intratesticular LH injection in vivo and tended to increase human chorionic gonadotropin (hCG)-stimulated T production by decapsulated testes in vitro. In the CBN-exposed mice, hCG-stimulated T production was enhanced, while CBD exposure had no effect. Prenatal THC exposure altered the negative feedback effects of exogenous gonadal steroids in castrated adults, with lower plasma T and FSH levels after 20 micrograms T than in castrated controls. In contrast, CBD-exposed mice had higher levels of LH in plasma post-castration. In CBN-exposed adults, two weeks post-castration the concentration of norepinephrine (NE) and dopamine (DA) in hypothalamus and remaining brain were reduced, while levels of serotonin (5-HT) and its metabolite, 5-HIAA, were elevated compared to that in castrated OIL-controls. Prenatal CBD-exposure also reduced NE and elevated 5-HT and 5-HIAA, but did not affect DA levels post-castration. Concentrations of brain biogenic amines were not influenced by prenatal THC exposure in the present study. A single prenatal exposure to psychoactive or non-psychoactive components of marihuana results in long term alterations in the function of the hypothalamo-pituitary-gonadal axis. Changes in the concentrations of brain biogenic amines may be related to these effects of prenatal cannabinoids on endocrine function in adult male mice.

[Research on the mechanism and regulation of overtraining-related the function of neutrophils by the inhibitor of NADPH oxidase and glutamine supplementation].

PubMed

Dong, Jing-Mei; Chen, Pei-Jie

2013-07-01

To investigate the method and mechanism for exercise-related immunosuppression via the inhibitor of NADPH oxidase diphenyleneiodonium(DPI) and glutamine supplementation and on the function of neutrophils after overtraining. Fifty male Wistar rats were randomly divided into five groups: a negative control group (C), an overtraining group (E), an overtraining + DPI intervention group (D), an overtraining+ glutamine supplementation group(G) and combined glutamine + DPI intervention group(DG). After 36 - 40 h from the last training, eight rats were randomly selected from each group, and blood was sampled from the orbital vein. ELISAs were used to measure serum cytokine levels and lipid peroxidation in blood plasma. Flow cytometry was used to measure neutrophil respiratory burst and phagocytosis. The activity of NADPH oxidase was assessed by chemiluminescence and the gene expression of gp91(phox) and p47(phox) of the NADPH-oxidase subunit was checked by Western blot. Compared with group C, the plasma concentrations of NO increased in group G, and the NO, cytokine-induced neutrophil chemoattractant (CINC) concentrations in group DG increased significantly. The respiratory burst and phagocytosis function of neutrophils were decreased in group E, but in group DG were increased when compared with those of group E. After overtraining the expression of gp91(phox) and p47(phox) was up regulated in group E. There were no significant changes in other groups except group DG, in which the expression of gp91(phox) was down regulated. Compared with group E, the expression of gp91(phox) and p47(phox) was up regulated in group D, group G and group DG. The activation of NADPH oxidase is responsible for the production of superoxide anions, which may be related to the decrease in neutrophil function after over training and is the mechanism of exercise-related immunosuppression. The DPI treatment combined glutamine supplementation can reverse the decrease neutrophils function after overtraining in vitro.

Exercise training protects against atherosclerotic risk factors through vascular NADPH oxidase, extracellular signal-regulated kinase 1/2 and stress-activated protein kinase/c-Jun N-terminal kinase downregulation in obese rats.

PubMed

Touati, Sabeur; Montezano, Augusto C I; Meziri, Fayçal; Riva, Catherine; Touyz, Rhian M; Laurant, Pascal

2015-02-01

Exercise training reverses atherosclerotic risk factors associated with metabolic syndrome and obesity. The aim of the present study was to determine the molecular anti-inflammatory, anti-oxidative and anti-atherogenic effects in aorta from rats with high-fat diet-induced obesity. Male Sprague-Dawley rats were placed on a high-fat (HFD) or control (CD) diet for 12 weeks. The HFD rats were then divided into four groups: (i) sedentary HFD-fed rats (HFD-S); (ii) exercise trained (motor treadmill 5 days/week, 60 min/day, 12 weeks) HFD-fed rats (HFD-Ex); (iii) modified diet (HFD to CD) sedentary rats (HF/CD-S); and (iv) an exercise-trained modified diet group (HF/CD-Ex). Tissue levels of NADPH oxidase (activity and expression), NADPH oxidase (Nox) 1, Nox2, Nox4, p47(phox) , superoxide dismutase (SOD)-1, angiotensin AT1 and AT2 receptors, phosphorylated mitogen-activated protein kinase (MAPK; extracellular signal-regulated kinase (ERK) 1/2, stress-activated protein kinase (SAPK)/c-Jun N-terminal kinase (JNK)) and vascular cell adhesion molecule-1 (VCAM-1) were determined in the aorta. Plasma cytokines (tumour necrosis factor (TNF)-α and interleukin (IL)-6) levels were also measured. Obesity was accompanied by increases in NADPH oxidase activity, p47(phox) translocation, Nox4 and VCAM-1 protein expression, MAPK (ERK1/2, SAPK/JNK) phosphorylation and plasma TNF-α and IL-6 levels. Exercise training and switching from the HFD to CD reversed almost all these molecular changes. In addition, training increased aortic SOD-1 protein expression and decreased ERK1/2 phosphorylation. These findings suggest that protective effects of exercise training on atherosclerotic risk factors induced by obesity are associated with downregulation of NADPH oxidase, ERK1/2 and SAPK/JNK activity and increased SOD-1 expression. © 2014 Wiley Publishing Asia Pty Ltd.

The design, synthesis and structure-activity relationships associated with C28 amine-based betulinic acid derivatives as inhibitors of HIV-1 maturation.

PubMed

Chen, Yan; Sit, Sing-Yuen; Chen, Jie; Swidorski, Jacob J; Liu, Zheng; Sin, Ny; Venables, Brian L; Parker, Dawn D; Nowicka-Sans, Beata; Lin, Zeyu; Li, Zhufang; Terry, Brian J; Protack, Tricia; Rahematpura, Sandhya; Hanumegowda, Umesh; Jenkins, Susan; Krystal, Mark; Dicker, Ira D; Meanwell, Nicholas A; Regueiro-Ren, Alicia

2018-05-15

The design and synthesis of a series of C28 amine-based betulinic acid derivatives as HIV-1 maturation inhibitors is described. This series represents a continuation of efforts following on from previous studies of C-3 benzoic acid-substituted betulinic acid derivatives as HIV-1 maturation inhibitors (MIs) that were explored in the context of C-28 amide substituents. Compared to the C-28 amide series, the C-28 amine derivatives exhibited further improvements in HIV-1 inhibitory activity toward polymorphisms in the Gag polyprotein as well as improved activity in the presence of human serum. However, plasma exposure of basic amines following oral administration to rats was generally low, leading to a focus on moderating the basicity of the amine moiety distal from the triterpene core. The thiomorpholine dioxide (TMD) 20 emerged from this study as a compound with the optimal antiviral activity and an acceptable pharmacokinetic profile in the C-28 amine series. Compared to the C-28 amide 3, 20 offers a 2- to 4-fold improvement in potency towards the screening viruses, exhibits low shifts in the EC 50 values toward the V370A and ΔV370 viruses in the presence of human serum or human serum albumin, and demonstrates improved potency towards the polymorphic T371A and V362I virus variants. Copyright © 2018 Elsevier Ltd. All rights reserved.

Metabolic Signature of Remote Ischemic Preconditioning Involving a Cocktail of Amino Acids and Biogenic Amines.

PubMed

Chao de la Barca, Juan Manuel; Bakhta, Oussama; Kalakech, Hussein; Simard, Gilles; Tamareille, Sophie; Catros, Véronique; Callebert, Jacques; Gadras, Cédric; Tessier, Lydie; Reynier, Pascal; Prunier, Fabrice; Mirebeau-Prunier, Delphine

2016-09-24

Remote ischemic preconditioning (RIPC) is an attractive therapeutic procedure for protecting the heart against ischemia/reperfusion injury. Despite evidence of humoral mediators transported through the circulation playing a critical role, their actual identities so far remain unknown. We sought to identify plasmatic RIPC-induced metabolites that may play a role. Rat plasma samples from RIPC and control groups were analyzed using a targeted metabolomic approach aimed at measuring 188 metabolites. Principal component analysis and orthogonal partial least-squares discriminant analysis were used to identify the metabolites that discriminated between groups. Plasma samples from 50 patients subjected to RIPC were secondarily explored to confirm the results obtained in rats. Finally, a combination of the metabolites that were significantly increased in both rat and human plasma was injected prior to myocardial ischemia/reperfusion in rats. In the rat samples, 124 molecules were accurately quantified. Six metabolites (ornithine, glycine, kynurenine, spermine, carnosine, and serotonin) were the most significant variables for marked differentiation between the RIPC and control groups. In human plasma, analysis confirmed ornithine decrease and kynurenine and glycine increase following RIPC. Injection of the glycine and kynurenine alone or in combination replicated the protective effects of RIPC seen in rats. We have hereby reported significant variations in a cocktail of amino acids and biogenic amines after remote ischemic preconditioning in both rat and human plasma. URL: http://www.clinicaltrials.gov. Unique identifier: NCT01390129. © 2016 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

Milestones and recent discoveries on cell death mediated by mitochondria and their interactions with biologically active amines.

PubMed

Grancara, Silvia; Ohkubo, Shinji; Artico, Marco; Ciccariello, Mauro; Manente, Sabrina; Bragadin, Marcantonio; Toninello, Antonio; Agostinelli, Enzo

2016-10-01

Mitochondria represent cell "powerhouses," being involved in energy transduction from the electrochemical gradient to ATP synthesis. The morphology of their cell types may change, according to various metabolic processes or osmotic pressure. A new morphology of the inner membrane and mitochondrial cristae, significantly different from the previous one, has been proposed for the inner membrane and mitochondrial cristae, based on the technique of electron tomography. Mitochondrial Ca(2+) transport (the transporter has been isolated) generates reactive oxygen species and induces the mitochondrial permeability transition of both inner and outer mitochondrial membranes, leading to induction of necrosis and apoptosis. In the mitochondria of several cell types (liver, kidney, and heart), mitochondrial oxidative stress is an essential step in the induction of cell death, although not in brain, in which the phenomenon is caused by a different mechanism. Mitochondrial permeability transition drives both apoptosis and necrosis, whereas mitochondrial outer membrane permeability is characteristic of apoptosis. Adenine nucleotide translocase remains the most important component involved in membrane permeability, with the opening of the transition pore, although other proteins, such as ATP synthase or phosphate carriers, have been proposed. Intrinsic cell death is triggered by the release from mitochondria of proteic factors, such as cytochrome c, apoptosis inducing factor, and Smac/DIABLO, with the activation of caspases upon mitochondrial permeability transition or mitochondrial outer membrane permeability induction. Mitochondrial permeability transition induces the permeability of the inner membrane in sites in contact with the outer membrane; mitochondrial outer membrane permeability forms channels on the outer membrane by means of various stimuli involving Bcl-2 family proteins. The biologically active amines, spermine, and agmatine, have specific functions on mitochondria which distinguish them from other amines. Enzymatic oxidative deamination of spermine by amine oxidases in tumor cells may produce reactive oxygen species, leading to transition pore opening and apoptosis. This process could be exploited as a new therapeutic strategy to combat cancer.

Identification of trace-amine-associated receptors (TAAR) in the rat aorta and their role in vasoconstriction by β-phenylethylamine.

PubMed

Fehler, Martina; Broadley, Kenneth J; Ford, William R; Kidd, Emma J

2010-10-01

Trace amines including tyramine and β-phenylethylamine (β-PEA) increase blood pressure and cause vasoconstriction which is attributed to indirect sympathomimetic actions. However, there is evidence that they may also have non-sympathomimetic mechanisms. This study examined whether β-PEA causes vasoconstriction of rat aorta by a sympathomimetic action or through the recently described trace-amine-associated receptors (TAAR). Concentration-response curves (CRCs) for β-PEA were constructed either cumulatively or non-cumulatively in rat isolated aortic rings. TAAR-1 and TAAR-4 protein expression was determined in rat aorta by Western blotting and TAAR-1 mRNA by reverse transcriptase polymerase chain reaction (RT-PCR). β-PEA caused concentration-related constriction of rat aorta. The contractions were unaffected by endothelium removal or the nitric oxide synthase inhibitor, N(ω)-nitro-L-arginine methyl ester (L-NAME, 100 μM) or the cyclooxygenase inhibitor, indomethacin (10 μM). Non-cumulative CRCs showed greater contractions and sensitivity to β-PEA than cumulative. The α(1)-adrenoceptor antagonist, prazosin, failed to inhibit either curve. The β-adrenoceptor antagonist, propranolol, the adrenergic neuronal transport inhibitor, cocaine, and the monoamine oxidase inhibitor, pargyline, also failed to alter the CRC. In the combined presence of prazosin, cocaine, pargyline, and the selective β(2)-adrenoceptor antagonist, ICI-118,551, the trace amine contractile potency order was tryptamine > β-PEA > octopamine > D: -amphetamine > tyramine. Western blotting and RT-PCR revealed the presence of TAAR-1 in rat aorta, but TAAR-4 was poorly expressed. Vasoconstriction of rat aorta by β-PEA appears not to be an indirect sympathomimetic action. The presence of TAAR-1 suggests that vasoconstriction may be via these receptors; however, the potency order differed from that reported for transfected cells expressing rat TAAR-1.

Reconstituted high-density lipoprotein suppresses leukocyte NADPH oxidase activation by disrupting lipid rafts.

PubMed

Peshavariya, Hitesh; Dusting, Gregory J; Di Bartolo, Belinda; Rye, Kerry-Anne; Barter, Philip J; Jiang, Fan

2009-08-01

Reconstituted discoidal high-density lipoprotein (rHDL) has potent vascular protective actions. Native HDL suppresses cellular generation of reactive oxygen species, whereas this antioxidant effect of rHDL is less clear. This study examined the effects of rHDL on NADPH oxidase, a major source of cellular superoxide generation, in both leukocytes and human umbilical vein endothelial cells. Superoxide was measured with lucigenin-enhanced chemiluminescence. Expression of NADPH oxidase sub-units was determined by real-time PCR. Pre-treatment of HL-60 cells with rHDL (10 and 25 microM) for 1 h significantly reduced phorbol 12-myristate 13-acetate-stimulated superoxide production. Treatment with rHDL for up to 24 h did not change the mRNA expression of NADPH oxidase sub-units. In HL-60 cells, depletion of cholesterol from the plasma membrane by methyl-beta-cyclodextrin mimicked the effect of rHDL, whereas cholesterol repletion blunted the effects of rHDL. Treatment with rHDL induced disruption of the lipid raft structures and blunted PMA-induced redistribution of p47phox into lipid rafts. In contrast, treatment of endothelial cells with rHDL for up to 18 h had no effect on either basal or tumour necrosis factor-alpha-stimulated NADPH oxidase activity, but markedly suppressed the cytokine-induced expression of proinflammatory adhesion molecules. The results suggest that rHDL inhibits NADPH oxidase activation in leukocytes, probably by interrupting the assembly of NADPH oxidase sub-units at the lipid rafts. This effect may contribute to the vascular protective actions of rHDL against inflammation-mediated oxidative damage.

Neurochemical binding profiles of novel indole and benzofuran MDMA analogues.

PubMed

Shimshoni, Jakob A; Winkler, Ilan; Golan, Ezekiel; Nutt, David

2017-01-01

3,4-Methylenedioxy-N-methylamphetamine (MDMA) has been shown to be effective in the treatment of post-traumatic stress disorder (PTSD) in numerous clinical trials. In the present study, we have characterized the neurochemical binding profiles of three MDMA-benzofuran analogues (1-(benzofuran-5-yl)-propan-2-amine, 5-APB; 1-(benzofuran-6-yl)-N-methylpropan-2-amine, 6-MAPB; 1-(benzofuran-5-yl)-N-methylpropan-2-amine, 5-MAPB) and one MDMA-indole analogue (1-(1H-indol-5-yl)-2-methylamino-propan-1-ol, 5-IT). These compounds were screened as potential second-generation anti-PTSD drugs, against a battery of human and non-human receptors, transporters, and enzymes, and their potencies as 5-HT 2 receptor agonist and monoamine uptake inhibitors determined. All MDMA analogues displayed high binding affinities for 5-HT 2a,b,c and NE α2 receptors, as well as significant 5-HT, DA, and NE uptake inhibition. 5-APB revealed significant agonist activity at the 5-HT 2a,b,c receptors, while 6-MAPB, 5-MAPB, and 5-IT exhibited significant agonist activity at the 5-HT 2c receptor. There was a lack of correlation between the results of functional uptake and the monoamine transporter binding assay. MDMA analogues emerged as potent and selective monoamine oxidase A inhibitors. Based on 6-MAPB favorable pharmacological profile, it was further subjected to IC 50 determination for monoamine transporters. Overall, all MDMA analogues displayed higher monoamine receptor/transporter binding affinities and agonist activity at the 5-HT 2a,c receptors as compared to MDMA.

Relative quantification of enantiomers of chiral amines by high-throughput LC-ESI-MS/MS using isotopic variants of light and heavy L-pyroglutamic acids as the derivatization reagents.

PubMed

Mochizuki, Toshiki; Taniguchi, Sayuri; Tsutsui, Haruhito; Min, Jun Zhe; Inoue, Koichi; Todoroki, Kenichiro; Toyo'oka, Toshimasa

2013-04-22

L-Pyroglutamic acid (L-PGA) was evaluated as a chiral labeling reagent for the enantioseparation of chiral amines in terms of separation efficiency by reversed-phase chromatography and detection sensitivity by ESI-MS/MS. Several amines and amino acid methyl esters were used as typical representatives of the chiral amines. Both enantiomers of the chiral amines were easily labeled with L-PGAS at room temperature for 60 min in the presence of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide and 1-hydroxy-1H-benzotriazole as the activation reagents. The resulting diastereomers were completely separated by reversed-phase chromatography using the small particle (1.7 μm) ODS column (Rs=1.6-6.8). A highly sensitive detection at a low-fmol level (1-4 fmol) was also obtained from the multiple reaction monitoring (MRM) chromatograms. Therefore, a high-throughput determination was achieved by the present UPLC-ESI-MS/MS method. An isotope labeling strategy using light and heavy L-PGAs for the differential analysis of chiral amines in different sample groups was also proposed in this paper. As a model study, the differential analysis of the R and S ratio of 1-phenylethylamine (PEA) was performed according to the proposed procedure using light and heavy reagents, i.e., L-PGA and L-PGA-d5. The R/S ratio of PEA, spiked at the different concentrations in rat plasma, was almost similar to the theoretical values. Consequently, the proposed strategy using light and heavy chiral labeling reagents seems to be applicable for the differential analysis of chiral amine enantiomers in different sample groups, such as healthy persons and disease patients. Copyright © 2013 Elsevier B.V. All rights reserved.

Exogenous polyamines elicit herbivore-induced volatiles in lima bean leaves: involvement of calcium, H2O2 and Jasmonic acid.

PubMed

Ozawa, Rika; Bertea, Cinzia M; Foti, Maria; Narayana, Ravishankar; Arimura, Gen-Ichiro; Muroi, Atsushi; Horiuchi, Jun-Ichiro; Nishioka, Takaaki; Maffei, Massimo E; Takabayashi, Junji

2009-12-01

We investigated the role of polyamines (PAs) in lima bean (Phaseolus lunatus) leaves on the production of herbivorous mite (Tetranychus urticae)-induced plant volatiles that attract carnivorous natural enemies of the herbivores. To do this, we focused on the effects of the exogenous PAs [cadaverine, putrescine, spermidine and spermine (Spm)] on the production of volatiles, H(2)O(2) and jasmonic acid (JA) and the levels of defensive genes, cytosolic calcium and reactive oxygen species (ROS). Among the tested PAs, Spm was the most active in inducing the production of volatile terpenoids known to be induced by T. urticae. An increase in JA levels was also found after Spm treatment, indicating that Spm induces the biosynthesis of JA, which has been shown elsewhere to regulate the production of some volatile terpenoids. Further, treatment with JA and Spm together resulted in greater volatile emission than that with JA alone. In a Y-tube olfactometer, leaves treated with Spm + JA attracted more predatory mites (Phytoseiulus persimilis) than those treated with JA alone. After treatment with Spm + JA, no effects were found on the enzyme activity of polyamine oxidase and copper amine oxidase. However, induction of calcium influx and ROS production, and increased enzyme activities and gene expression for NADPH oxidase complex, superoxide dismutase, catalase, ascorbate peroxidase, glutathione reductase and glutathione peroxidase were found after treatment with Spm + JA. These results indicate that Spm plays an important role in the production of T. urticae-induced lima bean leaf volatiles.

Mechanistic and Structural Analyses of the Role of His67 in the Yeast Polyamine Oxidase Fms1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adachi, Mariya S.; Taylor, Alexander B.; Hart, P. John

2012-07-25

The flavoprotein oxidase Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine and N1-acetylspermine to spermidine and 3-aminopropanal or N-acetyl-3-aminopropanal. Within the active site of Fms1, His67 is positioned to form hydrogen bonds with the polyamine substrate. This residue is also conserved in other polyamine oxidases. The catalytic properties of H67Q, H67N, and H67A Fms1 have been characterized to evaluate the role of this residue in catalysis. With both spermine and N1-acetylspermine as the amine substrate, the value of the first-order rate constant for flavin reduction decreases 2-3 orders of magnitude, with the H67Q mutation having the smallest effect andmore » H67N the largest. The k{sub cat}/K{sub O2} value changes very little upon mutation with N{sup 1}-acetylspermine as the amine substrate and decreases only an order of magnitude with spermine. The k{sub cat}/K{sub M}-pH profiles with N{sup 1}-acetylspermine are bell-shaped for all the mutants; the similarity to the profile of the wild-type enzyme rules out His67 as being responsible for either of the pK{sub a} values. The pH profiles for the rate constant for flavin reduction for all the mutant enzymes similarly show the same pK{sub a} as wild-type Fms1, about {approx}7.4; this pK{sub a} is assigned to the substrate N4. The k{sub cat}/K{sub O2}-pH profiles for wild-type Fms1 and the H67A enzyme both show a pK{sub a} of about {approx}6.9; this suggests His67 is not responsible for this pH behavior. With the H67Q, H67N, and H67A enzymes the k{sub cat} value decreases when a single residue is protonated, as is the case with the wild-type enzyme. The structure of H67Q Fms1 has been determined at a resolution of 2.4 {angstrom}. The structure shows that the mutation disrupts a hydrogen bond network in the active site, suggesting that His67 is important both for direct interactions with the substrate and to maintain the overall active site structure.« less

Antioxidative and antihypertensive effects of Welsh onion on rats fed with a high-fat high-sucrose diet.

PubMed

Yamamoto, Yukiko; Aoyama, Sakiko; Hamaguchi, Noriko; Rhi, Gyou-Sei

2005-07-01

The effects of Welsh onion on the development of hypertension and autoxidation were studied in 6-week-old male Sprague-Dawley rats. The rats were fed with a control diet or a high-fat high-sucrose (HFS) diet with or without 5% Welsh onion (green-leafy type or white-sheath type) for 4 weeks. The systolic blood pressure was elevated and the thiobarbituric acid reactive substances (TBARS) in plasma were increased in the rats fed with the HFS diet without Welsh onion. The rats fed with the HFS diet containing Welsh onion, especially the green-leafy type, had lower blood pressure. They also had a higher level of nitric oxide (NO) metabolites in both the urine and plasma, lower activity of NADH/NADPH oxidase in the aorta, and suppressed angiotensin II production. The effect of white Welsh onion on decreasing the blood pressure was not significant, although the effects on increasing NO metabolites in the urine and decreasing NADH oxidase activity in the aorta were significant. The TBARS value in the plasma was lowered in the rats fed with either green or white Welsh onion, but the in vitro radical scavenging and ferric reducing antioxidative activities were much higher with green Welsh onion than with the white type. These results suggest that the green-leafy Welsh onion, but not the white type, reduced superoxide generation by suppressing the angiotensine II production and then the NADH/NADPH oxidase activity, increasing the NO availability in the aorta, and consequently lowering the blood pressure in the rats fed with the HFS diet. The radical scavenging and reducing antioxidative activities of green Welsh onion may also be effective in decreasing superoxide.

Simultaneous Determination of Cyanide and Thiocyanate in Plasma by Chemical Ionization Gas Chromatography Mass-Spectrometry (CI-GC-MS)

DTIC Science & Technology

2012-09-04

3]. Once cyanide is introduced into cells, it inhibits cytochrome c oxidase, which subsequently causes cellular hypoxia, cytotoxic anoxia, and may...NAME OF RESPONSIBLE PERSON a. REPORT unclassified b. ABSTRACT unclassified c . THIS PAGE unclassified Standard Form 298 (Rev. 8-98) Prescribed by...Medical Center (Lackland Air Force Base, TX). Upon re- ceipt, the plasma was frozen and stored at −80 ° C until utilized for optimizing analytical

Vascular oxidative stress and endothelial dysfunction in patients with chronic heart failure: role of xanthine-oxidase and extracellular superoxide dismutase.

PubMed

Landmesser, Ulf; Spiekermann, Stephan; Dikalov, Sergey; Tatge, Helma; Wilke, Ragna; Kohler, Christoph; Harrison, David G; Hornig, Burkhard; Drexler, Helmut

2002-12-10

Impaired flow-dependent, endothelium-mediated vasodilation (FDD) in patients with chronic heart failure (CHF) results, at least in part, from accelerated degradation of nitric oxide by oxygen radicals. The mechanisms leading to increased vascular radical formation, however, remain unclear. Therefore, we determined endothelium-bound activities of extracellular superoxide dismutase (ecSOD), a major vascular antioxidant enzyme, and xanthine-oxidase, a potent radical producing enzyme, and their relation to FDD in patients with CHF. ecSOD and xanthine-oxidase activities, released from endothelium into plasma by heparin bolus injection, were determined in 14 patients with CHF and 10 control subjects. FDD of the radial artery was measured using high-resolution ultrasound and was assessed before and after administration of the antioxidant vitamin C (25 mg/min; IA). In patients with CHF, endothelium-bound ecSOD activity was substantially reduced (5.0+/-0.7 versus 14.4+/-2.6 U x mL(-1) x min(-1); P<0.01) and closely related to FDD (r=0.61). Endothelium-bound xanthine-oxidase activity was increased by >200% (38+/-10 versus 12+/-4 nmol O2*- x microL(-1); P<0.05) and inversely related to FDD (r=-0.35) in patients with CHF. In patients with low ecSOD and high xanthine-oxidase activity, a greater benefit of vitamin C on FDD was observed, ie, the portion of FDD inhibited by radicals correlated negatively with ecSOD (r=-0.71) but positively with xanthine-oxidase (r=0.75). These results demonstrate that both increased xanthine-oxidase and reduced ecSOD activity are closely associated with increased vascular oxidative stress in patients with CHF. This loss of vascular oxidative balance likely represents a novel mechanism contributing to endothelial dysfunction in CHF.

Nanocapillary Atmospheric Pressure Plasma Jet: A Tool for Ultrafine Maskless Surface Modification at Atmospheric Pressure.

PubMed

Motrescu, Iuliana; Nagatsu, Masaaki

2016-05-18

With respect to microsized surface functionalization techniques we proposed the use of a maskless, versatile, simple tool, represented by a nano- or microcapillary atmospheric pressure plasma jet for producing microsized controlled etching, chemical vapor deposition, and chemical modification patterns on polymeric surfaces. In this work we show the possibility of size-controlled surface amination, and we discuss it as a function of different processing parameters. Moreover, we prove the successful connection of labeled sugar chains on the functionalized microscale patterns, indicating the possibility to use ultrafine capillary atmospheric pressure plasma jets as versatile tools for biosensing, tissue engineering, and related biomedical applications.

Molecular aspects of monoamine oxidase B.

PubMed

Ramsay, Rona R

2016-08-01

Monoamine oxidases (MAO) influence the monoamine levels in brain by virtue of their role in neurotransmitter breakdown. MAO B is the predominant form in glial cells and in platelets. MAO B structure, function and kinetics are described as a background for the effect of alterations in its activity on behavior. The need to inhibit MAO B to combat decreased brain amines continues to drive the search for new drugs. Reversible and irreversible inhibitors are now designed using data-mining, computational screening, docking and molecular dynamics. Multi-target ligands designed to combat the elevated activity of MAO B in Alzheimer's and Parkinson's Diseases incorporate MAO inhibition (usually irreversible) as well as iron chelation, antioxidant or neuroprotective properties. The main focus of drug design is the catalytic activity of MAO, but the imidazoline I2 site in the entrance cavity of MAO B is also a pharmacological target. Endogenous regulation of MAO B expression is discussed briefly in light of new studies measuring mRNA, protein, or activity in healthy and degenerative samples, including the effect of DNA methylation on the expression. Overall, this review focuses on examples of recent research on the molecular aspects of the expression, activity, and inhibition of MAO B. Copyright © 2016 Elsevier Inc. All rights reserved.

Monoamine oxidase A mediates prostate tumorigenesis and cancer metastasis

PubMed Central

Wu, Jason Boyang; Shao, Chen; Li, Xiangyan; Li, Qinlong; Hu, Peizhen; Shi, Changhong; Li, Yang; Chen, Yi-Ting; Yin, Fei; Liao, Chun-Peng; Stiles, Bangyan L.; Zhau, Haiyen E.; Shih, Jean C.; Chung, Leland W.K.

2014-01-01

Tumors from patients with high-grade aggressive prostate cancer (PCa) exhibit increased expression of monoamine oxidase A (MAOA), a mitochondrial enzyme that degrades monoamine neurotransmitters and dietary amines. Despite the association between MAOA and aggressive PCa, it is unclear how MAOA promotes PCa progression. Here, we found that MAOA functions to induce epithelial-to-mesenchymal transition (EMT) and stabilize the transcription factor HIF1α, which mediates hypoxia through an elevation of ROS, thus enhancing growth, invasiveness, and metastasis of PCa cells. Knockdown and overexpression of MAOA in human PCa cell lines indicated that MAOA induces EMT through activation of VEGF and its coreceptor neuropilin-1. MAOA-dependent activation of neuropilin-1 promoted AKT/FOXO1/TWIST1 signaling, allowing FOXO1 binding at the TWIST1 promoter. Importantly, the MAOA-dependent HIF1α/VEGF-A/FOXO1/TWIST1 pathway was activated in high-grade PCa specimens, and knockdown of MAOA reduced or even eliminated prostate tumor growth and metastasis in PCa xenograft mouse models. Pharmacological inhibition of MAOA activity also reduced PCa xenograft growth in mice. Moreover, high MAOA expression in PCa tissues correlated with worse clinical outcomes in PCa patients. These findings collectively characterize the contribution of MAOA in PCa pathogenesis and suggest that MAOA has potential as a therapeutic target in PCa. PMID:24865426

Monoamine oxidase A mediates prostate tumorigenesis and cancer metastasis.

PubMed

Wu, Jason Boyang; Shao, Chen; Li, Xiangyan; Li, Qinlong; Hu, Peizhen; Shi, Changhong; Li, Yang; Chen, Yi-Ting; Yin, Fei; Liao, Chun-Peng; Stiles, Bangyan L; Zhau, Haiyen E; Shih, Jean C; Chung, Leland W K

2014-07-01

Tumors from patients with high-grade aggressive prostate cancer (PCa) exhibit increased expression of monoamine oxidase A (MAOA), a mitochondrial enzyme that degrades monoamine neurotransmitters and dietary amines. Despite the association between MAOA and aggressive PCa, it is unclear how MAOA promotes PCa progression. Here, we found that MAOA functions to induce epithelial-to-mesenchymal transition (EMT) and stabilize the transcription factor HIF1α, which mediates hypoxia through an elevation of ROS, thus enhancing growth, invasiveness, and metastasis of PCa cells. Knockdown and overexpression of MAOA in human PCa cell lines indicated that MAOA induces EMT through activation of VEGF and its coreceptor neuropilin-1. MAOA-dependent activation of neuropilin-1 promoted AKT/FOXO1/TWIST1 signaling, allowing FOXO1 binding at the TWIST1 promoter. Importantly, the MAOA-dependent HIF1α/VEGF-A/FOXO1/TWIST1 pathway was activated in high-grade PCa specimens, and knockdown of MAOA reduced or even eliminated prostate tumor growth and metastasis in PCa xenograft mouse models. Pharmacological inhibition of MAOA activity also reduced PCa xenograft growth in mice. Moreover, high MAOA expression in PCa tissues correlated with worse clinical outcomes in PCa patients. These findings collectively characterize the contribution of MAOA in PCa pathogenesis and suggest that MAOA has potential as a therapeutic target in PCa.

Synthesis of three novel fluorine-18 labeled analogues of L-deprenyl for positron emission tomography (PET) studies of monoamine oxidase B (MAO-B).

PubMed

Nag, Sangram; Lehmann, Lutz; Heinrich, Tobias; Thiele, Andrea; Kettschau, Georg; Nakao, Ryuji; Gulyás, Balázs; Halldin, Christer

2011-10-27

The aim in this project was to synthesize and to study fluorine-18 labeled analogues of l-deprenyl which bind selectively to the enzyme monoamine oxidase B (MAO-B). Three fluorinated l-deprenyl analogues have been generated in multistep organic syntheses. The most promising fluorine-18 compound N-[(2S)-1-[(18)F]fluoro-3-phenylpropan-2-yl]-N-methylprop-2-yn-1-amine (4c) was synthesized by a one-step fluorine-18 nucleophilic substitution reaction. Autoradiography on human brain tissue sections demonstrated specific binding for compound 4c to brain regions known to have a high content of MAO-B. In addition, the corresponding nonradioactive fluorine-19 compound (13) inhibited recombinant human MAO-B with an IC(50) of 170.5 ± 29 nM but did not inhibit recombinant human MAO-A (IC(50) > 2000 nM), demonstrating its specificity. Biodistribution of 4c in mice showed high initial brain uptake leveling at 5.2 ± 0.04%ID/g after 2 min post injection. In conclusion, compound 4c is a specific inhibitor of MAO-B with high initial brain uptake in mice and is, therefore, a candidate for further investigation in PET.

Design and synthesis of novel 2-(indol-5-yl)thiazole derivatives as xanthine oxidase inhibitors.

PubMed

Song, Jeong Uk; Choi, Sung Pil; Kim, Tae Hun; Jung, Cheol-Kyu; Lee, Joo-Youn; Jung, Sang-Hun; Kim, Geun Tae

2015-03-15

Xanthine oxidase (XO) inhibitors have been widely used for the treatment of gout. Indole rings are frequently used as active scaffold in designing inhibitors for enzymes. Herein, we describe the structure-activity relationship for novel xanthine oxidase inhibitors based on indole scaffold. A series of novel tri-substituted 2-(indol-5-yl)thiazole derivatives were synthesized, and their in vitro inhibitory activities against xanthine oxidase and in vivo efficacy lowering uric acid level in blood were measured. Among them, 2-(3-cyano-2-isopropylindol-5-yl)-4-methylthiazole-5-carboxylic acid exhibits the most potent XO inhibitory activity (IC50 value: 3.5nM) and the excellent plasma uric acid lowering activity. Study of structure activity relationship indicated that hydrophobic moiety (e.g., isopropyl) at 1-position and electron withdrawing group (e.g., CN) at 3-position of indole ring and small hydrophobic group (CH3) at 4-position of the thiazole ring enhanced the XO inhibitory activity. Hydrophobic substitution such as isopropyl at 1-position of the indole moiety without any substitution at 2-position has an essential role for enhancing bioavailability and therefore for high in vivo efficacy. Copyright © 2015 Elsevier Ltd. All rights reserved.

A semi-micromethod for determination of oxalate in human plasma.

PubMed

Porowski, Tadeusz; Gałasiński, Władysław

2003-01-01

An enzymatic semi-micromethod for oxalate determination in human plasma was elaborated. The principle of the method depends on the oxalate isolation from deproteinized plasma, following determination by the calorimetric oxalate oxidase-peroxidase-indamine system. This method protects against internal oxalate losses and excludes an interference of contaminations. Results, obtained by this method, were reliable and ideally suited for use as real normal values (less than or equal to 6 microM) of oxalate content in the plasma of healthy individuals. The elaborated method, which can assay plasma oxalate accurately in normal individuals as well as in hyperoxalemic conditions is superior to those previously used. The procedure of semi-micromethod does not require expensive equipments and apparatus: it is simple and easy to perform in every laboratory and takes little time.

Molecular dynamics, flexible docking, virtual screening, ADMET predictions, and molecular interaction field studies to design novel potential MAO-B inhibitors.

PubMed

Braun, Glaucia H; Jorge, Daniel M M; Ramos, Henrique P; Alves, Raquel M; da Silva, Vinicius B; Giuliatti, Silvana; Sampaio, Suley Vilela; Taft, Carlton A; Silva, Carlos H T P

2008-02-01

Monoamine oxidase is a flavoenzyme bound to the mitochondrial outer membranes of the cells, which is responsible for the oxidative deamination of neurotransmitter and dietary amines. It has two distinct isozymic forms, designated MAO-A and MAO-B, each displaying different substrate and inhibitor specificities. They are the well-known targets for antidepressant, Parkinson's disease, and neuroprotective drugs. Elucidation of the x-ray crystallographic structure of MAO-B has opened the way for the molecular modeling studies. In this work we have used molecular modeling, density functional theory with correlation, virtual screening, flexible docking, molecular dynamics, ADMET predictions, and molecular interaction field studies in order to design new molecules with potential higher selectivity and enzymatic inhibitory activity over MAO-B.

Polyamines control of cation transport across plant membranes: implications for ion homeostasis and abiotic stress signaling.

PubMed

Pottosin, Igor; Shabala, Sergey

2014-01-01

Polyamines are unique polycationic metabolites, controlling a variety of vital functions in plants, including growth and stress responses. Over the last two decades a bulk of data was accumulated providing explicit evidence that polyamines play an essential role in regulating plant membrane transport. The most straightforward example is a blockage of the two major vacuolar cation channels, namely slow (SV) and fast (FV) activating ones, by the micromolar concentrations of polyamines. This effect is direct and fully reversible, with a potency descending in a sequence Spm(4+) > Spd(3+) > Put(2+). On the contrary, effects of polyamines on the plasma membrane (PM) cation and K(+)-selective channels are hardly dependent on polyamine species, display a relatively low affinity, and are likely to be indirect. Polyamines also affect vacuolar and PM H(+) pumps and Ca(2+) pump of the PM. On the other hand, catabolization of polyamines generates H2O2 and other reactive oxygen species (ROS), including hydroxyl radicals. Export of polyamines to the apoplast and their oxidation there by available amine oxidases results in the induction of a novel ion conductance and confers Ca(2+) influx across the PM. This mechanism, initially established for plant responses to pathogen attack (including a hypersensitive response), has been recently shown to mediate plant responses to a variety of abiotic stresses. In this review we summarize the effects of polyamines and their catabolites on cation transport in plants and discuss the implications of these effects for ion homeostasis, signaling, and plant adaptive responses to environment.

Polyamines control of cation transport across plant membranes: implications for ion homeostasis and abiotic stress signaling

PubMed Central

Pottosin, Igor; Shabala, Sergey

2014-01-01

Polyamines are unique polycationic metabolites, controlling a variety of vital functions in plants, including growth and stress responses. Over the last two decades a bulk of data was accumulated providing explicit evidence that polyamines play an essential role in regulating plant membrane transport. The most straightforward example is a blockage of the two major vacuolar cation channels, namely slow (SV) and fast (FV) activating ones, by the micromolar concentrations of polyamines. This effect is direct and fully reversible, with a potency descending in a sequence Spm4+ > Spd3+ > Put2+. On the contrary, effects of polyamines on the plasma membrane (PM) cation and K+-selective channels are hardly dependent on polyamine species, display a relatively low affinity, and are likely to be indirect. Polyamines also affect vacuolar and PM H+ pumps and Ca2+ pump of the PM. On the other hand, catabolization of polyamines generates H2O2 and other reactive oxygen species (ROS), including hydroxyl radicals. Export of polyamines to the apoplast and their oxidation there by available amine oxidases results in the induction of a novel ion conductance and confers Ca2+ influx across the PM. This mechanism, initially established for plant responses to pathogen attack (including a hypersensitive response), has been recently shown to mediate plant responses to a variety of abiotic stresses. In this review we summarize the effects of polyamines and their catabolites on cation transport in plants and discuss the implications of these effects for ion homeostasis, signaling, and plant adaptive responses to environment. PMID:24795739

Mediterranean diets supplemented with virgin olive oil and nuts enhance plasmatic antioxidant capabilities and decrease xanthine oxidase activity in people with metabolic syndrome: The PREDIMED study.

PubMed

Sureda, Antoni; Bibiloni, Maria Del Mar; Martorell, Miquel; Buil-Cosiales, Pilar; Marti, Amelia; Pons, Antoni; Tur, Josep A; Martinez-Gonzalez, Miguel Ángel

2016-12-01

This study assessed plasmatic antioxidant capabilities and xanthine oxidase (XOX) activity in metabolic syndrome patients after 5 years intervention with Mediterranean diet (MeDiet) supplemented with extra-virgin olive oil or with nuts or with low-fat diet (the PREDIMED [PREvención con Dieta MEDiterránea] study). Seventy-five participants were randomly selected. Daily energy and nutrient intake were assessed with a validated 137-item food frequency questionnaire, and adherence to the MeDiet was assessed using a 14-item questionnaire. Catalase, superoxide dismutase (SOD), myeloperoxidase, XOX activities and protein levels, and protein carbonyl derivatives, nitrotyrosine, nitrite and nitrate levels were determined in overnight fasting venous blood samples. The plasma activity and protein levels of SOD and catalase were significantly higher and XOX activity was lower in MeDiet supplemented with extra-virgin olive oil and MeDiet supplemented with nuts than in the control group. Participants in both MeDiet groups showed higher plasma nitrate levels than in the control group. Adherence to the MeDiet showed a positive correlation with SOD and catalase plasma antioxidant activities. A MeDiet enriched with either virgin olive oil or nuts enhances the plasma antioxidant capabilities and decreases XOX activity in patients with the metabolic syndrome but we did not observe changes in myeloperoxidase or markers of oxidative damage. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Arctigenin reduces blood pressure by modulation of nitric oxide synthase and NADPH oxidase expression in spontaneously hypertensive rats.

PubMed

Liu, Ying; Wang, Guoyuan; Yang, Mingguang; Chen, Haining; zhao, Yan; Yang, Shucai; Sun, Changhao

2015-12-25

Arctigenin is a bioactive constituent from dried seeds of Arctium lappa L., which was traditionally used as medicine. Arctigenin exhibits various bioactivities, but its effects on blood pressure regulation are still not widely studied. In this study, we investigated antihypertensive effects of arctigenin by long-term treatment in spontaneously hypertensive rats (SHRs). Arctigenin (50 mg/kg) or vehicle was administered to SHRs or Wistar rats as negative control by oral gavage once a day for total 8 weeks. Nifedipine (3 mg/kg) was used as a positive drug control. After treatment, hemodynamic and physical parameters, vascular reactivity in aorta, the concentration of plasma arctigenin and serum thromboxane B2, NO release and vascular p-eNOS, p-Akt, caveolin-1 protein expression, and vascular superoxide anion generation and p47phox protein expression were detected and analyzed. The results showed that arctigenin significantly reduced systolic blood pressure and ameliorated endothelial dysfunction of SHRs. Arctigenin reduced the levels of thromboxane B2 in plasma and superoxide anion in thoracic aorta of SHRs. Furthermore, arctigenin increased the NO production by enhancing the phosphorylation of Akt and eNOS (Ser 1177), and inhibiting the expression of NADPH oxidase in thoracic aorta of SHRs. Our data suggested that antihypertensive mechanisms of arctigenin were associated with enhanced eNOS phosphorylation and decreased NADPH oxidase-mediated superoxide anion generation. Copyright © 2015 Elsevier Inc. All rights reserved.

Analysis of monoamine oxidase (MAO) enzymatic activity by high-performance liquid chromatography-diode array detection combined with an assay of oxidation with a peroxidase and its application to MAO inhibitors from foods and plants.

PubMed

Herraiz, Tomás; Flores, Andrea; Fernández, Lidia

2018-01-15

Monoamine oxidase (MAO) enzymes catalyze the oxidative deamination of biogenic amines and neurotransmitters and produce ammonia, aldehydes, and hydrogen peroxide which is involved in oxidative processes. Inhibitors of MAO-A and -B isozymes are useful as antidepressants and neuroprotectants. The assays of MAO usually measure amine oxidation products or hydrogen peroxide by spectrophotometric techniques. Those assays are often compromised by interfering compounds resulting in poor results. This research describes a new method that combines in the same assay the oxidative deamination of kynuramine to 4-hydroxyquinoline analyzed by HPLC-DAD with the oxidation of tetramethylbenzidine (TMB) (or Amplex Rex) by horseradish peroxidase (HRP) in presence of hydrogen peroxide. The new method was applied to study the inhibition of human MAO-A and -B by bioactive compounds including β-carboline alkaloids and flavonoids occurring in foods and plants. As determined by HPLC-DAD, β-carbolines, methylene blue, kaempferol and clorgyline inhibited MAO-A and methylene blue, 5-nitroindazole, norharman and deprenyl inhibited MAO-B, and all of them inhibited the oxidation of TMB in the same extent. The flavonoids catechin and cyanidin were not inhibitors of MAO by HPLC-DAD but highly inhibited the oxidation of TMB (or Amplex Red) by peroxidase whereas quercetin and resveratrol were moderate inhibitors of MAO-A by HPLC-DAD, but inhibited the peroxidase assay in a higher level. For some phenolic compounds, using the peroxidase-coupled assay to measure MAO activity led to mistaken results. The new method permits to discern between true inhibitors of MAO from those that are antioxidants and which interfere with peroxidase assays but do not inhibit MAO. For true inhibitors of MAO, inhibition as determined by HPLC-DAD correlated well with inhibition of the oxidation of TMB and this approach can be used to assess the in vitro antioxidant activity (less hydrogen peroxide production) resulting from MAO inhibition. Copyright © 2017 Elsevier B.V. All rights reserved.

Apple fruit copper amine oxidase isoforms: peroxisomal MdAO1 prefers diamines as substrates, whereas extracellular MdAO2 exclusively utilizes monoamines.

PubMed

Zarei, Adel; Trobacher, Christopher P; Cooke, Alison R; Meyers, Ashley J; Hall, J Christopher; Shelp, Barry J

2015-01-01

4-Aminobutyrate (GABA) accumulates in apple fruit during controlled atmosphere storage. A potential source of GABA is the polyamine putrescine, which can be oxidized via copper-containing amine oxidase (CuAO), resulting in the production 4-aminobutanal/Δ(1)-pyrroline, with the consumption of O2 and release of H2O2 and ammonia. Five putative CuAO genes (MdAO genes) were cloned from apple (Malus domestica Borkh. cv. Empire) fruit, and the deduced amino acid sequences found to contain the active sites typically conserved in CuAOs. Genes encoding two of these enzymes, MdAO1 and MdAO2, were highly expressed in apple fruit and selected for further analysis. Amino acid sequence analysis predicted the presence of a C-terminal peroxisomal targeting signal 1 tripeptide in MdAO1 and an N-terminal signal peptide and N-glycosylation site in MdAO2. Transient expression of green fluorescent fusion proteins in Arabidopsis protoplasts or onion epidermal cells revealed a peroxisomal localization for MdAO1 and an extracellular localization for MdAO2. The enzymatic activities of purified recombinant MdAO1 and MdAO2 were measured continuously as H2O2 production using a coupled reaction. MdAO1 did not use monoamines or polyamines and displayed high catalytic efficiency for 1,3-diaminopropane, putrescine and cadaverine, whereas MdAO2 exclusively utilized aliphatic and aromatic monoamines, including 2-phenylethylamine and tyramine. Together, these results indicate that MdAO1 may contribute to GABA production via putrescine oxidation in the peroxisome of apple fruit under controlled atmosphere conditions. MdAO2 seems to be involved in deamination of 2-phenylethylamine, which is a step in the biosynthesis of 2-phenylethanol, a contributor to fruit flavor and flower fragrance. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Plasma clearance of neostigmine and pyridostigmine in the dog.

PubMed Central

Baker, P R; Calvey, T N; Chan, K; Macnee, C M; Taylor, K

1978-01-01

1 The pharmacokinetics of neostigmine and pyridostigmine was studied in conscious dogs by the use of a cross-over design. 2 Both neostigmine and pyridostigmine were cleared from plasma in a biexponential manner. 3 The apparent volume of distribution of pyridostigmine was invariably greater than that of neostigmine, and its fast disposition half-life was approximately three times longer. 4 The whole body clearance and the urinary elimination of pyridostigmine was approximately twice that of neostigmine. 5 The slow disposition half-life of pyridostigmine was approximately three times longer than that of neostigmine, suggesting that the longer duration of action of pyridostigmine is related to the differential clearance of the two quarternary amines from plasma. PMID:667495

Regulation of the nitric oxide oxidase activity of myeloperoxidase by pharmacological agents.

PubMed

Maiocchi, Sophie L; Morris, Jonathan C; Rees, Martin D; Thomas, Shane R

2017-07-01

The leukocyte-derived heme enzyme myeloperoxidase (MPO) is released extracellularly during inflammation and impairs nitric oxide (NO) bioavailability by directly oxidizing NO or producing NO-consuming substrate radicals. Here, structurally diverse pharmacological agents with activities as MPO substrates/inhibitors or antioxidants were screened for their effects on MPO NO oxidase activity in human plasma and physiological model systems containing endogenous MPO substrates/antioxidants (tyrosine, urate, ascorbate). Hydrazide-based irreversible/reversible MPO inhibitors (4-ABAH, isoniazid) or the sickle cell anaemia drug, hydroxyurea, all promoted MPO NO oxidase activity. This involved the capacity of NO to antagonize MPO inhibition by hydrazide-derived radicals and/or the ability of drug-derived radicals to stimulate MPO turnover thereby increasing NO consumption by MPO redox intermediates or NO-consuming radicals. In contrast, the mechanism-based irreversible MPO inhibitor 2-thioxanthine, potently inhibited MPO turnover and NO consumption. Although the phenolics acetaminophen and resveratrol initially increased MPO turnover and NO consumption, they limited the overall extent of NO loss by rapidly depleting H 2 O 2 and promoting the formation of ascorbyl radicals, which inefficiently consume NO. The vitamin E analogue trolox inhibited MPO NO oxidase activity in ascorbate-depleted fluids by scavenging NO-consuming tyrosyl and urate radicals. Tempol and related nitroxides decreased NO consumption in ascorbate-replete fluids by scavenging MPO-derived ascorbyl radicals. Indoles or apocynin yielded marginal effects. Kinetic analyses rationalized differences in drug activities and identified criteria for the improved inhibition of MPO NO oxidase activity. This study reveals that widely used agents have important implications for MPO NO oxidase activity under physiological conditions, highlighting new pharmacological strategies for preserving NO bioavailability during inflammation. Copyright © 2017 Elsevier Inc. All rights reserved.

Properties of cupric ions in benzylamine oxidase from pig plasma as studied by magnetic-resonance and kinetic methods.

PubMed Central

Barker, R; Boden, N; Cayley, G; Charlton, S C; Henson, R; Holmes, M C; Kelly, I D; Knowles, P F

1979-01-01

Benzylamine oxidase from pig plasma has been studied by a variety of chemical and physical techniques. 1. Analytical ultracentrifugation, gel electrophoresis and isoelectric-focusing studies suggest that the enzyme is composed of two subunits with closely similar primary structures. 2. E.s.r. and n.m.r. measurements show that the enzyme contains two well-separated (greater than 0.6 nm) Cu2+ ions at chemically distinct sites. Each Cu2+ ion is coordinated by two water molecules, one 'axial' and the other 'equatorial'. Both water molecules undergo fast exchange (10(5)--10(8) s-1) with solvent and are deprotonated in the pH range 8--9, but only the equatorial water molecule is displaced by the inhibitors N3- and CN-. 3. Kinetic and e.s.r. measurements show that azide and cyanide compete against O2 binding and also make the two Cu2+ sites identical. It is concluded that Cu2+ must participate in the re-oxidation of reduced enzyme by molecular O2. PMID:218560

Phone camera detection of glucose blood level based on magnetic particles entrapped inside bubble wrap.

PubMed

Martinkova, Pavla; Pohanka, Miroslav

2016-12-18

Glucose is an important diagnostic biochemical marker of diabetes but also for organophosphates, carbamates, acetaminophens or salicylates poisoning. Hence, innovation of accurate and fast detection assay is still one of priorities in biomedical research. Glucose sensor based on magnetic particles (MPs) with immobilized enzymes glucose oxidase (GOx) and horseradish peroxidase (HRP) was developed and the GOx catalyzed reaction was visualized by a smart-phone-integrated camera. Exponential decay concentration curve with correlation coefficient 0.997 and with limit of detection 0.4 mmol/l was achieved. Interfering and matrix substances were measured due to possibility of assay influencing and no effect of the tested substances was observed. Spiked plasma samples were also measured and no influence of plasma matrix on the assay was proved. The presented assay showed complying results with reference method (standard spectrophotometry based on enzymes glucose oxidase and peroxidase inside plastic cuvettes) with linear dependence and correlation coefficient 0.999 in concentration range between 0 and 4 mmol/l. On the grounds of measured results, method was considered as highly specific, accurate and fast assay for detection of glucose.

Deposition of chemically reactive and repellent sites on biosensor chips for reduced non-specific binding.

PubMed

Gandhiraman, R P; Gubala, V; Le, N C H; Nam, Le Cao Hoai; Volcke, C; Doyle, C; James, B; Daniels, S; Williams, D E

2010-08-01

The performances of new polymeric materials with excellent optical properties and good machinability have led the biomedical diagnostics industry to develop cheap disposable biosensor platforms appropriate for point of care applications. Zeonor, a type of cycloolefin polymer (COP), is one such polymer that presents an excellent platform for biosensor chips. These polymer substrates have to be modified to have suitable physico-chemical properties for immobilizing proteins. In this work, we have demonstrated the amine functionalization of COP substrates, by plasma enhanced chemical vapour deposition (PECVD), through codeposition of ethylene diamine and 3-aminopropyltriethoxysilane precursors, for building chemistries on the plastic chip. The elemental composition, adhesion, ageing and reactivity of the plasma polymerized film were examined. The Si-O functionality present in amino silane contributed for a good interfacial adhesion of the coating to COP substrates and also acted as a network building layer for plasma polymerization. Wet chemical modification was then carried out on the amine functionalized chips to create chemically reactive isothiocyanate sites and protein repellent fluorinated sites on the same chip. The density of the reactive and repellent sites was altered by choosing appropriate mixtures of homofunctional phenyldiisothiocyanate (PDITC), pentafluoroisothiocyanate (5FITC) and phenylisothiocyanate (PITC) compounds. By tailoring the density of reactive binding sites and protein repellent sites, the non-specific binding of ssDNA has been decreased to a significant extent. Copyright 2010 Elsevier B.V. All rights reserved.

Electron spin resonance characterization of vascular xanthine and NAD(P)H oxidase activity in patients with coronary artery disease: relation to endothelium-dependent vasodilation.

PubMed

Spiekermann, Stephan; Landmesser, Ulf; Dikalov, Sergey; Bredt, Martin; Gamez, Graciela; Tatge, Helma; Reepschläger, Nina; Hornig, Burkhard; Drexler, Helmut; Harrison, David G

2003-03-18

Increased inactivation of nitric oxide by superoxide (O2*-) contributes to endothelial dysfunction in patients with coronary disease (CAD). We therefore characterized the vascular activities of xanthine oxidase and NAD(P)H oxidase, 2 major O2*--producing enzyme systems, and their relationship with flow-dependent, endothelium-mediated vasodilation (FDD) in patients with CAD. Xanthine- and NAD(P)H-mediated O*.- formation was determined in coronary arteries from 10 patients with CAD and 10 controls by using electron spin resonance spectroscopy. Furthermore, activity of endothelium-bound xanthine oxidase in vivo and FDD of the radial artery were determined in 21 patients with CAD and 10 controls. FDD was measured before and after infusion of the antioxidant vitamin C (25 mg/min i.a.) to determine the portion of FDD inhibited by radicals. In coronary arteries from patients with CAD, xanthine- and NAD(P)H-mediated O2*- formation was increased compared with controls (xanthine: 12+/-2 versus 7+/-1 nmol O2*-/ microg protein; NADH: 11+/-1 versus 7+/-1 nmol O2*-/ microg protein; and NADPH: 12+/-2 versus 9+/-1 nmol O2*-/ microg protein; each P<0.05). Endothelium-bound xanthine oxidase activity was increased by >200% in patients with CAD (25+/-4 versus 9+/-1 nmol O2*-/ microL plasma per min; P<0.05) and correlated inversely with FDD (r=-0.55; P<0.05) and positively with the effect of vitamin C on FDD (r=0.54; P<0.05). The present study represents the first electron spin resonance measurements of xanthine and NAD(P)H oxidase activity in human coronary arteries and supports the concept that increased activities of both enzymes contribute to increased vascular oxidant stress in patients with CAD. Furthermore, the present study suggests that increased xanthine oxidase activity contributes to endothelial dysfunction in patients with CAD and may thereby promote the atherosclerotic process.

MENTAL DEPRESSION AND KUNDALINI YOGA

PubMed Central

Devi, Sanjenbam Kunjeshwori; Chansauria, J. P. N.; Udupa, K.N.

1986-01-01

In cases of mental depression, the plasma serotonin, melatonin and glutamate levels are increased along with the lowering of urinary – 5 – hydroxyindole acetic acid, plasma monoamine oxidase and cortisol levels following three and six months Practice of Kundalini Yoga. The pulse rate and blood pressure in these patients are also lowered after Kundalini Yoga practice. Thus, the practice of Kundalini Yoga helps to maintain a perfect homeostasis by bringing an equilibrium between the sympathetic and parasympathetic activities and it can be used as a non – medical measure in treating patients with mental depression. PMID:22557558

Mental depression and kundalini yoga.

PubMed

Devi, S K; Chansauria, J P; Udupa, K N

1986-10-01

In cases of mental depression, the plasma serotonin, melatonin and glutamate levels are increased along with the lowering of urinary - 5 - hydroxyindole acetic acid, plasma monoamine oxidase and cortisol levels following three and six months Practice of Kundalini Yoga. The pulse rate and blood pressure in these patients are also lowered after Kundalini Yoga practice. Thus, the practice of Kundalini Yoga helps to maintain a perfect homeostasis by bringing an equilibrium between the sympathetic and parasympathetic activities and it can be used as a non - medical measure in treating patients with mental depression.

The rebirth of interest in renal tubular function.

PubMed

Lowenstein, Jerome; Grantham, Jared J

2016-06-01

The measurement of glomerular filtration rate by the clearance of inulin or creatinine has evolved over the past 50 years into an estimated value based solely on plasma creatinine concentration. We have examined some of the misconceptions and misunderstandings of the classification of renal disease and its course, which have followed this evolution. Furthermore, renal plasma flow and tubular function, which in the past were estimated by the clearance of the exogenous aryl amine, para-aminohippurate, are no longer measured. Over the past decade, studies in experimental animals with reduced nephron mass and in patients with reduced renal function have identified small gut-derived, protein-bound uremic retention solutes ("uremic toxins") that are poorly filtered but are secreted into the lumen by organic anion transporters (OATs) in the proximal renal tubule. These are not effectively removed by conventional hemodialysis or peritoneal dialysis. Residual renal function, urine produced in patients with advanced renal failure or undergoing dialysis treatment, may represent, at least in part, secretion of fluid and uremic toxins, such as indoxyl sulfate, mediated by proximal tubule OATs and might serve as a useful survival function. In light of this new evidence of the physiological role of proximal tubule OATs, we suggest that measurement of renal tubular function and renal plasma flow may be of considerable value in understanding and managing chronic kidney disease. Data obtained in normal subjects indicate that renal plasma flow and renal tubular function might be measured by the clearance of the endogenous aryl amine, hippurate. Copyright © 2016 the American Physiological Society.

Hepatocyte transplants improve liver function and encephalopathy in portacaval shunted rats.

PubMed

Fogel, Wieslawa Agnieszka; Stasiak, Anna; Maksymowicz, Michał; Kobos, Jozef; Unzeta, Mercedes; Mussur, Miroslaw

2014-07-01

Rats with portacaval shunt (PCS) are useful experimental models of human hepatic encephalopathy in chronic liver dysfunction. We have previously shown that PCS modifies amine neurotransmitter systems in the CNS and increases voluntary alcohol intake by rats. Hepatocyte transplantation, used in acute liver failure, has recently also been applied to chronic liver diseases, which prompted us to investigate whether the altered brain amine system and the drinking behavior in long-term shunted rats could be normalized by hepatocyte transplants. Hepatocytes, isolated from syngeneic donors by collagenase digestion, were injected (3 × 10(6) cells/rat) into the pancreatic tail region, 6 months after PCS. Hepatic function was evaluated by measuring urine urea and plasma L-histidine concentrations. A free choice test with two bottles (tap water and 10% ethyl alcohol) was performed for 3 days to assess the rats' preference for alcohol. The rats were euthanized 2 months posttransplantation. Brain histamine and 5-hydroxyindoleacetic acid (5-HIAA) levels were measured by radioenzymatic assay and by HPLC-EC, respectively, N-tele-methylhistamine by GC/MS while MAOA and MAOB activities by isotopic procedures. Portacaval shunt rats with hepatocyte transplants gave more urea than before transplantation, with lower plasma L-His levels and higher body weight versus the PCS counterparts. Also, those rats consumed less alcohol. The CNS amines and 5-HIAA concentrations, as well as MAO-B activity, being abnormally high in untreated PCS rats, significantly reduced after PCS hepatocyte treatment. The results support the therapeutic values of hepatocyte transplants in chronic liver diseases and the temporary character of PCS-exerted CNS dysfunctions. © 2014 John Wiley & Sons Ltd.

Colorimetric assay of heparin in plasma based on the inhibition of oxidase-like activity of citrate-capped platinum nanoparticles.

PubMed

You, Jyun-Guo; Liu, Yao-Wen; Lu, Chi-Yu; Tseng, Wei-Lung; Yu, Cheng-Ju

2017-06-15

We report citrate-capped platinum nanoparticles (Pt NPs) as oxidase mimetics for effectively catalyzing the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB), 2,2'-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid), dopamine, and methylene blue in the presence of O 2 . To confirm oxidase-like activity of citrate-capped Pt NPs, their activity toward oxygen reduction reaction was studied using cyclic voltammetry and rotating ring-disk electrode method. The results obtained showed that Pt NP NPs can catalyze the oxidation of organic substrates to the colored product and the reduction of oxygen to water through a four-electron exchange process. Because the aggregation of Pt NPs can inhibit their oxidase-like activity and protamine can recognize heparin, we prepared the protamine-modified Pt NPs through direct adsorption on the surface of citrate-capped Pt NPs. The electrostatic attraction between heparin and protamine-stabilized Pt NPs induced nanoparticle aggregation, inhibiting their catalytic activity. Therefore, the lowest detectable heparin concentrations through UV-vis absorption and by the naked eye were estimated to be 0.3 and 60nM, respectively. Moreover, the proposed system enabled the determination of the therapeutic heparin concentration in a single drop of blood. Copyright © 2016 Elsevier B.V. All rights reserved.

Effect of scopoletin on monoamine oxidases and brain amines.

PubMed

Basu, Mahua; Mayana, Kamlesh; Xavier, S; Balachandran, S; Mishra, Nibha

2016-02-01

Naturally, occurring compounds with MAO inhibitory property may provide promising lead molecules against neurodegenerative disorders. We report MAO inhibitory activity of a naturally occurring coumarin (validated chemical scaffold as MAO inhibitors), scopoletin. It selectively (and reversibly) inhibits human (Ki = 20.7 μM) and mouse (Ki = 22 μM) MAO-B, ∼3.5 times more selective towards MAO-B than MAO-A. Docking studies revealed its molecular recognition and explained the selectivity mechanism towards MAO isoforms. Scopoletin occupied the hydrophobic aromatic pockets showing favorable interactions for MAO-B; experimental Ki agreed with the predicted Ki. In vivo, scopoletin (80 mg/kg, i.p.) treatment significantly increases dopamine level and decreases its metabolite DOPAC in striatum. Overall, scopoletin is a partially selective MAO-B inhibitor that increases brain dopamine level. Copyright © 2016. Published by Elsevier Ltd.

Distribution, industrial applications, and enzymatic synthesis of D-amino acids.

PubMed

Gao, Xiuzhen; Ma, Qinyuan; Zhu, Hailiang

2015-04-01

D-Amino acids exist widely in microbes, plants, animals, and food and can be applied in pharmaceutical, food, and cosmetics. Because of their widespread applications in industry, D-amino acids have recently received more and more attention. Enzymes including D-hydantoinase, N-acyl-D-amino acid amidohydrolase, D-amino acid amidase, D-aminopeptidase, D-peptidase, L-amino acid oxidase, D-amino acid aminotransferase, and D-amino acid dehydrogenase can be used for D-amino acids synthesis by kinetic resolution or asymmetric amination. In this review, the distribution, industrial applications, and enzymatic synthesis methods are summarized. And, among all the current enzymatic methods, D-amino acid dehydrogenase method not only produces D-amino acid by a one-step reaction but also takes environment and atom economics into consideration; therefore, it is deserved to be paid more attention.

Antioxidant capacity of human blood plasma and human urine: simultaneous evaluation of the ORAC index and ascorbic acid concentration employing pyrogallol red as probe.

PubMed

Torres, P; Galleguillos, P; Lissi, E; López-Alarcón, C

2008-10-15

The oxygen radical absorbance capacity (ORAC) methodology has been employed to estimate the antioxidant capacity of human blood plasma and human urine using pyrogallol red (ORAC-PGR) as target molecule. Uric acid, reduced glutathione, human serum albumin, and ascorbic acid (ASC) inhibited the consumption of pyrogallol red, but only ASC generated an induction time. Human blood plasma and human urine protected efficiently pyrogallol red. In these assays, both biological fluids generated neat induction times that were removed by ascorbate oxidase. From these results, ORAC-PGR method could be proposed as a simple alternative to evaluate an ORAC index and, simultaneously, to estimate the concentration of ascorbic acid in human blood plasma or human urine.

Host-parasite co-metabolic activation of antitrypanosomal aminomethyl-benzoxaboroles

PubMed Central

Scullion, Paul; del Pino, Ricardo C.; Vincent, Isabel M.; Zhang, Yong-Kang; Alley, Michael R. K.; Jacobs, Robert T.; Read, Kevin D.

2018-01-01

Recent development of benzoxaborole-based chemistry gave rise to a collection of compounds with great potential in targeting diverse infectious diseases, including human African Trypanosomiasis (HAT), a devastating neglected tropical disease. However, further medicinal development is largely restricted by a lack of insight into mechanism of action (MoA) in pathogenic kinetoplastids. We adopted a multidisciplinary approach, combining a high-throughput forward genetic screen with functional group focused chemical biological, structural biology and biochemical analyses, to tackle the complex MoAs of benzoxaboroles in Trypanosoma brucei. We describe an oxidative enzymatic pathway composed of host semicarbazide-sensitive amine oxidase and a trypanosomal aldehyde dehydrogenase TbALDH3. Two sequential reactions through this pathway serve as the key underlying mechanism for activating a series of 4-aminomethylphenoxy-benzoxaboroles as potent trypanocides; the methylamine parental compounds as pro-drugs are transformed first into intermediate aldehyde metabolites, and further into the carboxylate metabolites as effective forms. Moreover, comparative biochemical and crystallographic analyses elucidated the catalytic specificity of TbALDH3 towards the benzaldehyde benzoxaborole metabolites as xenogeneic substrates. Overall, this work proposes a novel drug activation mechanism dependent on both host and parasite metabolism of primary amine containing molecules, which contributes a new perspective to our understanding of the benzoxaborole MoA, and could be further exploited to improve the therapeutic index of antimicrobial compounds. PMID:29425238

Emerging approaches in Parkinson’s disease – adjunctive role of safinamide

PubMed Central

Müller, Thomas

2016-01-01

Ongoing neuronal death in Parkinson’s disease (PD) causes an altered neurotransmission of various biogenic amines, particularly dopamine. As these changes do not follow a distinct pattern, they vary individually, and are differently pronounced. As a result, a heterogeneous onset of motor and nonmotor features occurs in each patient with PD during the whole course of the disease. PD actually describes a set of distinct diseases that manifest themselves in clinical syndromes with certain similarities but also great differences. This clinical picture responds to drugs with a broad spectrum of modes of actions better than to compounds with an exclusive focus on specific receptor subtypes. Therefore, safinamide is an ideal candidate for treatment of patients with PD, since its pharmacological profile includes reversible monoamine oxidase-B inhibition, blockade of voltage-dependent sodium channels, modulation of calcium channels, and inhibition of glutamate release. Safinamide is applied only once daily. Its oral dose ranges from 50 to 100 mg. Safinamide was well tolerated and safe in the clinical development program that demonstrated the amelioration of motor symptoms and OFF phenomena by safinamide when combined with dopamine agonists or levodopa. In the real world of maintenance of patients with PD, effects of safinamide application resemble therapy with classical monoamine oxidase inhibitors or amantadine in combination with other dopamine-substituting drugs. Safinamide is becoming increasingly available in the EU despite complex approval and pricing scenarios. PMID:27536120

Emerging approaches in Parkinson's disease - adjunctive role of safinamide.

PubMed

Müller, Thomas

2016-01-01

Ongoing neuronal death in Parkinson's disease (PD) causes an altered neurotransmission of various biogenic amines, particularly dopamine. As these changes do not follow a distinct pattern, they vary individually, and are differently pronounced. As a result, a heterogeneous onset of motor and nonmotor features occurs in each patient with PD during the whole course of the disease. PD actually describes a set of distinct diseases that manifest themselves in clinical syndromes with certain similarities but also great differences. This clinical picture responds to drugs with a broad spectrum of modes of actions better than to compounds with an exclusive focus on specific receptor subtypes. Therefore, safinamide is an ideal candidate for treatment of patients with PD, since its pharmacological profile includes reversible monoamine oxidase-B inhibition, blockade of voltage-dependent sodium channels, modulation of calcium channels, and inhibition of glutamate release. Safinamide is applied only once daily. Its oral dose ranges from 50 to 100 mg. Safinamide was well tolerated and safe in the clinical development program that demonstrated the amelioration of motor symptoms and OFF phenomena by safinamide when combined with dopamine agonists or levodopa. In the real world of maintenance of patients with PD, effects of safinamide application resemble therapy with classical monoamine oxidase inhibitors or amantadine in combination with other dopamine-substituting drugs. Safinamide is becoming increasingly available in the EU despite complex approval and pricing scenarios.

Monoamine Oxidase-A Inhibition and Associated Antioxidant Activity in Plant Extracts with Potential Antidepressant Actions

PubMed Central

Guillén, Hugo

2018-01-01

Monoamine oxidase (MAO) catalyzes the oxidative deamination of amines and neurotransmitters and is involved in mood disorders, depression, oxidative stress, and adverse pharmacological reactions. This work studies the inhibition of human MAO-A by Hypericum perforatum, Peganum harmala, and Lepidium meyenii, which are reported to improve and affect mood and mental conditions. Subsequently, the antioxidant activity associated with the inhibition of MAO is determined in plant extracts for the first time. H. perforatum inhibited human MAO-A, and extracts from flowers gave the highest inhibition (IC50 of 63.6 μg/mL). Plant extracts were analyzed by HPLC-DAD-MS and contained pseudohypericin, hypericin, hyperforin, adhyperforin, hyperfirin, and flavonoids. Hyperforin did not inhibit human MAO-A and hypericin was a poor inhibitor of this isoenzyme. Quercetin and flavonoids significantly contributed to MAO-A inhibition. P. harmala seed extracts highly inhibited MAO-A (IC50 of 49.9 μg/L), being a thousand times more potent than H. perforatum extracts owing to its content of β-carboline alkaloids (harmaline and harmine). L. meyenii root (maca) extracts did not inhibit MAO-A. These plants may exert protective actions related to antioxidant effects. Results in this work show that P. harmala and H. perforatum extracts exhibit antioxidant activity associated with the inhibition of MAO (i.e., lower production of H2O2). PMID:29568754

Inhibition of monoamine oxidase B (MAO-B) by Chinese herbal medicines.

PubMed

Lin, R D; Hou, W C; Yen, K Y; Lee, M H

2003-11-01

Monoamine oxidase (MAO) catalyzes the oxidative deamination of biogenic amines accompaned by the release of H2O2. Two subtypes, MAO-A and MAO-B, exist on the basis of their specificities to substrates and inhibitors. The regulation of MAO-B activity is important in the treatment of neurodegenerative diseases. Twenty-seven species of plants used in traditional Chinese medicines, selected from an enthnobotanical survey, were used in an investigation of their inhibitory effect on MAO-B in rat brain homogenates. The 50% aqueous methanol extracts of four active extracts, Arisaema amurense, Lilium brownii var. colchesteri, Lycium chinense, and Uncaria rhynchophylla, exhibited the best activity and selectivity towards MAO-B with IC50 values of 0.44, 0.29, 0.40, and 0.03 mg/ml, respectively. A kinetic study of MAO-B inhibition by the four extracts using the Lineweaver-Burk plot for each active extract revealed the IC50 concentrations, and results show that: Ki = 0.59 mg/ml for A. amurense for the mixed-type mode, Ki = 0.58 mg/ml for L. brownii var. colchesteri for the mixed-type mode, Ki = 5.01 mg/ml for L. chinense for the uncompetitive mode, and Ki = 0.02 mg/ml for U. rhynchophylla for the uncompetitive mode. These may therefore be candidates for use in delaying the progressive degeneration caused by neurological diseases.

Stress and Neuroendocrine Regulation of Immune Responses.

DTIC Science & Technology

1986-04-20

in spleen and thymus weights, and in T- cell responsivity to mitogens following * , rief periods of restraint or electric footshock, especially with...antigen (NDV), in addition to the previously * reported increases in plasma corticosterone . The cerebral responses are particularly i-vident in...hypophysectomized mice. in which we do not observe substantial increases in * lasma corticosterone . We have not observed consistent changes in cerebral amines

In vivo long-term kinetics of radiolabeled n,n-dimethyltryptamine and tryptamine.

PubMed

Vitale, Arturo A; Pomilio, Alicia B; Cañellas, Carlos O; Vitale, Martín G; Putz, Eva Maria; Ciprian-Ollivier, Jorge

2011-06-01

N,N-dimethyltryptamine (DMT), a strong psychodysleptic drug, has been found in higher plants, shamanic hallucinogenic beverages, and the urine of schizophrenic patients. The aim of this work was to gain better knowledge on the relationship between this drug and hallucinogenic processes by studying DMT behavior in comparison with tryptamine. (131)I-labeled DMT and tryptamine were injected into rabbits. γ-Camera and biodistribution studies were performed. Brain uptake, plasma clearance, and renal excretion were assessed for each indolealkylamine. DMT and tryptamine showed different behavior when brain uptake, residence time, and excretion were compared. Labeled DMT entered the brain 10 s after injection, crossed the blood-brain barrier, and bound to receptors; then it was partially renally excreted. It was detected in urine within 24 h after injection and remained in the brain, even after urine excretion ceased; up to 0.1% of the injected dose was detected at 7 d after injection in the olfactory bulb. In contrast, tryptamine was rapidly taken up in the brain and fully excreted 10 min after injection. To our knowledge, this is the first demonstration that exogenous DMT remains in the brain for at least 7 d after injection. Although labeled DMT and tryptamine behave as agonists for at least 5-hydroxytryptamine 2A receptor, 5-hydroxytryptamine 2C receptor, trace amine-associated receptor, and σ-1 putative receptor targets, binding to the latter can explain the different behavior of labeled DMT and tryptamine in the brain. The persistence in the brain can be further explained on the basis that DMT and other N,N-dialkyltryptamines are transporter substrates for both the plasma membrane serotonin transporter and the vesicle monoamine transporter 2. Furthermore, storage in vesicles prevents DMT degradation by monoamine oxidase. At high concentrations, DMT is taken up by the serotonin transporter and further stored in vesicles by the vesicle monoamine transporter 2, to be released under appropriate stimuli. Moreover, the (131)I-labeling proved to be a useful tool to perform long-term in vivo studies.

In vitro characterization of two different atmospheric plasma jet chemical functionalizations of titanium surfaces

NASA Astrophysics Data System (ADS)

Mussano, F.; Genova, T.; Verga Falzacappa, E.; Scopece, P.; Munaron, L.; Rivolo, P.; Mandracci, P.; Benedetti, A.; Carossa, S.; Patelli, A.

2017-07-01

Plasma surface activation and plasma polymers deposition are promising technologies capable to modulate biologically relevant surface features of biomaterials. The purpose of this study was to evaluate the biological effects of two different surface modifications, i.e. amine (NH2-Ti) and carboxylic/esteric (COOH/R-Ti) functionalities obtained from 3-aminopropyltriethoxysilane (3-APTES) and methylmethacrylate (MMA) precursors, respectively, through an atmospheric plasma jet RF-APPJ portable equipment. The coatings were characterized by Scanning Electron Microscopy, FT-IR spectroscopy, XPS and surface energy calculations. Stability in water and after UV sterilization were also verified. The pre-osteoblastic murine cell line MC3T3-E1 was used to perform the in-vitro tests. The treated samples showed a higher quantity of adsorbed proteins and improved osteoblast cells adhesion on the surfaces compared to the pristine titanium, in particular the COOH/R-Ti led to a nearly two-fold improvement. Cell proliferation on coated samples was initially (at 24 h) lower than on titanium control, while, at 48 h, COOH/R-Ti reached the proliferation rate of pristine titanium. Cells grown on NH2-Ti were more tapered and elongated in shape with lower areas than on COOH/R-Ti enriched surfaces. Finally, NH2-Ti significantly enhanced osteocalcin production, starting from 14 days, while COOH/R-Ti had this effect only from 21 days. Notably, NH2-Ti was more efficient than COOH/R-Ti at 21 days. The amine functionality elicited the most relevant osteogenic effect in terms of osteocalcin expression, thus establishing an interesting correlation between early cell morphology and later differentiation stages. Taken together, these data encourage the use of the functionalization procedures here reported in further studies.

Organization and mobility of CD11b/CD18 and targeting of superoxide on the surface of degranulated human neutrophils.

PubMed

Mukherjee, G; Rasmusson, B; Linner, J G; Quinn, M T; Parkos, C A; Magnusson, K E; Jesaitis, A J

1998-09-01

A monoclonal IgM, specifically recognizing both CD11b and CD18 of human neutrophils, was used to examine the organization and mobility of CD11b/CD18 in the plasma membrane of human neutrophils degranulated by dihydrocytochalasin B (dhCB) treatment and fMet-Leu-Phe (fMLF) stimulation. Subcellular fractionation analysis of untreated or dhCB-treated control neutrophils indicated that 20% of CD11b/CD18 cosedimented with plasma membrane and the remainder with specific granules. In contrast, fMLF stimulation of dhCB-treated cells caused a major reorganization of CD11b/CD18, in which 60-70% of CD11b/CD18 sedimented in dense plasma membrane fractions that were also enriched in superoxide-generating NADPH oxidase activity. Similarly pretreated neutrophils were fixed, immunogold labeled, and examined by scanning electron microscopy. Immunogold particles were distributed uniformly over the symmetrically ruffled surface of unstimulated neutrophils. On dhCB-treated cells, immunogold was mostly uniformly distributed on a smooth membrane with a small percentage of particles lining up into linear arrays. After fMLF + dhCB stimulation, CD11b/CD18 gold label was more abundant on the cell surface and formed large aggregates on polarized membrane protrusions. However, when cells were adhered to an albumin-coated quartz surface and stimulated with fMLF in the presence of dhCB, immunogold was excluded on the articulated and rounded cell body but concentrated on the periphery of adherent lamellae. Fluorescence photobleaching recovery indicated that in unstimulated cells 38 +/- 3% of CD11b/CD18 was mobile (R) with a diffusion constant D of 3.1 +/- 0.3 x 10(-10) cm2/s. Treatment with dhCB raised R and D 24 and 74%, respectively. Stimulation using 1 microM fMLF with dhCB lowered D and R to near control levels. Since NADPH oxidase and CD11b/CD18 cosediment in high-density plasma membrane domains after fMLF + dhCB stimulation, we speculate that a stimulus-induced reorganization of CD11b/CD18 and NADPH oxidase to common membrane domains may occur in fMLF + dhCB-degranulated neutrophils. Copyright 1998 Academic Press.

Effects of feeding endophyte-infected fescue seed to Holstein cows during the dry period on plasma nitric oxide (NO), xanthine oxidase (XO) and haptoglobin (Hp) status in newborn calves.

USDA-ARS?s Scientific Manuscript database

Fescue toxicosis in cattle, caused by ingestion of endophyte-infected fescue (EIF), is associated with decreased feed intake, growth, milk production and reproductive efficiency as well as decreased resistance to heat, transportation and immune stress. Increased inflammatory response to immune chal...

Decomposition Analyses Applied to a Complex Ultradian Biorhythm: The Oscillating NADH Oxidase Activity of Plasma Membranes Having a Potential Time-Keeping (Clock) Function

PubMed Central

Foster, Ken; Anwar, Nasim; Pogue, Rhea; Morré, Dorothy M.; Keenan, T. W.; Morré, D. James

2003-01-01

Seasonal decomposition analyses were applied to the statistical evaluation of an oscillating activity for a plasma membrane NADH oxidase activity with a temperature compensated period of 24 min. The decomposition fits were used to validate the cyclic oscillatory pattern. Three measured values, average percentage error (MAPE), a measure of the periodic oscillation, mean average deviation (MAD), a measure of the absolute average deviations from the fitted values, and mean standard deviation (MSD), the measure of standard deviation from the fitted values plus R-squared and the Henriksson-Merton p value were used to evaluate accuracy. Decomposition was carried out by fitting a trend line to the data, then detrending the data if necessary, by subtracting the trend component. The data, with or without detrending, were then smoothed by subtracting a centered moving average of length equal to the period length determined by Fourier analysis. Finally, the time series were decomposed into cyclic and error components. The findings not only validate the periodic nature of the major oscillations but suggest, as well, that the minor intervening fluctuations also recur within each period with a reproducible pattern of recurrence. PMID:19330112

Chemo-enzymatic Synthesis of Clickable Xylo-oligosaccharide Monomers from Hardwood 4-O-Methylglucuronoxylan.

PubMed

MacCormick, Benjamin; Vuong, Thu V; Master, Emma R

2018-02-12

A chemo-enzymatic pathway was developed to transform 4-O-methylglucuronic acid (MeGlcpA) containing xylo-oligosaccharides from beechwood into clickable monomers capable of polymerizing at room temperature and in aqueous conditions to form unique polytriazoles. While the gluco-oligosaccharide oxidase (GOOX) from Sarocladium strictum was used to oxidize C6-propargylated oligosaccharides, the acid-amine coupling reagents 1-ethyl-3-(3-(dimethylamino)propyl) carbodiimide (EDAC) and 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMT-MM) were employed and compared for their ability to append click functionalities to carboxylic acid groups of enzyme-treated oligosaccharides. While DMT-MM was a superior coupling reagent for this application, a triazine side product was observed during C-1 amidation. Resulting bifunctional xylo-oligosaccharide monomers were polymerized using a Cu(I) catalyst, forming a soft gel which was characterized by 1 H NMR, confirming the triazole product.

Copper import in Escherichia coli by the yersiniabactin metallophore system

PubMed Central

Koh, Eun-Ik; Robinson, Anne E.; Bandara, Nilantha; Rogers, Buck E.; Henderson, Jeffrey P.

2017-01-01

Copper plays a dual role as nutrient and toxin during bacterial infections. While uropathogenic Escherichia coli (UPEC) strains can use the copper-binding metallophore yersiniabactin (Ybt) to resist copper toxicity, Ybt also converts bioavailable copper to Cu(II)-Ybt in low copper conditions. Although E. coli have long been considered to lack a copper import pathway, we observed Ybt-mediated copper import in UPEC using canonical Fe(III)-Ybt transport proteins. UPEC removed copper from Cu(II)-Ybt with subsequent re-export of metal-free Ybt to the extracellular space. Copper released through this process became available to an E. coli cuproenzyme (the amine oxidase TynA), linking this import pathway to a nutrient acquisition function. Ybt-expressing E. coli thus engage in nutritional passivation, a strategy of minimizing a metal ion's toxicity while preserving its nutritional availability. Copper acquisition through this process may contribute to the marked virulence defect of Ybt transport-deficient UPEC. PMID:28759019

Optimization of orally bioavailable alkyl amine renin inhibitors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Zhenrong; Cacatian, Salvacion; Yuan, Jing

2010-09-17

Structure-guided drug design led to new alkylamine renin inhibitors with improved in vitro and in vivo potency. Lead compound 21a, has an IC{sub 50} of 0.83 nM for the inhibition of human renin in plasma (PRA). Oral administration of 21a at 10 mg/kg resulted in >20 h reduction of blood pressure in a double transgenic rat model of hypertension.

Simultaneous determination of multiclass pesticide residues in human plasma using a mini QuEChERS method.

PubMed

Srivastava, Anshuman; Rai, Satyajeet; Kumar Sonker, Ashish; Karsauliya, Kajal; Pandey, Chandra Prabha; Singh, Sheelendra Pratap

2017-06-01

Blood is one of the most assessable matrices for the determination of pesticide residue exposure in humans. Effective sample preparation/cleanup of biological samples is very important in the development of a sensitive, reproducible, and robust method. In the present study, a simple, cost-effective, and rapid gas chromatography-tandem mass spectrometry method has been developed and validated for simultaneous analysis of 31 multiclass (organophosphates, organochlorines, and synthetic pyrethroids) pesticide residues in human plasma by means of a mini QuEChERS (quick, easy, cheap, effective, rugged, and safe) method. We have adopted a modified version of the QuEChERS method, which is primarily used for pesticide residue analysis in food commodities. The QuEChERS method was optimized by use of different extraction solvents and different amounts and combinations of salts and sorbents (primary-secondary amines and C 18 ) for the dispersive solid-phase extraction step. The results show that a combination of ethyl acetate with 2% acetic acid, magnesium sulfate (0.4 g), and solid-phase extraction for sample cleanup with primary-secondary amines (50 mg) per 1-mL volume of plasma is the most suitable for generating acceptable results with high recoveries for all multiclass pesticides from human plasma. The mean recovery ranged from 74% to 109% for all the analytes. The limit of quantification and limit of detection of the method ranged from 0.12 to 13.53 ng mL -1 and from 0.04 to 4.10 ng mL -1 respectively. The intraday precision and the interday precision of the method were 6% or less and 11% or less respectively. This method would be useful for the analysis of a wide range of pesticides of interest in a small volume of clinical and/or forensic samples to support biomonitoring and toxicological applications. Graphical Abstract Pesticide residues analysis in human plasma using mini QuEChERS method.

Increased plasma membrane cholesterol in cystic fibrosis cells correlates with CFTR genotype and depends on de novo cholesterol synthesis

PubMed Central

2010-01-01

Background Previous observations demonstrate that Cftr-null cells and tissues exhibit alterations in cholesterol processing including perinuclear cholesterol accumulation, increased de novo synthesis, and an increase in plasma membrane cholesterol accessibility compared to wild type controls. The hypothesis of this study is that membrane cholesterol accessibility correlates with CFTR genotype and is in part influenced by de novo cholesterol synthesis. Methods Electrochemical detection of cholesterol at the plasma membrane is achieved with capillary microelectrodes with a modified platinum coil that accepts covalent attachment of cholesterol oxidase. Modified electrodes absent cholesterol oxidase serves as a baseline control. Cholesterol synthesis is determined by deuterium incorporation into lipids over time. Incorporation into cholesterol specifically is determined by mass spectrometry analysis. All mice used in the study are on a C57Bl/6 background and are between 6 and 8 weeks of age. Results Membrane cholesterol measurements are elevated in both R117H and ΔF508 mouse nasal epithelium compared to age-matched sibling wt controls demonstrating a genotype correlation to membrane cholesterol detection. Expression of wt CFTR in CF epithelial cells reverts membrane cholesterol to WT levels further demonstrating the impact of CFTR on these processes. In wt epithelial cell, the addition of the CFTR inhibitors, Gly H101 or CFTRinh-172, for 24 h surprisingly results in an initial drop in membrane cholesterol measurement followed by a rebound at 72 h suggesting a feedback mechanism may be driving the increase in membrane cholesterol. De novo cholesterol synthesis contributes to membrane cholesterol accessibility. Conclusions The data in this study suggest that CFTR influences cholesterol trafficking to the plasma membrane, which when depleted, leads to an increase in de novo cholesterol synthesis to restore membrane content. PMID:20487541

Self-assembling Gold Nanoparticle Monolayers in a Three-phase System - Overcoming Ligand Size Limitations

NASA Astrophysics Data System (ADS)

Yang, Guang; Nanda, Jagjit; Wang, Boya; Chen, Gang; Hallinan, Daniel T., Jr.

An effective self-assembly technique was developed to prepare centimeter-scale monolayer gold nanoparticle (Au NP) films of long-range order with hydrophobic ligands. Aqueous Au NPs were entrapped in the organic/aqueous interface where the Au NP surface was in situ modified with different types of amine ligands, including amine-terminated polystyrene. The Au NPs then spontaneously relocated to the air/water interface to form an NP monolayer. The spontaneous formation of an Au NP film at the organic/water interface was due to the minimization of the system Helmholtz free energy. Self-assembled Au NP films has a hexagonal close packed structure. The interparticle spacing was dictated by the amine ligand length. Thus-assembled Au NP monolayers exhibit tunable surface plasma resonance and excellent spacial homogeneity of surface-enhanced Raman-scattering. The ``air/water/oil'' self-assembly method developed in this study not only benefits the fundamental understanding of NP ligand conformations, but is also promising to scale up the manufacture of plasmonic nanoparticle devices with precisely designed optical properties. This study was financially supported by start-up funding supplied by the Florida State University and the FAMU-FSU College of Engineering.

Biochemical and hematological effects of lead ingestion in nestling American kestrels (Falco sparverius)

USGS Publications Warehouse

Hoffman, D.J.; Franson, J.C.; Pattee, O.H.; Bunck, C.M.; Murray, H.C.

1985-01-01

1. One-day old American kestrel (Faico sparverius) nestlings were orally dosed daily with 5 μl/g of corn oil (controls), 25, 125 or 625 mg/kg of metallic lead in corn oil for 10 days.2. Forty per cent of the nestlings receiving 625 mg/kg of lead died after 6 days and growth rates were significantly depressed in the two highest lead dosed groups. At 10 days hematocrit values were significantly lower in the two highest lead treated groups, and hemoglobin content and red blood cell (δ-aminolevulinic acid dehydratase (ALAD) activity was depressed in all lead treated groups. Plasma creatine phosphokinase decreased in the two highest treatment groups.3. Brain, liver and kidney ALAD activities, brain RNA to protein ratio and liver protein concentration decreased after lead exposure whereas liver DNA, DNA to RNA ratio and DNA to protein ratio increased. Brain monoamine oxidase and ATPase were not significantly altered.4. Measurements of the ontogeny of hematological variants and enzymes in normal development, using additional untreated nestlings, revealed decreases in red blood cell ALAD, plasma aspartate amino transferase, lactate dehydrogenase, brain DNA and RNA and liver DNA, whereas hematocrit, hemoglobin, plasma alkaline phosphatase, brain monoamine oxidase, brain ALAD and liver ALAD increased during the first 10 days of posthatching development.5. Biochemical and hematological alterations were more severe than those reported in adult kestrels or precocial young birds exposed to lead. Alterations may be due in part to delayed development.

Membraneless glucose/oxygen enzymatic fuel cells using redox hydrogel films containing carbon nanotubes.

PubMed

MacAodha, Domhnall; Ó Conghaile, Peter; Egan, Brenda; Kavanagh, Paul; Leech, Dónal

2013-07-22

Co-immobilisation of three separate multiple blue copper oxygenases, a Myceliophthora thermophila laccase, a Streptomyces coelicolor laccase and a Myrothecium verrucaria bilirubin oxidase, with an [Os(2,2'-bipyridine)2 (polyvinylimidazole)10Cl](+/2+) redox polymer in the presence of multi-walled carbon nanotubes (MWCNTs) on graphite electrodes results in enzyme electrodes that produce current densities above 0.5 mA cm(-2) for oxygen reduction at an applied potential of 0 V versus Ag/AgCl. Fully enzymatic membraneless fuel cells are assembled with the oxygen-reducing enzyme electrodes connected to glucose-oxidising anodes based on co-immobilisation of glucose oxidase or a flavin adenine dinucleotide-dependent glucose dehydrogenase with an [Os(4,4'-dimethyl-2,2'-bipyridine)2(polyvinylimidazole)10Cl](+/2+) redox polymer in the presence of MWCNTs on graphite electrodes. These fuel cells can produce power densities of up to 145 μW cm(-2) on operation in pH 7.4 phosphate buffer solution at 37 °C containing 150 mM NaCl, 5 mM glucose and 0.12 mM O2. The fuel cells based on Myceliophthora thermophila laccase enzyme electrodes produce the highest power density if combined with glucose oxidase-based anodes. Although the maximum power density of a fuel cell of glucose dehydrogenase and Myceliophthora thermophila laccase enzyme electrodes decreases from 110 μW cm(-2) in buffer to 60 μW cm(-2) on testing in artificial plasma, it provides the highest power output reported to date for a fully enzymatic glucose-oxidising, oxygen-reducing fuel cell in artificial plasma. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Polymerisation par plasma a pression atmospherique: Caracterisation des depots et leurs applications en biotechnologies

NASA Astrophysics Data System (ADS)

Girard-Lauriault, Pierre-Luc

Nitrogen (N)-containing polymer surfaces are attractive in numerous technological contexts, for example in biomedical applications. Here, we have used an atmospheric-pressure dielectric barrier discharge (DBD) apparatus to deposit novel families of N-rich plasma polymers, designated PP:N, using mixtures of three different hydrocarbon precursors (methane, ethylene, and acetylene) in nitrogen at varying respective gas flow ratios, typically parts per thousand. In preparation for subsequent cell-surface interaction studies, the first part of this research focuses on the chemical mapping of those materials, with specific attention to (semi)- quantitative analyses of functional groups. Well-established and some lesser-known analytical techniques have been combined to provide the best possible chemical and structural characterisations of these three families of PP:N thin films; namely, X-ray photoelectron spectroscopy (XPS), Near-edge X-ray absorption fine structure (NEXAFS), Fourier transform infrared spectroscopy (FTIR), contact angle goniometry (CAG), and elemental analysis (EA). High, "tunable" total nitrogen content was measured by both XPS and EA (between 6% and 25% by EA, or between 10% and 40% by XPS, which cannot detect hydrogen). Chemical derivatisation with 4-trifluoromethylbenzaldehyde (TFBA) enabled measurements of primary amine concentrations, the functionality of greatest bio-technological interest, which were found to account for 5 % to 20 % of the total bound nitrogen. By combining the above-mentioned complementary methods, we were further able to determine the complete chemical formulae, the degrees of unsaturation, and other major chemical functionalities in PP:N film structures. Several of these features are believed to be without precedents in the literature on hydrocarbon plasma polymers, for example measurements of absolute compositions (including hydrogen), and of unsaturation. It was shown that besides amines, nitriles, isonitriles and imines are the main nitrogenated functional groups in those materials. In a second part of this work, we have studied the interraction of these well-characterised surfaces with living cells. We have first demonstrated the adhesion, on both uniformly coated and micro-patterned PP:N deposits on BOPP, of three different cell types, namely, growth plate and articular chondrocytes, as well as U937 monocytes, the latter of which do not adhere at all to synthetic polymers used in tissue culture. In an effort to gain insight into cell adhesion mechanisms, we conducted a series of experiments where we cultured U937 monocytes on PP:N, as well as on two other families of chemically well-characterised N-rich thin films, the latter deposited by low pressure RF plasma and by vacuum ultra-violet (VUV) photo-polymerisation ("PVP:N" films). It was first shown that there exist sharply-defined ("critical") surface-chemical conditions that are necessary to induce cell adhesion. By comparing the extensively-characterised film chemistries at the " critical " conditions, we have clearly demonstrated the dominant role of primary amines in the cell adhesion mechanism. In the final aspect of this work, quantitative real-time reverse transcription-polymerase chain reaction (real-time RT-PCR) experiments were conducted using U937 cells that had been made to adhere on PP:N and PVP:N materials for up to 24h. We have shown that the adhesion of U937 monocytes to PP:N and PVP:N surfaces induced a transient expression of cytokines, markers of macrophage activation, as well as a sustained expression of PPARgamma and ICAM-I, implicated in the adhesion and retention of monocytes. Keywords: biomaterials; dielectric barrier discharges (DBD); deposition; plasma polymerisation; ESCA/XPS; NEXAFS; FTIR; primary amines; cell adhesion; gene expression.

Detection of glycoproteins in the Acanthamoeba plasma membrane

DOE Office of Scientific and Technical Information (OSTI.GOV)

Paatero, G.I.L.; Gahmberg, C.G.

1988-11-01

In the present study the authors have shown that glycoproteins are present in the plasma membrane of Acanthamoeba castellanii by utilizing different radioactive labeling techniques. Plasma membrane proteins in the amoeba were iodinated by {sup 125}I-lactoperoxidase labeling and the solubilized radiolabeled glycoproteins were separated by lectin-Sepharose affinity chromatography followed by polyacrylamide gel electrophoresis. The periodate/NaB{sup 3}H{sub 4} and galactose oxidase/NaB{sup 3}H{sub 4} labeling techniques were used for labeling of surface carbohydrates in the amoeba. Several surface-labeled glycoproteins were observed in addition to a diffusely labeled region with M{sub r} of 55,000-75,000 seen on electrophoresis, which could represent glycolipids. The presencemore » of glycoproteins in the plasma membrane of Acanthamoeba castellanii was confirmed by metabolic labeling with ({sup 35}S)methionine followed by lectin-Sepharose affinity chromatography and polyacrylamide gel electrophoresis.« less

Tailoring properties of reduced graphene oxide by oxygen plasma treatment

NASA Astrophysics Data System (ADS)

Kondratowicz, Izabela; Nadolska, Małgorzata; Şahin, Samet; Łapiński, Marcin; Prześniak-Welenc, Marta; Sawczak, Mirosław; Yu, Eileen H.; Sadowski, Wojciech; Żelechowska, Kamila

2018-05-01

We report an easily controllable, eco-friendly method for tailoring the properties of reduced graphene oxide (rGO) by means of oxygen plasma. The effect of oxygen plasma treatment time (1, 5 and 10 min) on the surface properties of rGO was evaluated. Physicochemical characterization using microscopic, spectroscopic and thermal techniques was performed. The results revealed that different oxygen-containing groups (e.g. carboxyl, hydroxyl) were introduced on the rGO surface enhancing its wettability. Furthermore, upon longer treatment time, other functionalities were created (e.g. quinones, lactones). Moreover, external surface of rGO was partially etched resulting in an increase of the material surface area and porosity. Finally, the oxygen plasma-treated rGO electrodes with bilirubin oxidase were tested for oxygen reduction reaction. The study showed that rGO treated for 10 min exhibited twofold higher current density than untreated rGO. The oxygen plasma treatment may improve the enzyme adsorption on rGO electrodes by introduction of oxygen moieties and increasing the porosity.

Effect of PCB and DES on rat monoamine oxidase, acetylcholinesterase, testosterone, and estradiol ontogeny

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vincent, D.R.; Bradshaw, W.S.; Booth, G.M.

1992-06-01

Diethylstilbestrol (DES) and polychlorinated biphenyl (PCB) have been documented as potentially hazardous environmental agents. In utero exposure to DES produces human vaginal adenocarcinoma, male reproductive tract lesions in mice, and has been correlated with personality changes in human males. PCB (Kanechlor) was found to be the major toxin in the {open_quotes}Yusho{close_quotes} rice oil poisoning in Japan in 1968. Other investigators have shown in rats that PCB (Arochlor) causes liver adenofibrosis, thyroid dysfunction, atypical mitochondria, and dilation of both smooth and rough endoplasmic reticulum. Matthews et al. (1978) also reported that 4, 4{prime} chlorinated biphenyl was the most potent inducer ofmore » monooxygenases, irrespective of chlorination at other sites. Although these compounds have been studied extensively in mammals, there is a paucity of data examining their effects when non-fetotoxic amounts are administered chronically and orally during gestation. The present study is part of a larger effort designed to establish a protocol for testing the developmental effects of xenobiotics such as DES and PCB. Levels of acetylcholinesterase (AChE) were measured as an indicator of the integrity of nerve transmission in the central nervous system. Monoamine oxidase (MAO) is a marker for the outer mitochondrial membrane and is an important amine metabolizing enzyme. Testosterone and estradiol are important sex steroids in mammals, and effects upon levels of the two hormones may signal anomalies in development of sex characteristics. 35 refs., 3 figs., 1 tab.« less

Fabrication of high performance bioanode based on fruitful association of dendrimer and carbon nanotube used for design O2/glucose membrane-less biofuel cell with improved bilirubine oxidase biocathode.

PubMed

Korani, Aazam; Salimi, Abdollah

2013-12-15

In this study, the preparation of an integrated modified electrode based on the covalent attachment of glucose dehydrogenase (GDH) enzyme and safranin O to amine-derivative multiwalled carbon nanotubes (MWCNTs-NH2) modified glassy carbon (GC) electrode using G2.5-carboxylated PAMAM dendrimer (Den) as linking agent is reported. The obtained results indicated that the proposed system has effective bioelectrocatalytic activity toward glucose oxidation at 100 mV with onset potential of -130 mV (vs. Ag/AgCl). The performance of the prepared hybrid system of GC/MWCNTs-NH2/Den/GDH/Safranin as anode in a membraneless enzyme-based glucose/O2 biofuel cell is further evaluated. The biocathode in this system was composed of bilirubin oxidase (BOX) enzyme immobilized onto a bilirubin modified carbon nanotube GC electrode. Immobilized BOX onto CNTs/bilirubin not only show direct electron transfer but also it has excellent electrocatalytic activity toward oxygen reduction at a positive potential of 610 mV. The open circuit voltage of the cell was 590 mV. The maximum current density was 0.5 mA cm(-2), while maximum power density of 108 μW cm(-2) was achieved at voltage of 330 mV. The immobilized enzymes in anode and cathode are very stable and output power of the BFC is approximately constant after 12 h continues operation. Copyright © 2013 Elsevier B.V. All rights reserved.

Linking Inflammation and Parkinson Disease: Hypochlorous Acid Generates Parkinsonian Poisons

PubMed Central

Jeitner, Thomas M.; Kalogiannis, Mike; Krasnikov, Boris F.; Gomlin, Irving; Peltier, Morgan R.; Moran, Graham R.

2016-01-01

Inflammation is a common feature of Parkinson Disease and other neurodegenerative disorders. Hypochlorous acid (HOCl) is a reactive oxygen species formed by neutrophils and other myeloperoxidase-containing cells during inflammation. HOCl chlorinates the amine and catechol moieties of dopamine to produce chlorinated derivatives collectively termed chlorodopamine. Here, we report that chlorodopamine is toxic to dopaminergic neurons both in vivo and in vitro. Intrastriatal administration of 90 nmol chlorodopamine to mice resulted in loss of dopaminergic neurons from the substantia nigra and decreased ambulation-results that were comparable to those produced by the same dose of the parkinsonian poison, 1-methyl-4-phenylpyridinium (MPP+). Chlorodopamine was also more toxic to differentiated SH SY5Y cells than HOCl. The basis of this selective toxicity is likely mediated by chlorodopamine uptake through the dopamine transporter, as expression of this transporter in COS-7 cells conferred sensitivity to chlorodopamine toxicity. Pharmacological blockade of the dopamine transporter also mitigated the deleterious effects of chlorodopamine in vivo. The cellular actions of chlorodopamine included inactivation of the α-ketoglutarate dehydrogenase complex, as well as inhibition of mitochondrial respiration. The latter effect is consistent with inhibition of cytochrome c oxidase. Illumination at 670 nm, which stimulates cytochrome c oxidase, reversed the effects of chlorodopamine. The observed changes in mitochondrial biochemistry were also accompanied by the swelling of these organelles. Overall, our findings suggest that chlorination of dopamine by HOCl generates toxins that selectively kill dopaminergic neurons in the substantia nigra in a manner comparable to MPP+. PMID:27026709

[Features of the biochemical action of paraquat on oxidative deamination of biogenic amines and other nitrogen compounds].

PubMed

Amanov, K; Mamadiev, M; Khuzhamberdiev, M A; Gorkin, V Z

1994-01-01

Intoxication of rats with the herbicide paraquat (1,1-dimethyl-4,4-bipyridilium dichloride) was accompanied by accumulation in lungs, brain, heart, liver or kidney of malonic dialdehyde (MDA) (the compounds reacting with 2-thiobarbituric acid), indicating that the intoxication stimulated lipid peroxidation (LPO) in biomembranes. Treatment of the intoxicated rats with the antioxidant diludin (2,6-dimethyl-3,5-diethoxycarbonyl-1,4-dihydropyridine) or with the nucleophilic reagents sodium ascorbate or thiosulphate normalized the content of MDA in lungs, brain, heart, liver or kidney demonstrating the reversibility of the LPO stimulation caused by paraquat. On incubation of mitochondrial fractions of homogenates of lungs, brain, heart, liver or kidney of the intoxicated rats (as compared with the corresponding fractions from the intact animals) a decrease was noted in deamination of the substrates of monoamine oxidases serotonin, tryptamine, benzylamine, tyramine; at the same time, deamination of glucosamine and gamma-aminobutyric acid was increased and deamination of putrescine and L-lysine appeared. These impairments in deamination of nitrogenous compounds caused by paraquat were reversible. All the impairments were normalized by the treatment of the experimental animals with the antioxidative and nucleophilic reagents; a decrease was noted in the rate of development of the lethal paraquat intoxication and appearance of morphological manifestations of normalization. The data obtained suggest that the reversible, qualitative modification ("transformation") of the monoamine oxidases of the type A might explain the peculiarities of the alterations in deamination of nitrogenous compounds in paraquat intoxication.

Transcriptome analyses provide insights into the difference of alkaloids biosynthesis in the Chinese goldthread (Coptis chinensis Franch.) from different biotopes.

PubMed

Chen, Hanting; Deng, Cao; Nie, Hu; Fan, Gang; He, Yang

2017-01-01

Coptis chinensis Franch., the Chinese goldthread ('Weilian' in Chinese), one of the most important medicinal plants from the family Ranunculaceae, and its rhizome has been widely used in Traditional Chinese Medicine for centuries. Here, we analyzed the chemical components and the transcriptome of the Chinese goldthread from three biotopes, including Zhenping, Zunyi and Shizhu. We built comprehensive, high-quality de novo transcriptome assemblies of the Chinese goldthread from short-read RNA-Sequencing data, obtaining 155,710 transcripts and 56,071 unigenes. More than 98.39% and 95.97% of core eukaryotic genes were found in the transcripts and unigenes respectively, indicating that this unigene set capture the majority of the coding genes. A total of 520,462, 493,718, and 507,247 heterozygous SNPs were identified in the three accessions from Zhenping, Zunyi, and Shizhu respectively, indicating high polymorphism in coding regions of the Chinese goldthread (∼1%). Chemical analyses of the rhizome identified six major components, including berberine, palmatine, coptisine, epiberberine, columbamine, and jatrorrhizine. Berberine has the highest concentrations, followed by coptisine, palmatine, and epiberberine sequentially for all the three accessions. The drug quality of the accession from Shizhu may be the highest among these accessions. Differential analyses of the transcriptome identified four pivotal candidate enzymes, including aspartate aminotransferaseprotein, polyphenol oxidase, primary-amine oxidase, and tyrosine decarboxylase, were significantly differentially expressed and may be responsible for the difference of alkaloids contents in the accessions from different biotopes.

Brain Monoamine Oxidase-A Activity Predicts Trait Aggression

PubMed Central

Alia-Klein, Nelly; Goldstein, Rita Z.; Kriplani, Aarti; Logan, Jean; Tomasi, Dardo; Williams, Benjamin; Telang, Frank; Shumay, Elena; Biegon, Anat; Craig, Ian W.; Henn, Fritz; Wang, Gene-Jack; Volkow, Nora D.; Fowler, Joanna S.

2008-01-01

The genetic deletion of monoamine oxidase A (MAO A, an enzyme which breaks down the monoamine neurotransmitters norepinephrine, serotonin and dopamine) produces aggressive phenotypes across species. Therefore, a common polymorphism in the MAO A gene (MAOA, MIM 309850, referred to as high or low based on transcription in non-neuronal cells) has been investigated in a number of externalizing behavioral and clinical phenotypes. These studies provide evidence linking the low MAOA genotype and violent behavior but only through interaction with severe environmental stressors during childhood. Here, we hypothesized that in healthy adult males the gene product of MAO A in the brain, rather than the gene per se, would be associated with regulating the concentration of brain amines involved in trait aggression. Brain MAO A activity was measured in-vivo in healthy non-smoking men with positron emission tomography using a radioligand specific for MAO A (clorgyline labeled with carbon 11). Trait aggression was measured with the Multidimensional Personality Questionnaire (MPQ). Here we report for the first time that brain MAO A correlates inversely with the MPQ trait measure of aggression (but not with other personality traits) such that the lower the MAO A activity in cortical and subcortical brain regions the higher the self-reported aggression (in both MAOA genotype groups) contributing to more than a third of the variability. Since trait aggression is a measure used to predict antisocial behavior, these results underscore the relevance of MAO A as a neurochemical substrate of aberrant aggression. PMID:18463263

Patterning of Biomolecules on Plasma-Enhanced Chemical Vapor Deposited Generated Surfaces

DTIC Science & Technology

2006-01-01

utilized conventional 1 -Ethyl- 3 -[ 3 - dimethylaminopropyl ]carbodiimide hydrochloride (EDC) to couple carboxyl groups to primary amines and thiol oxidation...RETURN YOUR FORM TO THE ABOVE ADDRESS. 1 . REPORT DATE (DD-MM-YY) 2. REPORT TYPE 3 . DATES COVERED (From - To) January 2006 Journal Article Preprint...morphology, allowing for robust surface deposited 3 materials. More specifically, bulk pp-benzene shows less than 1 % sol fraction when immersed in

Adenosine Amine Congener as a Cochlear Rescue Agent

PubMed Central

Vlajkovic, Srdjan M.; Chang, Hao; Paek, Song Yee; Chi, Howard H.-T.; Sreebhavan, Sreevalsan; Telang, Ravindra S.; Tingle, Malcolm; Housley, Gary D.; Thorne, Peter R.

2014-01-01

We have previously shown that adenosine amine congener (ADAC), a selective A1 adenosine receptor agonist, can ameliorate noise- and cisplatin-induced cochlear injury. Here we demonstrate the dose-dependent rescue effects of ADAC on noise-induced cochlear injury in a rat model and establish the time window for treatment. Methods. ADAC (25–300 μg/kg) was administered intraperitoneally to Wistar rats (8–10 weeks old) at intervals (6–72 hours) after exposure to traumatic noise (8–16 kHz, 110 dB sound pressure level, 2 hours). Hearing sensitivity was assessed using auditory brainstem responses (ABR) before and 12 days after noise exposure. Pharmacokinetic studies investigated ADAC concentrations in plasma after systemic (intravenous) administration. Results. ADAC was most effective in the first 24 hours after noise exposure at doses >50 μg/kg, providing up to 21 dB protection (averaged across 8–28 kHz). Pharmacokinetic studies demonstrated a short (5 min) half-life of ADAC in plasma after intravenous administration without detection of degradation products. Conclusion. Our data show that ADAC mitigates noise-induced hearing loss in a dose- and time-dependent manner, but further studies are required to establish its translation as a clinical otological treatment. PMID:25243188

Screening of Bothrops snake venoms for L-amino acid oxidase activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pessati, M.L.; Fontana, J.D.; Guimaraes, M.F.

1995-12-31

Toxins, enzymes, and biologically active peptides are the main components of snake venoms from the genus Bothrops. Following the venom inoculation, the local effects are hemorrhage, edema, and myonecrosis. Nineteen different species of Brazilian Bothrops were screened for protein content and L-amino acid oxidase activity. B. cotiara, formerly found in the South of Brazil, is now threatened with extinction. Its venom contains a highly hemorrhagic fraction and, as expected from the deep yellow color of the corresponding lyophilized powder, a high L-amino acid oxidase (LAO) activity was also characterized. Flavin adenine dinucleotide (FAD) is its associate coenzyme. B. cotiara venommore » LAO catalyzed the oxidative deamination of several L-amino acids, and the best substrates were methionine, leucine, tryptophan, and phenylalanine, hence, its potential application for the use in biosensors for aspartame determination and for the removal of amino acids from plasma. High levels for LAO were also found in other species than B. cotiara. In addition, the technique of isoelectric focusing (IEF) was employed as a powerful tool to study the iso- or multi-enzyme distribution for LAO activity in the B. cotiara snake venom.« less

Vitamin C prevents zidovudine-induced NAD(P)H oxidase activation and hypertension in the rat.

PubMed

Papparella, Italia; Ceolotto, Giulio; Berto, Laura; Cavalli, Maurizio; Bova, Sergio; Cargnelli, Gabriella; Ruga, Ezia; Milanesi, Ornella; Franco, Lorenzo; Mazzoni, Martina; Petrelli, Lucia; Nussdorfer, Gastone G; Semplicini, Andrea

2007-01-15

Cardiovascular risk is increased among HIV-infected patients receiving antiretroviral therapy due to the development of hypertension and metabolic abnormalities. In this study, we investigated the effects of long-term treatment with zidovudine (AZT) and vitamin C, alone and in combination, on blood pressure and on the chain of events linking oxidative stress to cardiac damage in the rat. Six adult Wistar Kyoto rats received AZT (1 mg/ml) in the drinking water for 8 months, six vitamin C (10 g/kg of food) and AZT, six vitamin C alone, and six served as controls. AZT increased systolic blood pressure, expression of gp91(phox) and p47(phox) subunits of NAD(P)H oxidase, and protein kinase C (PKC) delta activation and reduced antioxidant power of plasma and cardiac homogenates. AZT also caused morphological alterations in cardiac myocyte mitochondria, indicative of functional damage. All of these effects were prevented by vitamin C. Chronic AZT administration increases blood pressure and promotes cardiovascular damage through a NAD(P)H oxidase-dependent mechanism that involves PKC delta. Vitamin C antagonizes these adverse effects of AZT in the cardiovascular system.

Sensitive determination of four tetracycline antibiotics in pig plasma by field-amplified sample stacking open-tubular capillary electrochromatography with dimethylethanolamine aminated polychloromethyl styrene nano-latex coated capillary column.

PubMed

Guo, Yaxiao; Meng, Lei; Zhang, Yanhao; Tang, Wei; Zhang, Wenfen; Xia, Yan; Ban, Fuguo; Wu, Ningpeng; Zhang, Shusheng

2013-12-30

This paper described the preparation and application of a new dimethylethanolamine aminated polychloromethyl styrene nano-latex (DMEAPL) coated capillary column (ccc-DMEAPL) in the determination of four tetracycline antibiotics (TCA) including tetracycline (TC), oxytetracycline (OTC), doxycycline (DC) and chlorotetracycline (CTC) in pig plasma. The ccc-DMEAPL column was characterized with steady EOF values of ca. 1.5-5.2×10(-5)cm(2)/Vs at pH 1.8-6.3. The optimized conditions for field-amplified sample stacking open-tubular capillary electrochromatography (FASS-OT-CEC) were as following: background electrolyte, 10mmol/L Na2HPO4+15mmol/L citric acid (pH 3.2); ccc-DMEAPL, 50μm i.d.×50cm (effective length 41.5cm), separation voltage, 18kV; column temperature, 25°C; UV detection wavelength, 270nm; water-plug injection: 30mbar×10s; sample electrokinetic injection, 10kV×20s. The four TCA were extracted with the solution of 10mmol/L Na2HPO4+15mmol/L citric acid+4g/L EDTA-2Na (pH 3.2). The FASS-OT-CEC method was validated in terms of linearity, sensitivity, selectivity, precision and accuracy. The LODs ranged from 3 to 7ng/mL, the recoveries for the four TCA were all more than 80%. The developed method was successfully applied for the determination of TCs in the actual pig plasma samples. Copyright © 2013 Elsevier B.V. All rights reserved.

Oxygen plasma resistant phosphine oxide containing imide/arylene copolymers

NASA Technical Reports Server (NTRS)

Jensen, Brian J.

1993-01-01

A series of oxygen plasma resistant imide/arylene ether copolymers were prepared by reacting anhydride-terminated poly(amide acids) and amine-terminated polyarylene ethers containing phosphine oxide units. Inherent viscosities for these copolymers ranged from 0.42 to 0.80 dL/g. After curing, the resulting copolymers had glass transition temperatures ranging from 224 C to 228 C. Solution cast films of the block copolymers were tough and flexible with tensile strength, tensile moduli, and elongation at break up to 16.1 ksi, 439 ksi, and 23 percent, respectively at 25 C and 9.1 ksi, 308 ksi and 97 percent, respectively at 150 C. The copolymers show a significant improvement in resistance to oxygen plasma when compared to the commercial polyimide Kapton. The imide/arylene ether copolymers containing phosphine oxide units are suitable as coatings, films, adhesives, and composite matrices.

Colorimetric Glucose Assay Based on Magnetic Particles Having Pseudo-peroxidase Activity and Immobilized Glucose Oxidase.

PubMed

Martinkova, Pavla; Opatrilova, Radka; Kruzliak, Peter; Styriak, Igor; Pohanka, Miroslav

2016-05-01

Magnetic particles (MPs) are currently used as a suitable alternative for peroxidase in the construction of novel biosensors, analytic and diagnostic methods. Their better chemical and thermal stabilities predestine them as appropriate pseudo-enzymatic catalysts. In this point of view, our research was focused on preparation of simply and fast method for immobilization of glucose oxidase onto surface of MPs with peroxidase-like activity. Spectrophotometric method (wavelength 450 nm) optimized for glucose determination using modified MPs has been successfully developed. Concentration curve for optimization of method was assayed, and Michaelis-Menten constant (K m) calculated, maximum reaction rate (V max), limit of detection, and correlation coefficient were determined to be 0.13 mmol/l (2.34 mg/dl), 1.79 pkat, 3.74 µmol/l (0.067 mg/dl), and 0.996, respectively. Interferences of other sugars such as sucrose, sorbitol, deoxyribose, maltose, and fructose were determined as well as effect of substances presenting in plasma (ascorbic acid, reduced glutathione, trolox, and urea). Results in comparison with positive and negative controls showed no interferences of the other sugars and no influence of plasma substances to measuring of glucose. The constructed method showed corresponding results with linear dependence and a correlation coefficient of 0.997. Possibility of repeated use of modified MPs was successfully proved.

An improved amperometric L-lactate biosensor based on covalent immobilization of microbial lactate oxidase onto carboxylated multiwalled carbon nanotubes/copper nanoparticles/polyaniline modified pencil graphite electrode.

PubMed

Dagar, Kusum; Pundir, C S

2017-01-01

An improved amperometric l-lactate biosensor was constructed based on covalent immobilization of lactate oxidase (LOx) from Pediococcus species onto carboxylated multiwalled carbon nanotubes (cMWCNT)/copper nanoparticles (CuNPs)/polyaniline (PANI) hybrid film electrodeposited on the surface of a pencil graphite electrode (PGE). The enzyme electrode was characterized by cyclic voltammetry (CV), scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy and electrochemical impedance spectroscopy (EIS), while CuNPs synthesized by chemical reduction method, were characterized by transmission electron microscopy (TEM), UV spectrascopy and X-ray diffraction (XRD). The biosensor showed maximum response within 5s at pH 8.0 in 0.05M sodium phosphate buffer and 37°C, when operated at 20mVs -1 . The biosensor had a detection limit of 0.25μM with a wide working range between 1μM-2500μM. The biosensor was employed for measurement of l-lactic acid level in plasma of apparently healthy and diseased persons. Analytical recovery of added lactic acid in plasma was 95.5%. Within- and between-batch coefficients of variations were 6.24% and 4.19% respectively. There was a good correlation (R 2 =0.97) between plasma lactate values as measured by standard enzymatic spectrophotometric method and the present biosensor. The working enzyme electrode was used 180 times over a period of 140 days, when stored at 4°C. Copyright © 2016. Published by Elsevier Inc.

Selective reduction of N-oxides to amines: application to drug metabolism.

PubMed

Kulanthaivel, Palaniappan; Barbuch, Robert J; Davidson, Rita S; Yi, Ping; Rener, Gregory A; Mattiuz, Edward L; Hadden, Chad E; Goodwin, Lawrence A; Ehlhardt, William J

2004-09-01

Phase I oxidative metabolism of nitrogen-containing drug molecules to their corresponding N-oxides is a common occurrence. There are instances where liquid chromatography/tandem mass spectometry techniques are inadequate to distinguish this pathway from other oxidation processes, including C-hydroxylations and other heteroatom oxidations, such as sulfur to sulfoxide. Therefore, the purpose of the present study was to develop and optimize an efficient and practical chemical method to selectively convert N-oxides to their corresponding amines suitable for drug metabolism applications. Our results indicated that efficient conversion of N-oxides to amines could be achieved with TiCl(3) and poly(methylhydrosiloxane). Among them, we found TiCl(3) to be a facile and easy-to-use reagent, specifically applicable to drug metabolism. There are a few reports describing the use of TiCl(3) to reduce N-O bonds in drug metabolism studies, but this methodology has not been widely used. Our results indicated that TiCl(3) is nearly as efficient when the reductions were carried out in the presence of biological matrices, including plasma and urine. Finally, we have shown a number of examples where TiCl(3) can be successfully used to selectively reduce N-oxides in the presence of sulfoxides and other labile groups.

Antibacterial properties of chitosan-based coatings are affected by spacer-length and molecular weight

NASA Astrophysics Data System (ADS)

Vaz, Juliana M.; Taketa, Thiago B.; Hernandez-Montelongo, Jacobo; Chevallier, Pascale; Cotta, Monica A.; Mantovani, Diego; Beppu, Marisa M.

2018-07-01

Chitosan is a biopolymer with antibacterial properties, which are dependent on its molecular weight (Mw) and its degree of deacetylation (DDA). When grafted on surfaces as a coating, chitosan antibacterial efficiency is also dependent on the polymer chain conformation on the surface, as the amine groups, responsible of the antibacterial effect, should be available for contact with bacteria. To investigate this behavior, chitosans with different Mw were grafted onto plasma aminated surfaces through three different spacers: glutaric anhydride (GA), poly(ethylene-glycol) bis(carboxymethyl) ether (PEGb), and poly(ethylene-alt-maleic anhydride) (PA). The grafting efficiency was evaluated by X-ray Photoelectron Spectroscopy (XPS), contact angle and Rose Bengal test, while morphological features were assessed by profilometry analyses. Results evidenced a clear influence of the anchor arm length and of the Mw of chitosan both on the grafting efficiency and on the antibacterial behavior. PA CHIMW surface exhibited a better antibacterial response compared to GA and PEGb, which could be correlated to a denser coating coverage as seen by XPS and profilometry results. Further, PA CHIMW coating displayed a higher amine density, thus promoting the interaction with the bacteria cell wall. Based on these results, chitosan-based coatings can then be extended to a wide range of antibacterial applications.

Xanthine Oxidase Inhibition by Febuxostat Attenuates Experimental Atherosclerosis in Mice

PubMed Central

Nomura, Johji; Busso, Nathalie; Ives, Annette; Matsui, Chieko; Tsujimoto, Syunsuke; Shirakura, Takashi; Tamura, Mizuho; Kobayashi, Tsunefumi; So, Alexander; Yamanaka, Yoshihiro

2014-01-01

Atherosclerosis is a chronic inflammatory disease due to lipid deposition in the arterial wall. Multiple mechanisms participate in the inflammatory process, including oxidative stress. Xanthine oxidase (XO) is a major source of reactive oxygen species (ROS) and has been linked to the pathogenesis of atherosclerosis, but the underlying mechanisms remain unclear. Here, we show enhanced XO expression in macrophages in the atherosclerotic plaque and in aortic endothelial cells in ApoE−/− mice, and that febuxostat, a highly potent XO inhibitor, suppressed plaque formation, reduced arterial ROS levels and improved endothelial dysfunction in ApoE−/− mice without affecting plasma cholesterol levels. In vitro, febuxostat inhibited cholesterol crystal-induced ROS formation and inflammatory cytokine release in murine macrophages. These results demonstrate that in the atherosclerotic plaque, XO-mediated ROS formation is pro-inflammatory and XO-inhibition by febuxostat is a potential therapy for atherosclerosis. PMID:24686534

Therapeutic effects of proteoliposomes on X-linked chronic granulomatous disease: proof of concept using macrophages differentiated from patient-specific induced pluripotent stem cells

PubMed Central

Brault, Julie; Vaganay, Guillaume; Le Roy, Aline; Lenormand, Jean-Luc; Cortes, Sandra; Stasia, Marie José

2017-01-01

Chronic granulomatous disease (CGD) is a rare inherited immunodeficiency due to dysfunction of the phagocytic nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex leading to severe and recurrent infections in early childhood. The main genetic form is the X-linked CGD leading to the absence of cytochrome b558 composed of NOX2 and p22phox, the membrane partners of the NADPH oxidase complex. The first cause of death of CGD patients is pulmonary infections. Recombinant proteoliposome-based therapy is an emerging and innovative approach for membrane protein delivery, which could be an alternative local, targeted treatment to fight lung infections in CGD patients. We developed an enzyme therapy using recombinant NOX2/p22phox liposomes to supply the NADPH oxidase activity in X0-linked CGD (X0-CGD) macrophages. Using an optimized prokaryotic cell-free protein synthesis system, a recombinant cytochrome b558 containing functional hemes was produced and directly inserted into the lipid bilayer of specific liposomes. The size of the NOX2/p22phox liposomes was estimated to be around 700 nm. These proteoliposomes were able to generate reactive oxygen species (ROS) in an activated reconstituted cell-free NADPH oxidase activation assay in the presence of recombinant p47phox, p67phox and Rac, the cytosolic components of the NADPH oxidase complex. Furthermore, using flow cytometry and fluorescence microscopy, we demonstrated that cytochrome b558 was successfully delivered to the plasma membrane of X0-CGD-induced pluripotent stem cell (iPSC)-derived macrophages. In addition, NADPH oxidase activity was restored in X0-CGD iPSC-derived macrophages treated with NOX2/p22phox liposomes for 8 h without any toxicity. In conclusion, we confirmed that proteoliposomes provide a new promising technology for the delivery of functional proteins to the membrane of targeted cells. This efficient liposomal enzyme replacement therapy will be useful for future treatment of pulmonary infections in CGD patients refractory to conventional anti-infectious treatments. PMID:28356734

NADPH Oxidase Plays a Role on Ethanol-Induced Hypertension and Reactive Oxygen Species Generation in the Vasculature.

PubMed

Marchi, Katia Colombo; Ceron, Carla Speroni; Muniz, Jaqueline J; De Martinis, Bruno S; Tanus-Santos, José E; Tirapelli, Carlos Renato

2016-09-01

Investigate the role of NADPH oxidase on ethanol-induced hypertension and vascular oxidative stress. Male Wistar rats were treated with ethanol (20% v/v). Apocynin (10 mg/kg/day, i.p.) prevented ethanol-induced hypertension. The increased contractility of endothelium-intact and endothelium-denuded aortic rings from ethanol-treated rats to phenylephrine was prevented by apocynin. Ethanol consumption increased superoxide anion (O2 (-)) generation and lipid peroxidation and apocynin prevented these responses. The decrease on plasma and vascular nitrate/nitrite (NOx) levels induced by ethanol was not prevented by apocynin. Treatment with ethanol did not affect aortic levels of hydrogen peroxide (H2O2) or reduced glutathione (GSH). Ethanol did not alter the activities of xanthine oxidase (XO), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx). Ethanol increased the expression of Nox1, PKCδ, nNOS, SAPK/JNK and SOD2 in the rat aorta and apocynin prevented these responses. No difference on aortic expression of Nox2, Nox4, p47phox, Nox organizer 1 (Noxo1), eNOS and iNOS was detected after treatment with ethanol. Ethanol treatment did not alter the phosphorylation of SAPK/JNK, p38MAPK, c-Src, Rac1 or PKCδ. The major new finding of our study is that the increased vascular generation of reactive oxygen species (ROS) induced by ethanol is related to increased vascular Nox1/NADPH oxidase expression. This mechanism is involved in vascular dysfunction and hypertension induced by ethanol. Additionally, we conclude that ethanol consumption induces the expression of different proteins that regulate vascular contraction and growth and that NADPH oxidase-derived ROS play a role in such response. The key findings of our study are that ethanol-induced hypertension is mediated by NADPH oxidase. Moreover, increased vascular Nox1 expression is related to the generation of reactive oxygen species (ROS) by ethanol. Finally, ROS induced by ethanol increase the expression of the regulatory vascular proteins. © The Author 2016. Medical Council on Alcohol and Oxford University Press. All rights reserved.

Enzymatic measurement of free and esterified cholesterol levels in plasma and other biological preparations using the oxygen electrode in a modified glucose analyzer.

PubMed

Dietschy, J M; Weeks, L E; Delente, J J

1976-12-01

A method is described for assaying free and esterified cholesterol using the oxygen electrode in a modified glucose analyzer to measure the relative amount of oxygen utilization taking place during oxydation of free cholesterol by the enzyme, cholesterol oxidase. A second enzyme, cholesterol ester hydrolase, is utilized to generate free cholesterol from cholesterol esters. This assay procedure is rapid, specific, reproducible and applicable to the measurement of free and esterified cholesterol carried in the major plasma lipoprotein fractions of man and the rat and, in addition, it can be utilized for the assay of sterols in subcellular fractions of cells.

Influence of immobilization and forced swim stress on the neurotoxicity of lambda-cyhalothrin in rats: Effect on brain biogenic amines and BBB permeability.

PubMed

Shukla, Rajendra K; Dhuriya, Yogesh K; Chandravanshi, Lalit P; Gupta, Richa; Srivastava, Pranay; Pant, Aditya B; Kumar, Ajay; Pandey, Chandra M; Siddiqui, M Haris; Khanna, Vinay K

2017-05-01

Experimental studies have been carried out on rats to understand the influence of immobilization stress (IMS), a psychological stressor and forced swim stress (FSS), a physical stressor in the neurotoxicity of lambda-cyhalothrin (LCT), a new generation type II synthetic pyrethroid with extensive applications. No significant change in plasma corticosterone levels and blood brain barrier (BBB) permeability was observed in rats subjected to IMS (one session of 15min/day), FSS (one session of 3min/day) for 28days or LCT treatment (3.0mg/kg body weight, p.o. suspended in groundnut oil) for 3days (26th, 27th and 28th day) as compared to controls. Marginal changes in the levels of biogenic amines and their metabolites (NE, EPN, DA, HVA, DOPAC, 5-HT) in hypothalamus, frontal cortex, hippocampus, and corpus striatum were observed in rats subjected to IMS or FSS or LCT alone as compared to controls. It was interesting to note that pre-exposure to IMS or FSS followed by LCT treatment for 3days caused a marked increase in plasma corticosterone levels associated with disruption in the BBB permeability as compared to rats exposed to IMS or FSS or LCT alone. Pre-exposure to IMS or FSS followed by LCT treatment for 3days resulted to alter the levels of biogenic amines and their metabolites in hypothalamus, frontal cortex, hippocampus, and corpus striatum as compared to rats exposed to IMS or FSS or LCT alone. Although neurochemical changes were more intense in rats pre-exposed to IMS as compared to those subjected to FSS on LCT treatment, the results indicate that both psychological and physical stress could be important influencing factors in the neurotoxicity of LCT. Copyright © 2016. Published by Elsevier B.V.

Regulation of the NADPH Oxidase RBOHD During Plant Immunity.

PubMed

Kadota, Yasuhiro; Shirasu, Ken; Zipfel, Cyril

2015-08-01

Pathogen recognition induces the production of reactive oxygen species (ROS) by NADPH oxidases in both plants and animals. ROS have direct antimicrobial properties, but also serve as signaling molecules to activate further immune outputs. However, ROS production has to be tightly controlled to avoid detrimental effects on host cells, but yet must be produced in the right amount, at the right place and at the right time upon pathogen perception. Plant NADPH oxidases belong to the respiratory burst oxidase homolog (RBOH) family, which contains 10 members in the model plant Arabidopsis thaliana. The perception of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs) leads to a rapid, specific and strong production of ROS, which is dependent on RBOHD. RBOHD is mainly controlled by Ca(2+) via direct binding to EF-hand motifs and phosphorylation by Ca(2+)-dependent protein kinases. Recent studies have, however, revealed a critical role for a Ca(2+)-independent regulation of RBOHD. The plasma membrane-associated cytoplasmic kinase BIK1 (BOTRYTIS-INDUCED KINASE1), which is a direct substrate of the PRR complex, directly interacts with and phosphorylates RBOHD upon PAMP perception. Impairment of these phosphorylation events completely abolishes the function of RBOHD in immunity. These results suggest that RBOHD activity is tightly controlled by multilayered regulations. In this review, we summarize recent advances in our understanding of the regulatory mechanisms controlling RBOHD activation. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Identification of NADPH oxidase family members associated with cold stress in strawberry.

PubMed

Zhang, Yunting; Li, Yali; He, Yuwei; Hu, Wenjie; Zhang, Yong; Wang, Xiaorong; Tang, Haoru

2018-04-01

NADPH oxidase is encoded by a small gene family (Respiratory burst oxidase homologs, Rbohs ) and plays an important role in regulating various biological processes. However, little information about this gene family is currently available for strawberry. In this study, a total of seven Rboh genes were identified from strawberry through genomewide analysis. Gene structure analysis showed the number of exons ranged from 10 to 23, implying that this variation occurred in FvRboh genes by the insertion and distribution of introns; the order and approximate size of exons were relatively conserved. FvRbohC was predicted to localize to the thylakoid membrane of the chloroplast, while other members were computed to localize to the plasma membrane, indicating different functions. Amino acid sequence alignment, conserved domain, and motif analysis showed that all identified FvRbohs had typical features of plant Rbohs. Phylogenetic analysis of Rbohs from strawberry, grape, Arabidopsis, and rice suggested that the FvRbohs could be divided into five subgroups and showed a closer relationship with those from grape and Arabidopsis than those from rice. The expression patterns of FvRboh genes in root, stem, leaf, flower, and fruit revealed robust tissue specificity. The expression levels of FvRbohA and FvRbohD were quickly induced by cold stress, followed by an increase in NADPH oxidase activity, leading to O2- accumulation and triggering the antioxidant reaction by the transient increases in SOD activity. This suggested these two genes may be involved in cold stress and defense responses in strawberry.

Operational impact of using a vanadate oxidase method for direct bilirubin measurements at an academic medical center clinical laboratory.

PubMed

Dhungana, Neha; Morris, Cory; Krasowski, Matthew D

2017-08-01

The aim of this study was to compare the operational impact of using vanadate oxidase versus diazo direct bilirubin assays for an academic medical center patient population. Retrospective study was done over an approximately 3.5 year period. The main automated chemistry instrumentation was a Roche Diagnostics cobas 8000 line. The Roche Direct Bilirubin assay was compared to Diazyme Laboratories Direct Bilirubin Assay and Randox Laboratories Direct Bilirubin assay using manufacturer's guidelines for hemolysis index, lipemia index, and analytical measurement range (AMR). Retrospective data was analyzed for 47,333 serum/plasma specimens that had clinical orders for direct bilirubin. A total of 5943 specimens (12.6%) exceeded the hemolysis index limit for the Roche method compared to only 0.2% and 0.05% of specimens for the Diazyme and Randox methods, respectively. The impact was particularly large on patients less than 2 years old, for which 51.3% of specimens exceeded the hemolysis index for the Roche method. A total of 1671 specimens (3.5%) exceeded the lipemia index limit for the Roche method compared to less than 0.1% for the Randox method. Lastly, 988 (2.1%) of specimens had direct bilirubin concentrations exceeding the upper AMR limit of 10 mg/dL [171 µmol/L] for the Roche assay compared to less than 1% of specimens for the vanadate oxidase methods. Vanadate oxidase direct bilirubin methods offer advantages over diazo methods in terms of less interference by hemolysis and lipemia, as well as wider AMR. The advantages are particularly evident for neonatal and infant populations.

Reactive oxygen species are involved in regulating alpha1-adrenoceptor-activated vascular smooth muscle contraction.

PubMed

Tsai, Ming-Ho; Jiang, Meei Jyh

2010-08-23

Reactive oxygen species (ROS) were shown to mediate aberrant contractility in hypertension, yet the physiological roles of ROS in vascular smooth muscle contraction have remained elusive. This study aimed to examine whether ROS regulate alpha1-adrenoceptor-activated contraction by altering myosin phosphatase activities. Using endothelium-denuded rat tail artery (RTA) strips, effects of anti-oxidants on isometric force, ROS production, phosphorylation of the 20-kDa myosin light chain (MLC20), and myosin phosphatase stimulated by alpha1-adrenoceptor agonist phenylephrine were examined. An antioxidant, N-acetyl-L-cysteine (NAC), and two NADPH oxidase inhibitors, apocynin and VAS2870, dose-dependently inhibited contraction activated by phenylephrine. Phenylephrine stimulated superoxide anion production that was diminished by the pretreatment of apocynin, VAS2870, superoxide scavenger tiron or mitochondria inhibitor rotenone, but not by xanthine oxidase inhibitor allopurinol or cyclooxygenase inhibitor indomethacin. Concurrently, NADPH oxidase activity in RTA homogenates increased within 1 min upon phenylephrine stimulation, sustained for 10 min, and was abolished by the co-treatment with apocynin, but not allopurinol or rotenone. Phenylephrine-induced MLC20 phosphorylation was dose-dependently decreased by apocynin. Furthermore, apocynin inhibited phenylephrine-stimulated RhoA translocation to plasma membrane and phosphorylation of both myosin phosphatase regulatory subunit MYPT1Thr855 and myosin phosphatase inhibitor CPI-17Thr38. ROS, probably derived from NADPH oxidase and mitochondria, partially regulate alpha1-adrenoceptor-activated smooth muscle contraction by altering myosin phosphatase-mediated MLC20 phosphorylation through both RhoA/Rho kinase- and CPI-17-dependent pathways.

Complementary probes reveal that phosphatidylserine is required for the proper transbilayer distribution of cholesterol.

PubMed

Maekawa, Masashi; Fairn, Gregory D

2015-04-01

Cholesterol is an essential component of metazoan cellular membranes and it helps to maintain the structural integrity and fluidity of the plasma membrane. Here, we developed a cholesterol biosensor, termed D4H, based on the fourth domain of Clostridium perfringens theta-toxin, which recognizes cholesterol in the cytosolic leaflet of the plasma membrane and organelles. The D4H probe disassociates from the plasma membrane upon cholesterol extraction and after perturbations in cellular cholesterol trafficking. When used in combination with a recombinant version of the biosensor, we show that plasmalemmal phosphatidylserine is essential for retaining cholesterol in the cytosolic leaflet of the plasma membrane. In vitro experiments reveal that 1-stearoy-2-oleoyl phosphatidylserine can induce phase separation in cholesterol-containing lipid bilayers and shield cholesterol from cholesterol oxidase. Finally, the altered transbilayer distribution of cholesterol causes flotillin-1 to relocalize to endocytic organelles. This probe should be useful in the future to study pools of cholesterol in the cytosolic leaflet of the plasma membrane and organelles. © 2015. Published by The Company of Biologists Ltd.

Surface modification of poly(dimethylsiloxane) for microfluidic assay applications

NASA Astrophysics Data System (ADS)

Séguin, Christine; McLachlan, Jessica M.; Norton, Peter R.; Lagugné-Labarthet, François

2010-02-01

The surface of a poly(dimethylsiloxane) (PDMS) film was imparted with patterned functionalities at the micron-scale level. Arrays of circles with diameters of 180 and 230 μm were functionalized using plasma oxidation coupled with aluminum deposition, followed by silanization with solutions of 3-aminopropyltrimethoxy silane (3-APTMS) and 3-mercaptopropyltrimethoxy silane (3-MPTMS), to obtain patterned amine and thiol functionalities, respectively. The modification of the samples was confirmed using X-ray photoelectron spectroscopy (XPS), gold nanoparticle adhesion coupled with optical microscopy, as well as by derivatization with fluorescent dyes. To further exploit the novel surface chemistry of the modified PDMS, samples with surface amine functionalities were used to develop a protein assay as well as an array capable of cellular capture and patterning. The modified substrate was shown to successfully selectively immobilize fluorescently labeled immunoglobulin G (IgG) by tethering Protein A to the surface, and, for the cellular arrays, C2C12 rat endothelial cells were captured. Finally, this novel method of patterning chemical functionalities onto PDMS has been incorporated into microfluidic channels. Finally, we demonstrate the in situ chemical modification of the protected PDMS oxidized surface within a microfluidic device. This emphasizes the potential of our method for applications involving micron-scale assays since the aluminum protective layer permits to functionalize the oxidized PDMS surface several weeks after plasma treatment simply after etching away the metallic thin film.

Mechanisms for Cellular NO Oxidation and Nitrite Formation in Lung Epithelial Cells

PubMed Central

Zhao, Xue-Jun; Wang, Ling; Shiva, Sruti; Tejero, Jesus; Wang, Jun; Frizzell, Sam; Gladwin, Mark T.

2013-01-01

Airway lining fluid contains relatively high concentrations of nitrite and arterial blood levels of nitrite are higher than venous levels, suggesting the lung epithelium may represent an important source of nitrite in vivo. To investigate whether lung epithelial cells possess the ability to convert NO to nitrite by oxidation, and the effect of oxygen reactions on nitrite formation, the NO donor DETA NONOate was incubated with or without A549 cells or primary human bronchial epithelial (HBE) cells for 24 hrs under normoxic (21% O2) and hypoxic (1% O2) conditions. Nitrite production was significantly increased under all conditions in the presence of A549 or HBE cells, suggesting that both A549 and HBE cells have the capacity to oxidize NO to nitrite even under low oxygen conditions. The addition of oxy-hemoglobin (oxy-Hb) to the A549 cell media decreased the production of nitrite, consistent with NO scavenging limiting nitrite formation. Heat-denatured A549 cells produced much lower nitrite and bitrate, suggesting an enzymatic activity is required. This NO oxidation activity was found to be highest in membrane bound proteins with molecular sizes < 100 kDa. In addition, 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one] (ODQ) and cyanide inhibited formation of nitrite in A549 cells. It has been shown that ceruloplasmin (Cp) possesses an NO oxidase and nitrite synthase activity in plasma based on NO oxidation to nitrosonium cation (NO+). We observed that Cp is expressed intracellularly in lung epithelial A549 cells and secreted into medium under basal conditions and during cytokine stimulation. However, an analysis of Cp expression level and activity measured via ρ-phenylenediamine oxidase activity assay revealed very low activity compared with plasma, suggesting that there is insufficient Cp to contribute to detectable NO oxidation to nitrite in A549 cells. Additionally, Cp levels were knocked down using siRNA by more than 75% in A549 cells, with no significant change in either nitrite or cellular S-nitrosothiol (SNO) formation compared to scrambled siRNA control under basal conditions or cytokine stimulation. These data suggest that lung epithelial cells possess NO oxidase activity, which is enhanced in cell membrane associated proteins and not regulated by intracellular or secreted Cp, indicating that alternative NO oxidases determine hypoxic and normoxic nitrite formation from NO in human lung epithelial cells. PMID:23639566

Monoamine oxidase-A polymorphisms might modify the association between the dopamine D2 receptor gene and alcohol dependence.

PubMed

Huang, San-Yuan; Lin, Wei-Wen; Wan, Fang-Jung; Chang, Ai-Ju; Ko, Huei-Chen; Wang, Tso-Jen; Wu, Pei-Lin; Lu, Ru-Band

2007-05-01

Low monoamine oxidase (MAO) activity and the neurotransmitter dopamine are 2 important factors in the development of alcohol dependence. MAO is an important enzyme associated with the metabolism of biogenic amines. Therefore, the present study investigates whether the association between the dopamine D2 receptor (DRD2) gene and alcoholism is affected by different polymorphisms of the MAO type A (MAOA) gene. A total of 427 Han Chinese men in Taiwan (201 control subjects and 226 with alcoholism) were recruited for the study. Of the subjects with alcoholism, 108 had pure alcohol dependence (ALC) and 118 had both alcohol dependence and anxiety, depression or both (ANX/DEP ALC). All subjects were assessed with the Chinese Version of the Modified Schedule of Affective Disorders and Schizophrenia-Lifetime. Alcohol dependence, anxiety and major depressive disorders were diagnosed according to Diagnostic and Statistical Manual of Mental Disorders, fourth edition criteria. The genetic variant of the DRD2 gene was only associated with the ANX/DEP ALC phenotype, and the genetic variant of the MAOA gene was associated with pure ALC. Subjects carrying the MAOA 3-repeat allele and genotype A1/A1 of the DRD2 were 3.48 times (95% confidence interval = 1.47-8.25) more likely to be ANX/DEP ALC than the subjects carrying the MAOA 3-repeat allele and DRD2 A2/A2 genotype. The MAOA gene may modify the association between the DRD2 gene and ANX/DEP ALC phenotype.

Monoamine Oxidase Deficiency Causes Prostate Atrophy and Reduces Prostate Progenitor Cell Activity.

PubMed

Yin, Lijuan; Li, Jingjing; Liao, Chun-Peng; Jason Wu, Boyang

2018-04-10

Monoamine oxidases (MAOs) degrade a number of biogenic and dietary amines, including monoamine neurotransmitters, and play an essential role in many biological processes. Neurotransmitters and related neural events have been shown to participate in the development, differentiation, and maintenance of diverse tissues and organs by regulating the specialized cellular function and morphological structures of innervated organs such as the prostate. Here we show that mice lacking both MAO isoforms, MAOA and MAOB, exhibit smaller prostate mass and develop epithelial atrophy in the ventral and dorsolateral prostates. The cellular composition of prostate epithelium showed reduced CK5 + or p63 + basal cells, accompanied by lower Sca-1 expression in p63 + basal cells, but intact differentiated CK8 + luminal cells in MAOA/B-deficient mouse prostates. MAOA/B ablation also decreased epithelial cell proliferation without affecting cell apoptosis in mouse prostates. Using a human prostate epithelial cell line, we found that stable knockdown of MAOA and MAOB impaired the capacity of prostate stem cells to form spheres, coinciding with a reduced CD133 + /CD44 + /CD24 - stem cell population and less expression of CK5 and select stem cell markers, including ALDH1A1, TROP2, and CD166. Alternative pharmacological inhibition of MAOs also repressed prostate cell stemness. In addition, we found elevated expression of MAOA and MAOB in epithelial and/or stromal components of human prostate hyperplasia samples compared with normal prostate tissues. Taken together, our findings reveal critical roles for MAOs in the regulation of prostate basal progenitor cells and prostate maintenance. Stem Cells 2018. © AlphaMed Press 2018.

Safinamide for symptoms of Parkinson's disease.

PubMed

Müller, T

2015-11-01

Chronic and slow progression of neuronal death in Parkinson's disease is responsible for an altered neurotransmission of various biogenic amines, such as dopamine. Therefore, an individually different pronounced heterogeneity of motor and nonmotor symptoms characterizes each Parkinson's disease patient. Ideal candidates for the balance of these neurotransmitter deficits are compounds like safinamide with broad mechanisms of action such as reversible monoamine oxidase type B inhibition, blockage of voltage-dependent sodium channels, modulation of calcium channels and of glutamate release. Safinamide is administered one time daily with oral doses ranging from 50 to 100 mg. Safinamide was well tolerated and safe, ameliorated motor symptoms when combined with dopamine agonist only or additional levodopa in clinical trials. Safinamide is a novel instrument for the drug therapy of Parkinson's disease with better safety and tolerability particularly concerning diarrhea than inhibitors of catechol-O-methyltransferase, like entacapone, according to an indirect comparison within a meta-analysis with entacapone. Copyright 2015 Prous Science, S.A.U. or its licensors. All rights reserved.

Monoamine oxidase-B inhibitors in the treatment of Parkinson's disease: clinical-pharmacological aspects.

PubMed

Riederer, Peter; Müller, Thomas

2018-03-22

This invited narrative review emphasizes the role of MAO-B inhibition in the drug portfolio for dopamine substitution in patients with Parkinson's disease. Neuronal and glial MAO-B inhibition contributes to more stable levels of dopamine and other biogenic amines in the synaptic cleft. Accordingly, symptomatic effects of MAO-B inhibition for a limited amelioration of impaired motor behaviour and wearing-off phenomena in patients with Parkinson's disease are well proven, even when MAO-B inhibitors are only applied together with dopamine agonists. Delay of disease progression by MAO-B inhibition is under debate despite positive experimental findings. This discussion does not consider, that levodopa, respectively, dopamine agonists, are substrates, respectively, inhibitors of the ABCB1 (P-gp, MDR1, and CD243) transporter system. It supports toxin efflux over the blood-brain barrier. ABCB1 transporters have a limited capacity. MAO-B inhibitors do not weaken it. Treatment with MAO-B inhibitors is advantageous as it enables sparing of dopamine agonist and levodopa dosing.

Function-blocking antibodies to human vascular adhesion protein-1: a potential anti-inflammatory therapy.

PubMed

Kirton, Christopher M; Laukkanen, Marja-Leena; Nieminen, Antti; Merinen, Marika; Stolen, Craig M; Armour, Kathryn; Smith, David J; Salmi, Marko; Jalkanen, Sirpa; Clark, Michael R

2005-11-01

Human vascular adhesion protein-1 (VAP-1) is a homodimeric 170-kDa sialoglycoprotein that is expressed on the surface of endothelial cells and functions as a semicarbazide-sensitive amine oxidase and as an adhesion molecule. Blockade of VAP-1 has been shown to reduce leukocyte adhesion and transmigration in in vivo and in vitro models, suggesting that VAP-1 is a potential target for anti-inflammatory therapy. In this study we have constructed mouse-human chimeric antibodies by genetic engineering in order to circumvent the potential problems involved in using murine antibodies in man. Our chimeric anti-VAP-1 antibodies, which were designed to lack Fc-dependent effector functions, bound specifically to cell surface-expressed recombinant human VAP-1 and recognized VAP-1 in different cell types in tonsil. Furthermore, the chimeric antibodies prevented leukocyte adhesion and transmigration in vitro and in vivo. Hence, these chimeric antibodies have the potential to be used as a new anti-inflammatory therapy.

Amino Acid Isomerization in the Production of l-Phenylalanine from d-Phenylalanine by Bacteria1

PubMed Central

Chibata, Ichiro; Tosa, Tetsuya; Sano, Ryujiro

1965-01-01

To establish an advantageous method for the production of l-amino acids, microbial isomerization of d- and dl-amino acids to l-amino acids was studied. Screening experiments on a number of microorganisms showed that cell suspensions of Pseudomonas fluorescens and P. miyamizu were capable of isomerizing d- and dl-phenylalanines to l-phenylalanine. Various conditions suitable for isomerization by these organisms were investigated. Cells grown in a medium containing d-phenylalanine showed highest isomerization activity, and almost completely converted d- or dl-phenylalanine into l-phenylalanine within 24 to 48 hr of incubation. Enzymatic studies on this isomerizing system suggested that the isomerization of d- or dl-phenylalanine is not catalyzed by a single enzyme, “amino acid isomerase,” but the conversion proceeds by a two step system as follows: d-pheylalanine is oxidized to phenylpyruvic acid by d-amino acid oxidase, and the acid is converted to l-phenylalanine by transamination or reductive amination. PMID:14339270

Interaction between vitamin B6 metabolism, nitrogen metabolism and autoimmunity.

PubMed

Colinas, Maite; Fitzpatrick, Teresa B

2016-01-01

The essential micronutrient vitamin B6 is best known in its enzymatic cofactor form, pyridoxal 5'-phosphate (PLP). However, vitamin B6 comprises the amine pyridoxamine 5'-phosphate (PMP) and the alcohol pyridoxine 5'-phosphate (PNP) in addition to PLP, as well as their corresponding non-phosphorylated forms. The different B6 forms (called vitamers) are enzymatically interconverted in a ubiquitous salvage pathway. Recently, we have shown that balancing the ratio of the different B6 vitamers in particular PMP by the PMP/PNP oxidase PDX3 is essential for growth and development in Arabidopsis thaliana. Intriguingly, nitrate to ammonium conversion is impaired in pdx3 mutants, such that the mutants become ammonium-dependent, suggesting an interaction between vitamin B6 and nitrogen metabolism. In addition, we found a strong up-regulation of genes related to plant defense. Here, we further show that pdx3 mutants display a temperature-sensitive phenotype that is typical of autoimmune mutants and is possibly connected to the impaired nitrogen metabolism.

The Arabidopsis SKU5 gene encodes an extracellular glycosyl phosphatidylinositol-anchored glycoprotein involved in directional root growth

NASA Technical Reports Server (NTRS)

Sedbrook, John C.; Carroll, Kathleen L.; Hung, Kai F.; Masson, Patrick H.; Somerville, Chris R.

2002-01-01

To investigate how roots respond to directional cues, we characterized a T-DNA-tagged Arabidopsis mutant named sku5 in which the roots skewed and looped away from the normal downward direction of growth on inclined agar surfaces. sku5 roots and etiolated hypocotyls were slightly shorter than normal and exhibited a counterclockwise (left-handed) axial rotation bias. The surface-dependent skewing phenotype disappeared when the roots penetrated the agar surface, but the axial rotation defect persisted, revealing that these two directional growth processes are separable. The SKU5 gene belongs to a 19-member gene family designated SKS (SKU5 Similar) that is related structurally to the multiple-copper oxidases ascorbate oxidase and laccase. However, the SKS proteins lack several of the conserved copper binding motifs characteristic of copper oxidases, and no enzymatic function could be assigned to the SKU5 protein. Analysis of plants expressing SKU5 reporter constructs and protein gel blot analysis showed that SKU5 was expressed most strongly in expanding tissues. SKU5 was glycosylated and modified by glycosyl phosphatidylinositol and localized to both the plasma membrane and the cell wall. Our observations suggest that SKU5 affects two directional growth processes, possibly by participating in cell wall expansion.

Proton-coupled electron transfer and the role of water molecules in proton pumping by cytochrome c oxidase

PubMed Central

Sharma, Vivek; Enkavi, Giray; Vattulainen, Ilpo; Róg, Tomasz; Wikström, Mårten

2015-01-01

Molecular oxygen acts as the terminal electron sink in the respiratory chains of aerobic organisms. Cytochrome c oxidase in the inner membrane of mitochondria and the plasma membrane of bacteria catalyzes the reduction of oxygen to water, and couples the free energy of the reaction to proton pumping across the membrane. The proton-pumping activity contributes to the proton electrochemical gradient, which drives the synthesis of ATP. Based on kinetic experiments on the O–O bond splitting transition of the catalytic cycle (A → PR), it has been proposed that the electron transfer to the binuclear iron–copper center of O2 reduction initiates the proton pump mechanism. This key electron transfer event is coupled to an internal proton transfer from a conserved glutamic acid to the proton-loading site of the pump. However, the proton may instead be transferred to the binuclear center to complete the oxygen reduction chemistry, which would constitute a short-circuit. Based on atomistic molecular dynamics simulations of cytochrome c oxidase in an explicit membrane–solvent environment, complemented by related free-energy calculations, we propose that this short-circuit is effectively prevented by a redox-state–dependent organization of water molecules within the protein structure that gates the proton transfer pathway. PMID:25646428

NADPH oxidase activation contributes to native low-density lipoprotein-induced proliferation of human aortic smooth muscle cells.

PubMed

Park, Il Hwan; Hwang, Hye Mi; Jeon, Byeong Hwa; Kwon, Hyung-Joo; Hoe, Kwang Lae; Kim, Young Myeong; Ryoo, Sungwoo

2015-06-12

Elevated plasma concentration of native low-density lipoprotein (nLDL) is associated with vascular smooth muscle cell (VSMC) activation and cardiovascular disease. We investigated the mechanisms of superoxide generation and its contribution to pathophysiological cell proliferation in response to nLDL stimulation. Lucigenin-induced chemiluminescence was used to measure nLDL-induced superoxide production in human aortic smooth muscle cells (hAoSMCs). Superoxide production was increased by nicotinamide adenine dinucleotide phosphate (NADPH) and decreased by NADPH oxidase inhibitors in nLDL-stimulated hAoSMC and hAoSMC homogenates, as well as in prepared membrane fractions. Extracellular signal-regulated kinase 1/2 (Erk1/2), protein kinase C-θ (PKCθ) and protein kinase C-β (PKCβ) were phosphorylated and maximally activated within 3 min of nLDL stimulation. Phosphorylated Erk1/2 mitogen-activated protein kinase, PKCθ and PKCβ stimulated interactions between p47phox and p22phox; these interactions were prevented by MEK and PKC inhibitors (PD98059 and calphostin C, respectively). These inhibitors decreased nLDL-dependent superoxide production and blocked translocation of p47phox to the membrane, as shown by epifluorescence imaging and cellular fractionation experiments. Proliferation assays showed that a small interfering RNA against p47phox, as well as superoxide scavenger and NADPH oxidase inhibitors, blocked nLDL-induced hAoSMC proliferation. The nLDL stimulation in deendothelialized aortic rings from C57BL/6J mice increased dihydroethidine fluorescence and induced p47phox translocation that was blocked by PD98059 or calphostin C. Isolated aortic SMCs from p47phox(-/-) mice (mAoSMCs) did not respond to nLDL stimulation. Furthermore, NADPH oxidase 1 (Nox1) was responsible for superoxide generation and cell proliferation in nLDL-stimulated hAoSMCs. These data demonstrated that NADPH oxidase activation contributed to cell proliferation in nLDL-stimulated hAoSMCs.

Biodegradation and detoxification of textile azo dyes by bacterial consortium under sequential microaerophilic/aerobic processes

PubMed Central

Lade, Harshad; Kadam, Avinash; Paul, Diby; Govindwar, Sanjay

2015-01-01

Release of textile azo dyes to the environment is an issue of health concern while the use of microorganisms has proved to be the best option for remediation. Thus, in the present study, a bacterial consortium consisting of Providencia rettgeri strain HSL1 and Pseudomonas sp. SUK1 has been investigated for degradation and detoxification of structurally different azo dyes. The consortium showed 98-99 % decolorization of all the selected azo dyes viz. Reactive Black 5 (RB 5), Reactive Orange 16 (RO 16), Disperse Red 78 (DR 78) and Direct Red 81 (DR 81) within 12 to 30 h at 100 mg L-1 concentration at 30 ± 0.2 °C under microaerophilic, sequential aerobic/microaerophilic and microaerophilic/aerobic processes. However, decolorization under microaerophilic conditions viz. RB 5 (0.26 mM), RO 16 (0.18 mM), DR 78 (0.20 mM) and DR 81 (0.23 mM) and sequential aerobic/microaerophilic processes viz. RB 5 (0.08 mM), RO 16 (0.06 mM), DR 78 (0.07 mM) and DR 81 (0.09 mM) resulted into the formation of aromatic amines. In distinction, sequential microaerophilic/ aerobic process doesn’t show the formation of amines. Additionally, 62-72 % reduction in total organic carbon content was observed in all the dyes decolorized broths under sequential microaerophilic/aerobic processes suggesting the efficacy of method in mineralization of dyes. Notable induction within the levels of azoreductase and NADH-DCIP reductase (97 and 229 % for RB 5, 55 and 160 % for RO 16, 63 and 196 % for DR 78, 108 and 258 % for DR 81) observed under sequential microaerophilic/aerobic processes suggested their critical involvements in the initial breakdown of azo bonds, whereas, a slight increase in the levels of laccase and veratryl alcohol oxidase confirmed subsequent oxidation of formed amines. Also, the acute toxicity assay with Daphnia magna revealed the nontoxic nature of the dye-degraded metabolites under sequential microaerophilic/aerobic processes. As biodegradation under sequential microaerophilic/aerobic process completely detoxified all the selected textile azo dyes, further efforts should be made to implement such methods for large scale dye wastewater treatment technologies. PMID:26417357

Reversible effects of sphingomyelin degradation on cholesterol distribution and metabolism in fibroblasts and transformed neuroblastoma cells.

PubMed Central

Pörn, M I; Slotte, J P

1990-01-01

Plasma-membrane sphingomyelin appears to be one of the major determinants of the preferential allocation of cell cholesterol into the plasma-membrane compartment, since removal of sphingomyelin leads to a dramatic redistribution of cholesterol within the cell [Slotte & Bierman (1988) Biochem. J. 250, 653-658]. In the present study we examined the long-term effects of sphingomyelin degradation on cholesterol redistribution in cells and determined the reversibility of the process. In a human lung fibroblast-cell line, removal of 80% of the sphingomyelin led to a rapid and transient up-regulation (3-fold) of acyl-CoA:cholesterol acyltransferase (ACAT) activity, and also, within 30 h, to the translocation of about 50% of the cell non-esterified cholesterol from a cholesterol oxidase-susceptible compartment (i.e. the cell surface) to oxidase-resistant compartments. At 49 h after the initial sphingomyelin degradation, the cell sphingomyelin level was back to 45% of the control level, and the direction of cell cholesterol flow was toward the cell surface, although the original distribution was not achieved. In a transformed neuroblastoma cell line (SH-SY5Y), the depletion of sphingomyelin led to a similarly rapid and transient up-regulation of ACAT activity, and to the translocation of about 25% of cell-surface cholesterol into internal membranes (within 3 h). The flow of cholesterol back to the cholesterol oxidase-susceptible pool was rapid, and a pretreatment cholesterol distribution was reached within 20-49 h. Also, the resynthesis of sphingomyelin was faster in SH-SY5Y neuroblastoma cells and reached control levels within 24 h. The findings of the present study show that the cellular redistribution of cholesterol, as induced by sphingomyelin degradation, is reversible and suggest that the normalization of cellular cholesterol distribution is linked to the re-synthesis of sphingomyelin. PMID:2222406

The Use of Multiscale Molecular Simulations in Understanding a Relationship between the Structure and Function of Biological Systems of the Brain: The Application to Monoamine Oxidase Enzymes

PubMed Central

Vianello, Robert; Domene, Carmen; Mavri, Janez

2016-01-01

HIGHLIGHTS Computational techniques provide accurate descriptions of the structure and dynamics of biological systems, contributing to their understanding at an atomic level.Classical MD simulations are a precious computational tool for the processes where no chemical reactions take place.QM calculations provide valuable information about the enzyme activity, being able to distinguish among several mechanistic pathways, provided a carefully selected cluster model of the enzyme is considered.Multiscale QM/MM simulation is the method of choice for the computational treatment of enzyme reactions offering quantitative agreement with experimentally determined reaction parameters.Molecular simulation provide insight into the mechanism of both the catalytic activity and inhibition of monoamine oxidases, thus aiding in the rational design of their inhibitors that are all employed and antidepressants and antiparkinsonian drugs. Aging society and therewith associated neurodegenerative and neuropsychiatric diseases, including depression, Alzheimer's disease, obsessive disorders, and Parkinson's disease, urgently require novel drug candidates. Targets include monoamine oxidases A and B (MAOs), acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and various receptors and transporters. For rational drug design it is particularly important to combine experimental synthetic, kinetic, toxicological, and pharmacological information with structural and computational work. This paper describes the application of various modern computational biochemistry methods in order to improve the understanding of a relationship between the structure and function of large biological systems including ion channels, transporters, receptors, and metabolic enzymes. The methods covered stem from classical molecular dynamics simulations to understand the physical basis and the time evolution of the structures, to combined QM, and QM/MM approaches to probe the chemical mechanisms of enzymatic activities and their inhibition. As an illustrative example, the later will focus on the monoamine oxidase family of enzymes, which catalyze the degradation of amine neurotransmitters in various parts of the brain, the imbalance of which is associated with the development and progression of a range of neurodegenerative disorders. Inhibitors that act mainly on MAO A are used in the treatment of depression, due to their ability to raise serotonin concentrations, while MAO B inhibitors decrease dopamine degradation and improve motor control in patients with Parkinson disease. Our results give strong support that both MAO isoforms, A and B, operate through the hydride transfer mechanism. Relevance of MAO catalyzed reactions and MAO inhibition in the context of neurodegeneration will be discussed. PMID:27471444

Co-expression of cytochrome b561 and ascorbate oxidase in leaves of wild watermelon under drought and high light conditions.

PubMed

Nanasato, Yoshihiko; Akashi, Kinya; Yokota, Akiho

2005-09-01

Despite carrying out C3 photosynthesis, wild watermelon (Citrullus lanatus sp.) exhibits exceedingly good tolerance to severe drought at high light intensities. However, the mechanism(s) by which this plant protects itself from photodamage has yet to be elucidated. In this study, we characterized wild watermelon cytochrome b561 (cyt b561), which potentially mediates regeneration of apoplastic ascorbate by transferring electrons from cytosolic ascorbate across the plasma membrane. Two cDNA species for wild watermelon cyt b561, designated CLb561A and CLb561B, were isolated. Levels of both CLb561A mRNA and protein were significantly elevated in the leaves during drought at a light intensity of 700 micromol photons m(-2) s(-1). The transcript of CLb561B was detected to a much lesser extent, but no CLb561B protein was produced under any condition used in this study. A transient expression assay with the CLb561A::green fluorescent protein fusion construct showed clear fluorescence on the plasma membrane of onion epidermal cells. The CLb561A protein was enriched in the plasma membrane fraction in leaves of transgenic tobacco expressing CLb561A. Moreover, the high activity of apoplastic ascorbate oxidase (AO), which was able to dispose of cyt b561-transferred reducing equivalents, increased in leaves of wild watermelon grown at high light intensity, but not lower light intensities. Taken together, these observations suggest the occurrence of a novel pathway for excess light energy dissipation in wild watermelon leaves, where excessive energy absorbed by chloroplasts can be transported to and dissipated safely in the apoplasts through the cooperative action of cyt b561 and AO.

The plasma membrane-associated NADH oxidase of spinach leaves responds to blue light

NASA Technical Reports Server (NTRS)

Morre, D. James; Penel, Claude; Greppin, Hubert; Morre, Dorothy M.

2002-01-01

The plasma membrane-associated NADH oxidase (NOX) of spinach leaf disks is characterized by oscillations in activity with a regular period length of ca. 24 min. Within a single population of plants exposed to light at the same time, NOX activities of all plants function synchronously. Exposure of plants transferred from darkness to blue light (495 nm, 2 min, 50 micromoles m-2 s-1) resulted in a complex response pattern but with a new maximum in the rate of NOX activity 36 (24+12) min after illumination and then with maxima in the rate of NOX activity every 24 min thereafter. Transient maxima in NOX activity were observed as well after 9.3 + /- 1.4 and 20.7 +/- 2.1 min. The blue light response differed from the response to red (650 nm, 10 min, 50 micromoles m-2 s-1) or white light where activity maxima were initiated 12 min after the light exposure followed by maxima every 24 min thereafter. Green or yellow light was ineffective. The light response was independent of the time in the 24-min NOX cycle when the light was given. The net effects of blue and red light were ultimately the same with a new maximum in the rate of NOX activity at 12+24=36 min (and every 24 min thereafter), but the mechanisms appear to be distinct.

Bacopa monniera leaf extract ameliorates hypobaric hypoxia induced spatial memory impairment.

PubMed

Hota, Sunil Kumar; Barhwal, Kalpana; Baitharu, Iswar; Prasad, Dipti; Singh, Shashi Bala; Ilavazhagan, Govindasamy

2009-04-01

Hypobaric hypoxia induced memory impairment has been attributed to several factors including increased oxidative stress, depleted mitochondrial bioenergetics, altered neurotransmission and apoptosis. This multifactorial response of the brain to hypobaric hypoxia limits the use of therapeutic agents that target individual pathways for ameliorating hypobaric hypoxia induced memory impairment. The present study aimed at exploring the therapeutic potential of a bacoside rich leaf extract of Bacopa monniera in improving the memory functions in hypobaric conditions. The learning ability was evaluated in male Sprague Dawley rats along with memory retrieval following exposure to hypobaric conditions simulating an altitude of 25,000 ft for different durations. The effect of bacoside administration on apoptosis, cytochrome c oxidase activity, ATP levels, and oxidative stress markers and on plasma corticosterone levels was investigated. Expression of NR1 subunit of N-methyl-d-aspartate receptors, neuronal cell adhesion molecules and was also studied along with CREB phosphorylation to elucidate the molecular mechanisms of bacoside action. Bacoside administration was seen to enhance learning ability in rats along with augmentation in memory retrieval and prevention of dendritic atrophy following hypoxic exposure. In addition, it decreased oxidative stress, plasma corticosterone levels and neuronal degeneration. Bacoside administration also increased cytochrome c oxidase activity along with a concomitant increase in ATP levels. Hence, administration of bacosides could be a useful therapeutic strategy in ameliorating hypobaric hypoxia induced cognitive dysfunctions and other related neurological disorders.

Silymarin alleviates hepatic oxidative stress and protects against metabolic disorders in high-fat diet-fed mice.

PubMed

Feng, Bin; Meng, Ran; Huang, Bin; Shen, Shanmei; Bi, Yan; Zhu, Dalong

2016-01-01

Silymarin is a potent antioxidant medicine and has been widely used for the treatment of liver diseases over 30 years. Recent studies suggest that silymarin may benefit patients with glucose intolerance. However, the mechanism underlying the action of silymarin is not clarified. The aim of this work was to assess the impact of silymarin on glucose intolerance in high-fat diet (HFD)-fed mice, and explore the potential therapeutic mechanisms. C57BL/6 mice were fed with HFD for 12 weeks, randomized, and treated orally with vehicle saline or silymarin (30 mg/kg) daily for 30 days. We found that silymarin significantly improved HFD-induced body weight gain, glucose intolerance, and insulin resistance in mice. Silymarin treatment reduced HFD-increased oxidative stress indicators (reactive oxygen species, lipid peroxidation, protein oxidation) and restored HFD-down-regulated activities of antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase) in the plasma and/or liver of the HFD-fed mice. Furthermore, silymarin decreased HFD-up-regulated hepatic NADPH oxidase expression and NF-κB activation in mice. Additionally, silymarin treatment mitigated HFD-increased plasma IL-1β, TNF-α levels, and HFD-enhanced hepatic NO, TLR4, and iNOS expression in mice. These novel data indicate that silymarin has potent anti-diabetic actions through alleviating oxidative stress and inflammatory response, partially by inhibiting hepatic NADPH oxidase expression and the NF-κB signaling.

Oral administration of L-arginine in patients with angina or following myocardial infarction may be protective by increasing plasma superoxide dismutase and total thiols with reduction in serum cholesterol and xanthine oxidase

PubMed Central

Tripathi, Pratima; Chandra, M

2009-01-01

Administration of L-arginine has been shown to control ischemic injury by producing nitric oxide which dilates the vessels and thus maintains proper blood flow to the myocardium. In the present study attempt has been made to determine whether oral administration of L-arginine has any effect on oxidant/antioxidant homeostasis in ischemic myocardial patients [represented by the patients of acute angina (AA) and acute myocardial infarction (MI)]. L-arginine has antioxidant and antiapoptotic properties, decreases endothelin-1 expression and improves endothelial function, thereby controlling oxidative injury caused during myocardial ischemic syndrome. Effect of L-arginine administration on the status of free radical scavenging enzymes, pro-oxidant enzyme and antioxidants viz. total thiols, carbonyl content and plasma ascorbic acid levels in the patients has been evaluated. We have observed that L-arginine administration (three grams per day for 15 days) resulted in increased activity of free radical scavenging enzyme superoxide dismutase (SOD) and increase in the levels of total thiols (T-SH) and ascorbic acid with concomitant decrease in lipid per-oxidation, carbonyl content, serum cholesterol and the activity of proxidant enzyme, xanthine oxidase (XO). These findings suggest that the supplementation of L-arginine along with regular therapy may be beneficial to the patients of ischemic myocardial syndromes. PMID:20716909

In vitro metabolism and interactions of pyridostigmine bromide, N,N-diethyl-m-toluamide, and permethrin in human plasma and liver microsomal enzymes.

PubMed

Abu-Qare, A W; Abou-Donia, M B

2008-03-01

1. The in vitro human plasma activity and liver microsomal metabolism of pyridostigmine bromide (PB), a prophylactic treatment against organophosphate nerve agent attack, N,N-diethyl-m-toluamide (DEET), an insect repellent, and permethrin, a pyrethroid insecticide, either alone or in combination were investigated. 2. The three chemicals disappeared from plasma in the following order: permethrin > PB > DEET. The combined incubation of DEET with either permethrin or PB had no effect on permethrin or PB. Binary incubation with permethrin decreased the metabolism of PB and its disappearance from plasma and binary incubation with PB decreased the metabolism of permethrin and its clearance from plasma. Incubation with PB and/or permethrin shortened the DEET terminal half-life in plasma. These agents behaved similarly when studied in liver microsomal assays. The combined incubation of DEET with PB or permethrin (alone or in combination) diminished DEET metabolism in microsomal systems. 3. The present study evidences that PB and permethrin are metabolized by both human plasma and liver microsomal enzymes and that DEET is mainly metabolized by liver oxidase enzymes. Combined exposure to test chemicals increases their neurotoxicity by impeding the body's ability to eliminate them because of the competition for detoxifying enzymes.

Noise as a Public Health Problem: Proceedings of the International Congress (4th) Held at Turin (Italy) on June 21-25, 1983. Volume 1.

DTIC Science & Technology

1983-11-01

22 , 821-825. Peters, D.A.V., Prenatal stress: effects on brain biogenic amine and plasma corticosterone levels. Pharmacol., Biochem. and Behav., 17...527 Late Postnatal Effect on Corticosterone Blood Level, of an Acoustical Stimulation Previously Applied to Mice During Pre or Early Postnatal...evaluated threshold shifts and/or hair cell destruction as indicatozs of damage. The aim has particularly been focused on the effect of systema- tically

Endocrine and Metabolic Changes during Exhaustive Multifactorial Military Stress. Results from Studies during the Ranger Training Course of the Norwegian Military Academy

DTIC Science & Technology

2001-03-01

biochimiques. Moyens possibles de recuperation rapide.] To order the complete compilation report, use : ADA389235 The component part is provided here to...heparin, or also substances necessary to conserve the hormones investigated, such as the peptidase inhibitor aprotinin. The blood for preparation of...plasma were analyzed using a radioenzymatic method (s-adenosyl methionin as a tritiated methyl donor transferred to the amines by catechol-o-methyl

Synthesis and Preclinical Evaluation of QS-21 Variants Leading to Simplified Vaccine Adjuvants and Mechanistic Probes

PubMed Central

Chea, Eric K.; Fernández-Tejada, Alberto; Damani, Payal; Adams, Michelle M.; Gardner, Jeffrey R.; Livingston, Philip O.; Ragupathi, Govind; Gin, David Y.

2012-01-01

QS-21 is a potent immunostimulatory saponin that is currently under clinical investigation as an adjuvant in various vaccines to treat infectious diseases, cancers, and congnitive disorders. Herein we report the design, synthesis, and preclinical evaluation of simplified QS-21 congeners to define key structural features that are critical for adjuvant activity. Truncation of the linear tetrasaccharide domain revealed that a trisaccharide variant is equipotent to QS-21 while the corresponding disaccharide and monosaccharide congeners are more toxic or less potent, respectively. Modification of the acyl domain in the trisaccharide series revealed that a terminal carboxylic acid is well-tolerated while a terminal amine results in reduced adjuvant activity. Acylation of the terminal amine can restore adjuvant activity and enables the synthesis of fluorescently-labeled QS-21 variants. Cellular studies with these probes revealed that, contrary to conventional wisdom, the most highly adjuvant active of these fluorescently-labeled saponins does not simply associate with the plasma membrane, but rather is internalized by dendritic cells. PMID:22866694

Amine-Rich Organic Thin Films for Cell Culture: Possible Electrostatic Effects in Cell-Surface Interactions

NASA Astrophysics Data System (ADS)

Wertheimer, Michael R.; St-Georges-Robillard, Amélie; Lerouge, Sophie; Mwale, Fackson; Elkin, Bentsian; Oehr, Christian; Wirges, Werner; Gerhard, Reimund

2012-11-01

In recent communications from these laboratories, we observed that amine-rich thin organic layers are very efficient surfaces for the adhesion of mammalian cells. We prepare such deposits by plasma polymerization at low pressure, atmospheric pressure, or by vacuum-ultraviolet photo-polymerization. More recently, we have also investigated a commercially available material, Parylene diX AM. In this article we first briefly introduce literature relating to electrostatic interactions between cells, proteins, and charged surfaces. We then present certain selected cell-response results that pertain to applications in orthopedic and cardiovascular medicine: we discuss the influence of surface properties on the observed behaviors of two particular cell lines, human U937 monocytes, and Chinese hamster ovary cells. Particular emphasis is placed on possible electrostatic attractive forces due to positively charged R-NH3+ groups and negatively charged proteins and cells, respectively. Experiments carried out with electrets, polymers with high positive or negative surface potentials are added for comparison.

Amine-functionalized PVA-co-PE nanofibrous membrane as affinity membrane with high adsorption capacity for bilirubin.

PubMed

Wang, Wenwen; Zhang, Hao; Zhang, Zhifeng; Luo, Mengying; Wang, Yuedan; Liu, Qiongzhen; Chen, Yuanli; Li, Mufang; Wang, Dong

2017-02-01

In this study, poly(vinyl alcohol-co-ethylene) (PVA-co-PE) nanofibrous membrane was activated by sodium hydroxide and cyanuric chloride, and then the activated membranes were functionalized by 1,3-propanediamine, hexamethylenediamine and diethylenetriamine to be affinity membranes for bilirubin removal, respectively. The chemical structures and morphologies of membranes were investigated by SEM, FTIR and XPS. And the adsorption ability of different amine-functionalized nanofibrous membranes for bilirubin was characterized. Furthermore, the effects of temperature, initial concentration of bilirubin, NaCl concentration and BSA concentration on the adsorption capacity for bilirubin of diethylenetriamine-functionalized nanofibrous membrane were studied. Results indicated that the adsorption capacity for bilirubin of diethylenetriamine-functionalized nanofibrous membrane could reach 85mg/g membrane when the initial bilirubin concentration was 200mg/L while the adsorption capacity could be increased to 110mg/g membrane if the initial bilirubin concentration was more than 400mg/L. The dynamic adsorption of diethylenetriamine-functionalized nanofibrous membrane showed that the ligands of amine groups on the membrane surface could be used as far as possible by recirculating the plasma with certain flow rates. Therefore, the diethylenetriamine-functionalized PVA-co-PE nanofibrous membrane possessed high adsorption capacity for bilirubin and it can be candidate as affinity membrane for bilirubin removal. Copyright © 2016. Published by Elsevier B.V.

NecroX-7 prevents oxidative stress-induced cardiomyopathy by inhibition of NADPH oxidase activity in rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Joonghoon; Park, Eok; Ahn, Bong-Hyun

2012-08-15

Oxidative stress is one of the causes of cardiomyopathy. In the present study, NecroXs, novel class of mitochondrial ROS/RNS scavengers, were evaluated for cardioprotection in in vitro and in vivo model, and the putative mechanism of the cardioprotection of NecroX-7 was investigated by global gene expression profiling and subsequent biochemical analysis. NecroX-7 prevented tert-butyl hydroperoxide (tBHP)-induced death of H9C2 rat cardiomyocytes at EC{sub 50} = 0.057 μM. In doxorubicin (DOX)-induced cardiomyopathy in rats, NecroX-7 significantly reduced the plasma levels of creatine kinase (CK-MB) and lactate dehydrogenase (LDH) which were increased by DOX treatment (p < 0.05). Microarray analysis revealed thatmore » 21 genes differentially expressed in tBHP-treated H9C2 cells were involved in ‘Production of reactive oxygen species’ (p = 0.022), and they were resolved by concurrent NecroX-7 treatment. Gene-to-gene networking also identified that NecroX-7 relieved cell death through Ncf1/p47phox and Rac2 modulation. In subsequent biochemical analysis, NecroX-7 inhibited NADPH oxidase (NOX) activity by 53.3% (p < 0.001). These findings demonstrate that NecroX-7, in part, provides substantial protection of cardiomyopathy induced by tBHP or DOX via NOX-mediated cell death. -- Highlights: ► NecroX-7 prevented tert-butyl hydroperoxide-induced in vitro cardiac cell death. ► NecroX-7 ameliorated doxorubicin-induced in vivo cardiomyopathy. ► NecroX-7 prevented oxidative stress and necrosis-enriched transcriptional changes. ► NecroX-7 effectively inhibited NADPH oxidase activation. ► Cardioprotection of Necro-7 was brought on by modulation of NADPH oxidase activity.« less

Immobilization of biomolecules to plasma polymerized pentafluorophenyl methacrylate.

PubMed

Duque, Luis; Menges, Bernhard; Borros, Salvador; Förch, Renate

2010-10-11

Thin films of plasma polymerized pentafluorophenyl methacrylate (pp-PFM) offer highly reactive ester groups throughout the structure of the film that allow for subsequent reactions with different aminated reagents and biological molecules. The present paper follows on from previous work on the plasma deposition of pentafluorophenyl methacrylate (PFM) for optimum functional group retention (Francesch, L.; Borros, S.; Knoll, W.; Foerch, R. Langmuir 2007, 23, 3927) and reactivity in aqueous solution (Duque, L.; Queralto, N.; Francesch, L.; Bumbu, G. G.; Borros, S.; Berger, R.; Förch, R. Plasma Process. Polym. 2010, accepted for publication) to investigate the binding of a biologically active peptide known to induce cellular adhesion (IKVAV) and of biochemically active proteins such as BSA and fibrinogen. Analyses of the films and of the immobilization of the biomolecules were carried out using infrared reflection absorption spectroscopy (IRRAS), X-ray photoelectron spectroscopy (XPS), and atomic force microscopy (AFM). The attachment of the biomolecules on pulsed plasma polymerized pentafluorophenyl methacrylate was monitored using surface plasmon resonance spectroscopy (SPR). SPR analysis confirmed the presence of immobilized biomolecules on the plasma polymer and was used to determine the mass coverage of the peptide and proteins adsorbed onto the films. The combined analysis of the surfaces suggests the covalent binding of the peptide and proteins to the surface of the pp-PFM.

Discovery of isatin and 1H-indazol-3-ol derivatives as d-amino acid oxidase (DAAO) inhibitors.

PubMed

Szilágyi, Bence; Kovács, Péter; Ferenczy, György G; Rácz, Anita; Németh, Krisztina; Visy, Júlia; Szabó, Pál; Ilas, Janez; Balogh, György T; Monostory, Katalin; Vincze, István; Tábi, Tamás; Szökő, Éva; Keserű, György M

2018-05-01

d-Amino acid oxidase (DAAO) is a potential target in the treatment of schizophrenia as its inhibition increases brain d-serine level and thus contributes to NMDA receptor activation. Inhibitors of DAAO were sought testing [6+5] type heterocycles and identified isatin derivatives as micromolar DAAO inhibitors. A pharmacophore and structure-activity relationship analysis of isatins and reported DAAO inhibitors led us to investigate 1H-indazol-3-ol derivatives and nanomolar inhibitors were identified. The series was further characterized by pK a and isothermal titration calorimetry measurements. Representative compounds exhibited beneficial properties in in vitro metabolic stability and PAMPA assays. 6-fluoro-1H-indazol-3-ol (37) significantly increased plasma d-serine level in an in vivo study on mice. These results show that the 1H-indazol-3-ol series represents a novel class of DAAO inhibitors with the potential to develop drug candidates. Copyright © 2018 Elsevier Ltd. All rights reserved.

Development of a Portable Blood Sugar Apparatus and GOD Enzyme Strip.

PubMed

Zhen-Cheng, Chen; Yu-Qian, Zhao; Jing-Tian, Tang; Ling-Yun, Li

2005-01-01

A pocket blood sugar apparatus tested by enzyme electrode, which was designed using carbon and silver plasma as conducting materials. Both the work and reference electrodes are applied to the parts of enzyme electrode. The glucose oxidase is taken as the medium of blood sugar measuring. And the range of measuring glucose is about 50mg/dL - 500mgl/dL. It has better linear for the results and fit coefficient arrives at 0.985. Its sensitivity of measurement is higher than current glucose biosensor obviously. A single chip microcomputer, AD mu C812, is used for central control processor of the instrument parts. It measures the output of microampere level currency, which is conduced by blood sugar reacting with the glucose oxidase on the enzyme electrode. And at the same time, the microampere level currency is amplified, processed. Then the results are displayed on LCD. The apparatus are better for measuring blood sugar, and the results are consistent with what the large biochemical instruments get.

Quantitation of glycerophosphorylcholine by flow injection analysis using immobilized enzymes.

PubMed

Mancini, A; Del Rosso, F; Roberti, R; Caligiana, P; Vecchini, A; Binaglia, L

1996-09-20

A method for quantitating glycerophosphorylcholine by flow injection analysis is reported in the present paper. Glycerophosphorylcholine phosphodiesterase and choline oxidase, immobilized on controlled porosity glass beads, are packed in a small reactor inserted in a flow injection manifold. When samples containing glycerophosphorylcholine are injected, glycerophosphorylcholine is hydrolyzed into choline and sn-glycerol-3-phosphate. The free choline produced in this reaction is oxidized to betain and hydrogen peroxide. Hydrogen peroxide is detected amperometrically. Quantitation of glycerophosphorylcholine in samples containing choline and phosphorylcholine is obtained inserting ahead of the reactor a small column packed with a mixed bed ion exchange resin. The time needed for each determination does not exceed one minute. The present method, applied to quantitate glycerophosphorylcholine in samples of seminal plasma, gave results comparable with those obtained using the standard enzymatic-spectrophotometric procedure. An alternative procedure, making use of co-immobilized glycerophosphorylcholine phosphodiesterase and glycerol-3-phosphate oxidase for quantitating glycerophosphorylcholine, glycerophosphorylethanolamine and glycerophosphorylserine is also described.

Cataplexy and monoamine oxidase deficiency in Norrie disease.

PubMed

Vossler, D G; Wyler, A R; Wilkus, R J; Gardner-Walker, G; Vlcek, B W

1996-05-01

Norrie disease (ND) is an X-linked recessive disorder causing ocular atrophy, mental retardation, deafness, and dysmorphic features. Virtually absent monoamine oxidase (MAO) type-A and -B activity has been found in some boys with chromosome deletions. We report the coexistence of cataplexy and abnormal REM sleep organization with ND. Three related boys, referred for treatment of medically refractory atonic spells and apneas, underwent extended EEG-video-polysomnographic monitoring. They demonstrated attacks of cataplexy and inappropriate periods of REM sleep during which they were unarousable. One boy also had generalized tonic-clonic seizures. Previous testing revealed that all three have complete ND gene deletions. In all subjects, platelet MAO-B activity was absent, serum serotonin levels were markedly increased, and plasma catecholamine levels were normal. Data from the canine narcolepsy syndrome model implicate abnormal catecholaminergic and cholinergic activities in the pathogenesis of cataplexy. Our findings suggest that abnormal MAO activity or an imbalance between serotonin and other neurotransmitter levels may be involved in the pathogenesis of human cataplexy.

Ionization Suppression and Recovery in Direct Biofluid Analysis Using Paper Spray Mass Spectrometry

NASA Astrophysics Data System (ADS)

Vega, Carolina; Spence, Corina; Zhang, Chengsen; Bills, Brandon J.; Manicke, Nicholas E.

2016-04-01

Paper spray mass spectrometry is a method for the direct analysis of biofluid samples in which extraction of analytes from dried biofluid spots and electrospray ionization occur from the paper on which the dried sample is stored. We examined matrix effects in the analysis of small molecule drugs from urine, plasma, and whole blood. The general method was to spike stable isotope labeled analogs of each analyte into the spray solvent, while the analyte itself was in the dried biofluid. Intensity of the labeled analog is proportional to ionization efficiency, whereas the ratio of the analyte intensity to the labeled analog in the spray solvent is proportional to recovery. Ion suppression and recovery were found to be compound- and matrix-dependent. Highest levels of ion suppression were obtained for poor ionizers (e.g., analytes lacking basic aliphatic amine groups) in urine and approached -90%. Ion suppression was much lower or even absent for good ionizers (analytes with aliphatic amines) in dried blood spots. Recovery was generally highest in urine and lowest in blood. We also examined the effect of two experimental parameters on ion suppression and recovery: the spray solvent and the sample position (how far away from the paper tip the dried sample was spotted). Finally, the change in ion suppression and analyte elution as a function of time was examined by carrying out a paper spray analysis of dried plasma spots for 5 min by continually replenishing the spray solvent.

Acrolein generation stimulates hypercontraction in isolated human blood vessels

PubMed Central

Conklin, D.J.; Bhatnagar, A.; Cowley, H.R.; Johnson, G.H.; Wiechmann, R.J.; Sayre, L.M.; Trent, M.B.; Boor, P.J.

2012-01-01

Increased risk of vasospasm, a spontaneous hyperconstriction, is associated with atherosclerosis, cigarette smoking, and hypertension—all conditions involving oxidative stress, lipid peroxidation, and inflammation. To test the role of the lipid peroxidation- and inflammation-derived aldehyde, acrolein, in human vasospasm, we developed an ex vivo model using human coronary artery bypass graft (CABG) blood vessels and a demonstrated acrolein precursor, allylamine. Allylamine induces hypercontraction in isolated rat coronary artery in a semicarbazide-sensitive amine oxidase activity (SSAO) dependent manner. Isolated human CABG blood vessels (internal mammary artery, radial artery, saphenous vein) were used to determine: (1) vessel responses and sensitivity to acrolein, allylamine, and H2O2 exposure (1 μM–1 mM), (2) SSAO dependence of allylamine-induced effects using SSAO inhibitors (semicarbazide, 1 mM; MDL 72274-E, active isomer; MDL 72274-Z, inactive isomer; 100 μM), (3) the vasoactive effects of two other SSAO amine substrates, benzylamine and methylamine, and (4) the contribution of extracellular Ca2+ to hypercontraction. Acrolein or allylamine but not H2O2, benzylamine, or methylamine stimulated spontaneous and pharmacologically intractable hypercontraction in CABG blood vessels that was similar to clinical vasospasm. Allylamine-induced hypercontraction and blood vessel SSAO activity were abolished by pretreatment with semicarbazide or MDL 72274-E but not by MDL 72274-Z. Allylamine-induced hypercontraction also was significantly attenuated in Ca2+-free buffer. In isolated aorta of spontaneously hypertensive rat, allylamine-induced an SSAO-dependent contraction and enhanced norepinephrine sensitivity but not in Sprague–Dawley rat aorta. We conclude that acrolein generation in the blood vessel wall increases human susceptibility to vasospasm, an event that is enhanced in hypertension. PMID:17095030

Update on the pharmacology of selective inhibitors of MAO-A and MAO-B: focus on modulation of CNS monoamine neurotransmitter release.

PubMed

Finberg, John P M

2014-08-01

Inhibitors of monoamine oxidase (MAO) were initially used in medicine following the discovery of their antidepressant action. Subsequently their ability to potentiate the effects of an indirectly-acting sympathomimetic amine such as tyramine was discovered, leading to their limitation in clinical use, except for cases of treatment-resistant depression. More recently, the understanding that: a) potentiation of indirectly-acting sympathomimetic amines is caused by inhibitors of MAO-A but not by inhibitors of MAO-B, and b) that reversible inhibitors of MAO-A cause minimal tyramine potentiation, has led to their re-introduction to clinical use for treatment of depression (reversible MAO-A inhibitors and new dose form MAO-B inhibitor) and treatment of Parkinson's disease (MAO-B inhibitors). The profound neuroprotective properties of propargyl-based inhibitors of MAO-B in preclinical experiments have drawn attention to the possibility of employing these drugs for their neuroprotective effect in neurodegenerative diseases, and have raised the question of the involvement of the MAO-mediated reaction as a source of reactive free radicals. Despite the long-standing history of MAO inhibitors in medicine, the way in which they affect neuronal release of monoamine neurotransmitters is still poorly understood. In recent years, the detailed chemical structure of MAO-B and MAO-A has become available, providing new possibilities for synthesis of mechanism-based inhibitors. This review describes the latest advances in understanding the way in which MAO inhibitors affect the release of the monoamine neurotransmitters dopamine, noradrenaline and serotonin (5-HT) in the CNS, with an accent on the importance of these effects for the clinical actions of the drugs. Copyright © 2014 Elsevier Inc. All rights reserved.

Synthesis, biological assessment and molecular modeling of new multipotent MAO and cholinesterase inhibitors as potential drugs for the treatment of Alzheimer's disease.

PubMed

Samadi, Abdelouahid; Chioua, Mourad; Bolea, Irene; de Los Ríos, Cristóbal; Iriepa, Isabel; Moraleda, Ignacio; Bastida, Agatha; Esteban, Gerard; Unzeta, Mercedes; Gálvez, Enrique; Marco-Contelles, José

2011-09-01

The synthesis, biological evaluation and molecular modeling of new multipotent inhibitors of type I and type II, able to simultaneously inhibit monoamine oxidases (MAO) as well as acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE), is described. Compounds of type I were prepared by sequential reaction of 2,6-dichloro-4-phenylpyridine-3,5-dicarbonitrile (14) [or 2,6-dichloropyridine-3,5-dicarbonitrile (15)] with prop-2-yn-1-amine (or N-methylprop-2-yn-1-amine) and 2-(1-benzyl-piperidin-4-yl)alkylamines 22-25. Compounds of type II were prepared by Friedländer type reaction of 6-amino-5-formyl-2-(methyl(prop-2-yn-1-yl)amino)nicotinonitriles 32 and 33 with 4-(1-benzylpiperidin-4-yl)butan-2-one (31). The biological evaluation of molecules 1-11 showed that most of these compounds are potent, in the nanomolar range, and selective AChEI, with moderate and equipotent selectivity for MAO-A and MAO-B inhibition. Kinetic studies of compound 8 proved that this is a EeAChE mixed type inhibitor (IC(50) = 16 ± 2; Ki = 12 ± 3 nM). Molecular modeling investigation on compound 8 confirmed its dual AChE inhibitory profile, binding simultaneously at the catalytic active site (CAS) and at the peripheric anionic site (PAS). In overall, compound 11, as a potent and selective dual AChEI, showing a moderate and selective MAO-A inhibitory profile, can be considered as an attractive multipotent drug for further development on two key pharmacological targets playing key roles in the therapy of Alzheimer's disease. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

Metabolism of β-valine via a CoA-dependent ammonia lyase pathway.

PubMed

Otzen, Marleen; Crismaru, Ciprian G; Postema, Christiaan P; Wijma, Hein J; Heberling, Matthew M; Szymanski, Wiktor; de Wildeman, Stefaan; Janssen, Dick B

2015-11-01

Pseudomonas species strain SBV1 can rapidly grow on medium containing β-valine as a sole nitrogen source. The tertiary amine feature of β-valine prevents direct deamination reactions catalyzed by aminotransferases, amino acid dehydrogenases, and amino acid oxidases. However, lyase- or aminomutase-mediated conversions would be possible. To identify enzymes involved in the degradation of β-valine, a PsSBV1 gene library was prepared and used to complement the β-valine growth deficiency of a closely related Pseudomonas strain. This resulted in the identification of a gene encoding β-valinyl-coenzyme A ligase (BvaA) and two genes encoding β-valinyl-CoA ammonia lyases (BvaB1 and BvaB2). The BvaA protein demonstrated high sequence identity to several known phenylacetate CoA ligases. Purified BvaA enzyme did not convert phenyl acetic acid but was able to activate β-valine in an adenosine triphosphate (ATP)- and CoA-dependent manner. The substrate range of the enzyme appears to be narrow, converting only β-valine and to a lesser extent, 3-aminobutyrate and β-alanine. Characterization of BvaB1 and BvaB2 revealed that both enzymes were able to deaminate β-valinyl-CoA to produce 3-methylcrotonyl-CoA, a common intermediate in the leucine degradation pathway. Interestingly, BvaB1 and BvaB2 demonstrated no significant sequence identity to known CoA-dependent ammonia lyases, suggesting they belong to a new family of enzymes. BLAST searches revealed that BvaB1 and BvaB2 show high sequence identity to each other and to several enoyl-CoA hydratases, a class of enzymes that catalyze a similar reaction with water instead of amine as the leaving group.

Evaluation Physical Characteristics and Comparison Antimicrobial and Anti-Inflammation Potentials of Dental Root Canal Sealers Containing Hinokitiol In Vitro

PubMed Central

Shih, Yin-Hua; Lin, Dan-Jae; Chang, Kuo-Wei; Hsia, Shih-Min; Ko, Shun-Yao; Lee, Shyh-Yuan; Hsue, Shui-Sang; Wang, Tong-Hong; Chen, Yi-Ling; Shieh, Tzong-Ming

2014-01-01

Hinokitiol displays potent antimicrobial activity. It has been used in toothpaste and oral-care gel to improve the oral lichen planus and reduce halitosis. The aim of this study was to evaluate the antimicrobial activity of 3 different dental root canal sealers with hinokitiol (sealers+H) and their physical and biological effects. AH Plus (epoxy amine resin-based, AH), Apexit Plus (calcium-hydroxide-based, AP), and Canals (zinc-oxide-eugenol-based, CA), were used in this study. The original AH and CA exhibited strong anti-methicillin-resistant Staphylococcus aureus (anti-MRSA) activity, but AP did not. The setting time, working time, flowability, film thickness, and solubility of each sealer+0.2%H complied with ISO 6876:2001. CA+0.2%H exhibited high cytotoxicity, but the others sealers+0.2%H did not. Because hinokitiol combined with Zn2+ in CA creates a synergistic effect. The physical tests of AP+0.5%–1%H complied with ISO 6876:2001, improved antimicrobial activity, inhibited inflammation genes cyclooxygenase-2 (COX-2) and hypoxia-inducible factor-1α (HIF-1α) mRNA in MG-63 cells and human gingival fibroblasts (HGF), and down-regulated lysyl oxidase (LOX) mRNA of HGF. In summary, AH and CA demonstrated strong antimicrobial activity, but AP did not. Application of hinokitiol increases AH anti-MRSA activity should less than 0.2% for keep well flowability. AP+0.5%–1% hinokitiol exhibited strong physical, antibacterial, and anti-inflammation potentials, and inhibited S. aureus abscess formation. Applying an appreciable proportion of hinokitiol to epoxy-amine-resin-based and calcium-hydroxide-based root canal sealers is warranted, but the enhanced cytotoxicity and synergistic effect must be considered. PMID:24915566

Oxygen activation at the plasma membrane: relation between superoxide and hydroxyl radical production by isolated membranes.

PubMed

Heyno, Eiri; Mary, Véronique; Schopfer, Peter; Krieger-Liszkay, Anja

2011-07-01

Production of reactive oxygen species (hydroxyl radicals, superoxide radicals and hydrogen peroxide) was studied using EPR spin-trapping techniques and specific dyes in isolated plasma membranes from the growing and the non-growing zones of hypocotyls and roots of etiolated soybean seedlings as well as coleoptiles and roots of etiolated maize seedlings. NAD(P)H mediated the production of superoxide in all plasma membrane samples. Hydroxyl radicals were only produced by the membranes of the hypocotyl growing zone when a Fenton catalyst (FeEDTA) was present. By contrast, in membranes from other parts of the seedlings a low rate of spontaneous hydroxyl radical formation was observed due to the presence of small amounts of tightly bound peroxidase. It is concluded that apoplastic hydroxyl radical generation depends fully, or for the most part, on peroxidase localized in the cell wall. In soybean plasma membranes from the growing zone of the hypocotyl pharmacological tests showed that the superoxide production could potentially be attributed to the action of at least two enzymes, an NADPH oxidase and, in the presence of menadione, a quinone reductase.

Advanced oxidation protein products induce chondrocyte apoptosis via receptor for advanced glycation end products-mediated, redox-dependent intrinsic apoptosis pathway.

PubMed

Wu, Qian; Zhong, Zhao-Ming; Zhu, Si-Yuan; Liao, Cong-Rui; Pan, Ying; Zeng, Ji-Huan; Zheng, Shuai; Ding, Ruo-Ting; Lin, Qing-Song; Ye, Qing; Ye, Wen-Bin; Li, Wei; Chen, Jian-Ting

2016-01-01

Pro-inflammatory cytokine-induced chondrocyte apoptosis is a primary cause of cartilage destruction in the progression of rheumatoid arthritis (RA). Advanced oxidation protein products (AOPPs), a novel pro-inflammatory mediator, have been confirmed to accumulate in patients with RA. However, the effect of AOPPs accumulation on chondrocyte apoptosis and the associated cellular mechanisms remains unclear. The present study demonstrated that the plasma formation of AOPPs was enhanced in RA rats compared with normal. Then, chondrocyte were treated with AOPPs-modified rat serum albumin (AOPPs-RSA) in vitro. Exposure of chondrocyte to AOPPs activated nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and increased expression of NADPH oxidase subunits, which was mediated by receptor for advanced glycation end products (RAGE), but not scavenger receptor CD36. Moreover, AOPPs challenge triggered NADPH oxidase-dependent ROS generation which induced mitochondrial dysfunction and endoplasmic reticulum stress resulted in activation of caspase family that eventually lead to apoptosis. Lastly, blockade of RAGE, instead of CD36, largely attenuated these signals. Our study demonstrated first time that AOPPs induce chondrocyte apoptosis via RAGE-mediated and redox-dependent intrinsic apoptosis pathway in vitro. These data implicates that AOPPs may represent a novel pathogenic factor that contributes to RA progression. Targeting AOPPs-triggered cellular mechanisms might emerge as a promising therapeutic option for patients with RA.

Aqueous two-phase partition applied to the isolation of plasma membranes and Golgi apparatus from cultured mammalian cells.

PubMed

Morré, D M; Morre, D J

2000-06-23

Partitioning in dextran-poly(ethylene)glycol (PEG) aqueous-aqueous phase systems represents a mature technology with many applications to separations of cells and to the preparation of membranes from mammalian cells. Most applications to membrane isolation and purification have focused on plasma membranes, plasma membrane domains and separation of right side-out and inside-out plasma membrane vesicles. The method exploits a combination of membrane properties, including charge and hydrophobicity. Purification is based upon differential distributions of the constituents in a sample between the two principal compartments of the two phases (upper and lower) and at the interface. The order of affinity of animal cell membranes for the upper phase is: endoplasmic reticulum

Aqueous two-phase partition applied to the isolation of plasma membranes and Golgi apparatus from cultured mammalian cells

NASA Technical Reports Server (NTRS)

Morre, D. M.; Morre, D. J.

2000-01-01

Partitioning in dextran-poly(ethylene)glycol (PEG) aqueous-aqueous phase systems represents a mature technology with many applications to separations of cells and to the preparation of membranes from mammalian cells. Most applications to membrane isolation and purification have focused on plasma membranes, plasma membrane domains and separation of right side-out and inside-out plasma membrane vesicles. The method exploits a combination of membrane properties, including charge and hydrophobicity. Purification is based upon differential distributions of the constituents in a sample between the two principal compartments of the two phases (upper and lower) and at the interface. The order of affinity of animal cell membranes for the upper phase is: endoplasmic reticulum

Chemoselective organocatalytic aerobic oxidation of primary amines to secondary imines.

PubMed

Wendlandt, Alison E; Stahl, Shannon S

2012-06-01

Biomimetic aerobic oxidation of primary benzylic amines has been achieved by using a quinone catalyst. Excellent selectivity is observed for primary, unbranched benzylic amines relative to secondary/tertiary amines, branched benzylic amines, and aliphatic amines. The exquisite selectivity for benzylic amines enables oxidative self-sorting within dynamic mixtures of amines and imines to afford high yields of cross-coupled imine products.

A molecular key for building hyphae aggregates: the role of the newly identified Streptomyces protein HyaS.

PubMed

Koebsch, Ilona; Overbeck, Jens; Piepmeyer, Sophie; Meschke, Holger; Schrempf, Hildgund

2009-05-01

Streptomycetes produce many metabolites with medical and biotechnological applications. During fermentations, their hyphae build aggregates, a process in which the newly identified protein HyaS plays an important role. The corresponding hyaS gene is present within all investigated Streptomyces species. Reporter fusions indicate that transcription of hyaS occurs within substrate hyphae of the Streptomyces lividans wild type (WT). The HyaS protein is dominantly associated with the substrate hyphae. The WT strain forms cylindrically shaped clumps of densely packed substrate hyphae, often fusing to higher aggregates (pellets), which remain stably associated during shaking. Investigations by electron microscopy suggest that HyaS induces tight fusion-like contacts among substrate hyphae. In contrast, the pellets of the designed hyaS disruption mutant ΔH are irregular in shape, contain frequently outgrowing bunches of hyphae, and fuse less frequently. ΔH complemented with a plasmid carrying hyaS resembles the WT phenotype. Biochemical studies indicate that the C-terminal region of HyaS has amine oxidase activity. Investigations of ΔH transformants, each carrying a specifically mutated gene, lead to the conclusion that the in situ oxidase activity correlates with the pellet-inducing role of HyaS, and depends on the presence of certain histidine residues. Furthermore, the level of undecylprodigiosin, a red pigment with antibiotic activity, is influenced by the engineered hyaS subtype within a strain. These data present the first molecular basis for future manipulation of pellets, and concomitant production of secondary metabolites during biotechnological processes. © 2009 The Authors. Journal compilation © 2009 Society for Applied Microbiology and Blackwell Publishing Ltd.

Linking Inflammation and Parkinson Disease: Hypochlorous Acid Generates Parkinsonian Poisons.

PubMed

Jeitner, Thomas M; Kalogiannis, Mike; Krasnikov, Boris F; Gomolin, Irving; Peltier, Morgan R; Moran, Graham R

2016-06-01

Inflammation is a common feature of Parkinson Disease and other neurodegenerative disorders. Hypochlorous acid (HOCl) is a reactive oxygen species formed by neutrophils and other myeloperoxidase-containing cells during inflammation. HOCl chlorinates the amine and catechol moieties of dopamine to produce chlorinated derivatives collectively termed chlorodopamine. Here, we report that chlorodopamine is toxic to dopaminergic neurons both in vivo and in vitro Intrastriatal administration of 90 nmol chlorodopamine to mice resulted in loss of dopaminergic neurons from the substantia nigra and decreased ambulation-results that were comparable to those produced by the same dose of the parkinsonian poison, 1-methyl-4-phenylpyridinium (MPP+). Chlorodopamine was also more toxic to differentiated SH SY5Y cells than HOCl. The basis of this selective toxicity is likely mediated by chlorodopamine uptake through the dopamine transporter, as expression of this transporter in COS-7 cells conferred sensitivity to chlorodopamine toxicity. Pharmacological blockade of the dopamine transporter also mitigated the deleterious effects of chlorodopamine in vivo The cellular actions of chlorodopamine included inactivation of the α-ketoglutarate dehydrogenase complex, as well as inhibition of mitochondrial respiration. The latter effect is consistent with inhibition of cytochrome c oxidase. Illumination at 670 nm, which stimulates cytochrome c oxidase, reversed the effects of chlorodopamine. The observed changes in mitochondrial biochemistry were also accompanied by the swelling of these organelles. Overall, our findings suggest that chlorination of dopamine by HOCl generates toxins that selectively kill dopaminergic neurons in the substantia nigra in a manner comparable to MPP+. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Platelet monoamine oxidase type B, MAOB intron 13 and MAOA-uVNTR polymorphism and symptoms of post-traumatic stress disorder.

PubMed

Svob Strac, Dubravka; Kovacic Petrovic, Zrnka; Nikolac Perkovic, Matea; Umolac, Danica; Nedic Erjavec, Gordana; Pivac, Nela

2016-07-01

Post-traumatic stress disorder (PTSD), a disorder that develops following exposure to traumatic experience(s), is frequently associated with agitation, aggressive behavior and psychotic symptoms. Monoamine oxidase (MAO) degrades different biogenic amines and regulates mood, emotions and behavior, and has a role in the pathophysiology of various neuropsychiatric disorders. The aim of the study was to investigate the association between different symptoms occurring in PTSD [PTSD symptom severity assessed by the Clinician Administered PTSD Scale (CAPS), agitation and selected psychotic symptoms assessed by the Positive and Negative Syndrome Scale (PANSS)] and platelet MAO-B activity and/or genetic variants of MAOB rs1799836 and MAOA-uVNTR polymorphisms in 249 Croatian male veterans with PTSD. Our study revealed slightly higher platelet MAO-B activity in veterans with PTSD with more severe PTSD symptoms and in veterans with agitation, and significantly higher platelet MAO-B activity in veterans with more pronounced psychotic symptoms compared to veterans with less pronounced psychotic symptoms. Platelet MAO-B activity was associated with smoking but not with age. Genetic variants of MAOB rs1799836 and MAOA-uVNTR were not associated with agitation and selected psychotic symptoms in veterans with PTSD. A marginally significant association was found between MAOB rs1799836 polymorphism and severity of PTSD symptoms, but it was not confirmed since carriers of G or A allele of MAOB rs1799836 did not differ in their total CAPS scores. These findings suggest an association of platelet MAO-B activity, but a lack of association of MAOB rs1799836 and MAOA-uVNTR, with selected psychotic symptoms in ethnically homogenous veterans with PTSD.

Model studies of the Cu(B) site of cytochrome c oxidase utilizing a Zn(II) complex containing an imidazole-phenol cross-linked ligand.

PubMed

Pesavento, Russell P; Pratt, Derek A; Jeffers, Jerry; van der Donk, Wilfred A

2006-07-21

Cytochrome c oxidase, the enzyme complex responsible for the four-electron reduction of O2 to H2O, contains an unusual histidine-tyrosine cross-link in its bimetallic heme a3-CuB active site. We have synthesised an unhindered, tripodal chelating ligand, BPAIP, containing the unusual ortho-imidazole-phenol linkage, which mimics the coordination environment of the CuB center. The ligand was used to investigate the physicochemical (pKa, oxidation potential) and coordination properties of the imidazole-phenol linkage when bound to a dication. Zn(II) coordination lowers the pKa of the phenol by 0.6 log units, and increases the potential of the phenolate/phenoxyl radical couple by approximately 50 mV. These results are consistent with inductive withdrawal of electron density from the phenolic ring. Spectroscopic data and theoretical calculations (DFT) were used to establish that the cationic complex [Zn(BPAIP)Br]+ has an axially distorted trigonal bipyramidal structure, with three coordinating nitrogen ligands (two pyridine and one imidazole) occupying the equatorial plane and the bromide and the tertiary amine nitrogen of the tripod in the axial positions. Interestingly, the Zn-Namine bonding interaction is weak or absent in [Zn(BPAIP)Br]+ and the complex gains stability in basic solutions, as indicated by 1H NMR spectroscopy. These observations are supported by theoretical calculations (DFT), which suggest that the electron-donating capacity of the equatorial imidazole ligand can be varied by modulation of the protonation and/or redox state of the cross-linked phenol. Deprotonation of the phenol makes the equatorial imidazole a stronger sigma-donor, resulting in an increased Zn-Nimd interaction and thereby leading to distortion of the axial ligand axis toward a more tetrahedral geometry.

Potent selective monoamine oxidase B inhibition by maackiain, a pterocarpan from the roots of Sophora flavescens.

PubMed

Lee, Hyun Woo; Ryu, Hyung Won; Kang, Myung-Gyun; Park, Daeui; Oh, Sei-Ryang; Kim, Hoon

2016-10-01

Monoamine oxidase (MAO) catalyzes the oxidation of monoamines and its two isoforms, MAO-A and MAO-B, break down neurotransmitter amines. Of the compounds isolated from the roots of Sophora flavescens, (-)-maackiain (4), a pterocarpan, was found to potently and selectively inhibit human MAO-B, with an IC50 of 0.68μM, and to have a selectivity index of 126.2 for MAO-B. As compared with other herbal natural products, the IC50 value of 4 for MAO-B is one of the lowest reported to date. Genistein (1) highly, effectively and non-selectively inhibited MAO-A and MAO-B with IC50 values of 3.9μM and 4.1μM, respectively. (-)-4-Hydroxy-3-methoxy-8,9-methylenedioxypterocarpan (2) effectively and non-selectively inhibited MAO-A and MAO-B with IC50 values of 20.3μM and 10.3μM, respectively. In addition, compound 4 reversibly and competitively inhibited MAO-B with a Ki value of 0.054μM. Molecular docking simulation revealed that the binding affinity of 4 for MAO-B (-26.6kcal/mol) was greater than its affinity for MAO-A (-8.3kcal/mol), which was in-line with our inhibitory activity findings. Furthermore, Cys172 of MAO-B was found to be a key residue for hydrogen bonding with compound 4. The findings of this study suggest compound 4 be viewed as a new potent, selective, and reversible MAO-B inhibitor, and that compounds 1 and 2 be considered useful lead compounds for the developments of nonselective and reversible MAO inhibitors for the treatment of disorders like Parkinson's disease, Alzheimer disease, and depression. Copyright © 2016 Elsevier Ltd. All rights reserved.

Monoamine Oxidase B Prompts Mitochondrial and Cardiac Dysfunction in Pressure Overloaded Hearts

PubMed Central

Kaludercic, Nina; Carpi, Andrea; Nagayama, Takahiro; Sivakumaran, Vidhya; Zhu, Guangshuo; Lai, Edwin W.; Bedja, Djahida; De Mario, Agnese; Chen, Kevin; Gabrielson, Kathleen L.; Lindsey, Merry L.; Pacak, Karel; Takimoto, Eiki; Shih, Jean C.; Kass, David A.; Di Lisa, Fabio

2014-01-01

Abstract Aims: Monoamine oxidases (MAOs) are mitochondrial flavoenzymes responsible for neurotransmitter and biogenic amines catabolism. MAO-A contributes to heart failure progression via enhanced norepinephrine catabolism and oxidative stress. The potential pathogenetic role of the isoenzyme MAO-B in cardiac diseases is currently unknown. Moreover, it is has not been determined yet whether MAO activation can directly affect mitochondrial function. Results: In wild type mice, pressure overload induced by transverse aortic constriction (TAC) resulted in enhanced dopamine catabolism, left ventricular (LV) remodeling, and dysfunction. Conversely, mice lacking MAO-B (MAO-B−/−) subjected to TAC maintained concentric hypertrophy accompanied by extracellular signal regulated kinase (ERK)1/2 activation, and preserved LV function, both at early (3 weeks) and late stages (9 weeks). Enhanced MAO activation triggered oxidative stress, and dropped mitochondrial membrane potential in the presence of ATP synthase inhibitor oligomycin both in neonatal and adult cardiomyocytes. The MAO-B inhibitor pargyline completely offset this change, suggesting that MAO activation induces a latent mitochondrial dysfunction, causing these organelles to hydrolyze ATP. Moreover, MAO-dependent aldehyde formation due to inhibition of aldehyde dehydrogenase 2 activity also contributed to alter mitochondrial bioenergetics. Innovation: Our study unravels a novel role for MAO-B in the pathogenesis of heart failure, showing that both MAO-driven reactive oxygen species production and impaired aldehyde metabolism affect mitochondrial function. Conclusion: Under conditions of chronic hemodynamic stress, enhanced MAO-B activity is a major determinant of cardiac structural and functional disarrangement. Both increased oxidative stress and the accumulation of aldehyde intermediates are likely liable for these adverse morphological and mechanical changes by directly targeting mitochondria. Antioxid. Redox Signal. 20, 267–280. PMID:23581564

Serotonin Receptor 6 Mediates Defective Brain Development in Monoamine Oxidase A-deficient Mouse Embryos

PubMed Central

Wang, Chi Chiu; Man, Gene Chi Wai; Chu, Ching Yan; Borchert, Astrid; Ugun-Klusek, Aslihan; Billett, E. Ellen; Kühn, Hartmut; Ufer, Christoph

2014-01-01

Monoamine oxidases A and B (MAO-A and MAO-B) are enzymes of the outer mitochondrial membrane that metabolize biogenic amines. In the adult central nervous system, MAOs have important functions for neurotransmitter homeostasis. Expression of MAO isoforms has been detected in the developing embryo. However, suppression of MAO-B does not induce developmental alterations. In contrast, targeted inhibition and knockdown of MAO-A expression (E7.5–E10.5) caused structural abnormalities in the brain. Here we explored the molecular mechanisms underlying defective brain development induced by MAO-A knockdown during in vitro embryogenesis. The developmental alterations were paralleled by diminished apoptotic activity in the affected neuronal structures. Moreover, dysfunctional MAO-A expression led to elevated levels of embryonic serotonin (5-hydroxytryptamine (5-HT)), and we found that knockdown of serotonin receptor-6 (5-Htr6) expression or pharmacologic inhibition of 5-Htr6 activity rescued the MAO-A knockdown phenotype and restored apoptotic activity in the developing brain. Our data suggest that excessive 5-Htr6 activation reduces activation of caspase-3 and -9 of the intrinsic apoptotic pathway and enhances expression of antiapoptotic proteins Bcl-2 and Bcl-XL. Moreover, we found that elevated 5-HT levels in MAO-A knockdown embryos coincided with an enhanced activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and a reduction of proliferating cell numbers. In summary, our findings suggest that excessive 5-HT in MAO-A-deficient mouse embryos triggers cellular signaling cascades via 5-Htr6, which suppresses developmental apoptosis in the brain and thus induces developmental retardations. PMID:24497636

Functional purification of the monocarboxylate transporter of the yeast Candida utilis.

PubMed

Baltazar, Fátima; Cássio, Fernanda; Leão, Cecília

2006-08-01

Plasma membranes of the yeast, Candida utilis, were solubilized with octyl-beta-D-glucopyranoside and a fraction enriched in the lactate carrier was obtained with DEAE-Sepharose anion-exchange chromatography, after elution with 0.4 M NaCl. The uptake of lactic acid into proteoliposomes, containing the purified protein fraction and cytochrome c oxidase, was dependent on a proton-motive force and the transport specificity was consistent with the one of C. utilis intact cells. Overall, we have obtained a plasma membrane fraction enriched in the lactate carrier of C. utilis in which the transport properties were preserved. Given the similarities between the lactate transport of C. utilis and the one of mammalian cells, this purified system could be further explored to screen for specific lactate inhibitors, with potential therapeutic applications.

Mechanisms for cellular NO oxidation and nitrite formation in lung epithelial cells.

PubMed

Zhao, Xue-Jun; Wang, Ling; Shiva, Sruti; Tejero, Jesus; Myerburg, Mike M; Wang, Jun; Frizzell, Sam; Gladwin, Mark T

2013-08-01

Airway lining fluid contains relatively high concentrations of nitrite, and arterial blood levels of nitrite are higher than venous levels, suggesting the lung epithelium may represent an important source of nitrite in vivo. To investigate whether lung epithelial cells possess the ability to convert NO to nitrite by oxidation, and the effect of oxygen reactions on nitrite formation, the NO donor DETA NONOate was incubated with or without A549 cells or primary human bronchial epithelial (HBE) cells for 24 h under normoxic (21% O2) and hypoxic (1% O2) conditions. Nitrite production was significantly increased under all conditions in the presence of A549 or HBE cells, suggesting that both A549 and HBE cells have the capacity to oxidize NO to nitrite even under low-oxygen conditions. The addition of oxyhemoglobin to the A549 cell medium decreased the production of nitrite, consistent with NO scavenging limiting nitrite formation. Heat-denatured A549 cells produced much lower nitrite and nitrate, suggesting an enzymatic activity is required. This NO oxidation activity was highest in membrane-bound proteins with molecular size <100kDa. In addition, 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one and cyanide inhibited formation of nitrite in A549 cells. It has been shown that ceruloplasmin (Cp) possesses an NO oxidase and nitrite synthase activity in plasma based on NO oxidation to nitrosonium cation. We observed that Cp is expressed intracellularly in lung epithelial A549 cells and secreted into the medium under basal conditions and during cytokine stimulation. However, an analysis of Cp expression level and activity measured via p-phenylenediamine oxidase activity assay revealed very low activity compared with plasma, suggesting that there is insufficient Cp to contribute to detectable NO oxidation to nitrite in A549 cells. Additionally, Cp levels were knocked down using siRNA by more than 75% in A549 cells, with no significant change in either nitrite or cellular S-nitrosothiol formation compared to scrambled siRNA control under basal conditions or cytokine stimulation. These data suggest that lung epithelial cells possess NO oxidase activity, which is enhanced in cell-membrane-associated proteins and not regulated by intracellular or secreted Cp, indicating that alternative NO oxidases determine hypoxic and normoxic nitrite formation from NO in human lung epithelial cells. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Analysis of 4-methylthioamphetamine in clinical specimens.

PubMed

Elliott, S P

2001-07-01

There has been much publicity, particularly in Europe, regarding a new phenylethylamine-based compound called 4-methylthioamphetamine (4-MTA), also known as para-methylthioamphetamine (p-MTA), MTA or 'Flatliner'. Chemically, 4-MTA is an amphetamine derivative and is a potent, non-neurotoxic serotonin-releasing agent and reversible inhibitor of rat monoamine oxidase-A. Its effects, therefore, appear different from those of amphetamine. Analysis of various plasma and urine specimens in three clinical cases implicating 'Ecstasy' ingestion revealed the presence of 4-MTA. Presumed metabolites were also detected, with one compound identified as being 4-MTA sulphoxide. The concentrations of 4-MTA measured in the plasma ranged from 0.131 mg/L to 0.760 mg/L. In one patient the 4-MTA concentration was determined in a series of plasma samples and this allowed a presumptive half-life of approximately 7 h to be estimated. This paper describes the first reported data regarding possible pharmacokinetics of 4-MTA in humans and presents the first reported non-fatal instances of 4-MTA intoxication in the UK.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Peden, D.B.; Swiersz, M.; Ohkubo, K.

Uric acid, an important scavenger of ozone, has been identified as the major low molecular weight antioxidant in baseline and cholinergically induced nasal secretions. The purpose of this study was to determine the specific tissue source of uric acid in airway secretions. The secretion of uric acid is increased by cholinergic stimulation and correlates closely with the secretion of lactoferrin (a nasal glandular protein), suggesting that submucosal glands are involved. Indeed, nasal turbinate tissue was found to contain uric acid. However, careful analysis of nasal turbinate tissue failed to reveal the presence of xanthine oxidase, the enzyme responsible for uricmore » acid synthesis. These data suggest that uric acid might be taken up secondarily by glands from plasma. This possibility was strengthened by the observation that lowering the plasma urate level with probenecid concomitantly lowered urate secretion. These findings are consistent with the hypotheses that the principal source of uric acid in nasal secretions is plasma and that uric acid is taken up, concentrated, and secreted by nasal glands.« less

Protecting-group-free synthesis of amines: synthesis of primary amines from aldehydes via reductive amination.

PubMed

Dangerfield, Emma M; Plunkett, Catherine H; Win-Mason, Anna L; Stocker, Bridget L; Timmer, Mattie S M

2010-08-20

New methodology for the protecting-group-free synthesis of primary amines is presented. By optimizing the metal hydride/ammonia mediated reductive amination of aldehydes and hemiacetals, primary amines were selectively prepared with no or minimal formation of the usual secondary and tertiary amine byproduct. The methodology was performed on a range of functionalized aldehyde substrates, including in situ formed aldehydes from a Vasella reaction. These reductive amination conditions provide a valuable synthetic tool for the selective production of primary amines in fewer steps, in good yields, and without the use of protecting groups.

Polyketone polymer: a new support for direct enzyme immobilization.

PubMed

Agostinelli, E; Belli, F; Tempera, G; Mura, A; Floris, G; Toniolo, L; Vavasori, A; Fabris, S; Momo, F; Stevanato, R

2007-01-20

Polyketone polymer -[-CO-CH(2)-CH(2)-](n)-, obtained by copolymerization of ethene and carbon monoxide, is utilized for immobilization of three different enzymes, one peroxidase from horseradish (HRP) and two amine oxidases, from bovine serum (BSAO) and lentil seedlings (LSAO). The easy immobilization procedure is carried out in diluted buffer, at pH 7.0 and 3 degrees C, gently mixing the proteins with the polymer. No bifunctional reagents and spacer arms are required for the immobilization, which occurs exclusively via a large number of hydrogen bonds between the carbonyl groups of the polymer and the -NH groups of the polypeptidic chain. Experiments demonstrate a high linking capacity of polymer for BSAO and an extraordinary strong linkage for LSAO. Moreover, activity measurements demonstrate that immobilized LSAO totally retains the catalytic characteristics of the free enzyme, where only a limited increase of K(M) value is observed. Finally, the HRP-activated polymer is successfully used as active packed bed of an enzymatic reactor for continuous flow conversion and flow injection analysis of hydrogen peroxide containing solutions.

N-(/sup 11/C)-methyl-p-substituted phentermine analogs as potential brain blood flow agents for positron tomography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kizuka, H.; Elmaleh, D.R.; Boudreaux, G.J.

The addition of a methyl group to the ..cap alpha..-position of amphetamine increases both the lipophilicity of the agent and its resistance to metabolism by monoamine oxidase. In addition, since tritium substituted phenteramine analog studies suggested that the p-halo phentermines had a greater concentration in the brain and prolonged retention time, the authors evaluated the biological behavior of positron labeled ..cap alpha..-methylamphetamine (phenteramine) in rats, dogs and monkeys. The N-(/sup 11/C) methyl analogs of p-chloro (I) and p-fluoro (II) phentermines were prepared by methylation of their primary amines using /sup 11/Ch/sub 3/I. Biodistribution studies in rats shows brain uptake ismore » in the range of 1% dose/gr at 5 and 15 min for both agents. The activity in blood and eyes is low. Sequential images of the dogs' brain over 1 hour revealed a clearance of <15%. Images of the monkey brain were also obtained using a MGH positron camera PCR-I.« less

Microchip-Based Organophosphorus Detection Using Bienzyme Bioelectrocatalysis

NASA Astrophysics Data System (ADS)

Han, Yong Duk; Jeong, Chi Yong; Lee, Jun Hee; Lee, Dae-Sik; Yoon, Hyun C.

2012-06-01

We have developed a microsystem for the detection of organophosphorus (OP) compounds using acetylcholine esterase (AchE) and choline oxidase (ChOx) bienzyme bioelectrocatalysis. Because AchE is irreversibly inhibited by OP pesticides, the change in AchE activity with OP treatment can be traced to determine OP concentration. Polymer-associated ChOx immobilization on the working electrode surface and magnetic microparticle (MP)-assisted AchE deposition methods were employed to create an AchE-ChOx bienzyme-modified biosensing system. ChOx was immobilized on the micropatterned electrodes using poly(L-lysine), glutaraldehyde, and amine-rich interfacial surface. AchE was immobilized on the MP surface via Schiff's base formation, and the enzyme-modified MPs were deposited on the working electrode using a magnet under the microfluidic channel. The bioelectrocatalytic reaction between AchE-ChOx bienzyme cascade and the ferrocenyl electron shuttle was successfully used to detect OP with the developed microchip. This provides a self-contained and relatively easy method for OP detection. It requires minimal time and a small sample size, and has potential analytic applications in pesticides and chemical warfare agents.

Copper(II) complex of new non-innocent O-aminophenol-based ligand as biomimetic model for galactose oxidase enzyme in aerobic oxidation of alcohols

NASA Astrophysics Data System (ADS)

Safaei, Elham; Bahrami, Hadiseh; Pevec, Andrej; Kozlevčar, Bojan; Jagličić, Zvonko

2017-04-01

Mononuclear copper(II) complex of tetra-dentate o-aminophenol-based ligand (H2LBAPP) has been synthesized and characterized. The three dentate precursor (HLBAP) of the final ligand was synthesized first, while the title four-dentate copper bound ligand was synthesized in situ, isolated only in the final copper species [CuLBAPP]. This copper coordination complex reveals a distorted square-planar geometry around the copper(II) centre by one oxygen and three nitrogen atoms from the coordinating ligand. The ligand is thus twice deprotonated via hydroxy and amine groups. The complex is red, non-typical for copper(II), but the effective magnetic moment of 1.86 B M. and a single isotropic symmetry EPR signal with g 2.059 confirm a S = 1/2 diluted spin system, without copper-copper magnetic coupling. Electrochemical oxidation of this complex yields the corresponding Cu(II)-phenyl radical species. Finally, the title complex CuLBAPP has shown good and selective catalytic activity towards alcohol to aldehyde oxidation, at aerobic room temperature conditions, for a set of different alcohols.

Fungal Laccases and Their Applications in Bioremediation

PubMed Central

Viswanath, Buddolla; Rajesh, Bandi; Janardhan, Avilala; Kumar, Arthala Praveen; Narasimha, Golla

2014-01-01

Laccases are blue multicopper oxidases, which catalyze the monoelectronic oxidation of a broad spectrum of substrates, for example, ortho- and para-diphenols, polyphenols, aminophenols, and aromatic or aliphatic amines, coupled with a full, four-electron reduction of O2 to H2O. Hence, they are capable of degrading lignin and are present abundantly in many white-rot fungi. Laccases decolorize and detoxify the industrial effluents and help in wastewater treatment. They act on both phenolic and nonphenolic lignin-related compounds as well as highly recalcitrant environmental pollutants, and they can be effectively used in paper and pulp industries, textile industries, xenobiotic degradation, and bioremediation and act as biosensors. Recently, laccase has been applied to nanobiotechnology, which is an increasing research field, and catalyzes electron transfer reactions without additional cofactors. Several techniques have been developed for the immobilization of biomolecule such as micropatterning, self-assembled monolayer, and layer-by-layer techniques, which immobilize laccase and preserve their enzymatic activity. In this review, we describe the fungal source of laccases and their application in environment protection. PMID:24959348

Quantitation of plasma thiamine, related metabolites and plasma protein oxidative damage markers in children with autism spectrum disorder and healthy controls.

PubMed

Anwar, Attia; Marini, Marina; Abruzzo, Provvidenza Maria; Bolotta, Alessandra; Ghezzo, Alessandro; Visconti, Paola; Thornalley, Paul J; Rabbani, Naila

2016-11-01

To assess thiamine and related metabolite status by analysis of plasma and urine in autistic children and healthy controls, correlations to clinical characteristics and link to plasma protein markers of oxidative damage. 27 children with autism (21 males and 6 females) and 21 (15 males and 6 females) age-matched healthy control children were recruited. The concentration of thiamine and related phosphorylated metabolites in plasma and urine and plasma protein content of dityrosine, N-formylkynurenine and 3-nitrotyrosine was determined. Plasma thiamine and thiamine monophosphate concentrations were similar in both study groups (median [lower-upper quartile]): autistic children - 6.60 nM (4.48-8.91) and 7.00 nM (5.51-8.55), and healthy controls - 6.82 nM (4.47-7.02) and 6.82 nM (5.84-8.91), respectively. Thiamine pyrophosphate (TPP) was decreased 24% in autistic children compared to healthy controls: 6.82 nM (5.81-8.52) versus 9.00 nM (8.41-10.71), p < .01. Urinary excretion of thiamine and fractional renal clearance of thiamine did not change between the groups. No correlation was observed between clinical markers and the plasma and urine thiamine concentration. Plasma protein dityrosine content was increased 88% in ASD. Other oxidative markers were unchanged. Autistic children had normal plasma and urinary thiamine levels whereas plasma TPP concentration was decreased. The latter may be linked to abnormal tissue handling and/or absorption from gut microbiota of TPP which warrants further investigation. Increased plasma protein dityrosine may reflect increased dual oxidase activity in response to change in mucosal immunity and host-microbe homeostasis.

Random and independent sampling of endogenous tryptic peptides from normal human EDTA plasma by liquid chromatography micro electrospray ionization and tandem mass spectrometry.

PubMed

Dufresne, Jaimie; Florentinus-Mefailoski, Angelique; Ajambo, Juliet; Ferwa, Ammara; Bowden, Peter; Marshall, John

2017-01-01

Normal human EDTA plasma samples were collected on ice, processed ice cold, and stored in a freezer at - 80 °C prior to experiments. Plasma test samples from the - 80 °C freezer were thawed on ice or intentionally warmed to room temperature. Protein content was measured by CBBR binding and the release of alcohol soluble amines by the Cd ninhydrin assay. Plasma peptides released over time were collected over C18 for random and independent sampling by liquid chromatography micro electrospray ionization and tandem mass spectrometry (LC-ESI-MS/MS) and correlated with X!TANDEM. Fully tryptic peptides by X!TANDEM returned a similar set of proteins, but was more computationally efficient, than "no enzyme" correlations. Plasma samples maintained on ice, or ice with a cocktail of protease inhibitors, showed lower background amounts of plasma peptides compared to samples incubated at room temperature. Regression analysis indicated that warming plasma to room temperature, versus ice cold, resulted in a ~ twofold increase in the frequency of peptide identification over hours-days of incubation at room temperature. The type I error rate of the protein identification from the X!TANDEM algorithm combined was estimated to be low compared to a null model of computer generated random MS/MS spectra. The peptides of human plasma were identified and quantified with low error rates by random and independent sampling that revealed 1000s of peptides from hundreds of human plasma proteins from endogenous tryptic peptides.

Small Molecular, Macromolecular and Cellular Chloramines React with Thiocyanate to Give the Human Defense Factor Hypothiocyanite†

PubMed Central

Xulu, Bheki A.; Ashby, Michael T.

2010-01-01

Thiocyanate reacts non-catalytically with myeloperoxidase-derived HOCl to produce hypothiocyanite (OSCN−), thereby potentially limiting the propensity of HOCl to inflict host tissue damage that can lead to inflammatory diseases. However, the efficiency with which SCN− captures HOCl in vivo depends on the concentration of SCN− relative to other chemical targets. In blood plasma, where the concentration of SCN− is relatively low, proteins may be the principal initial targets of HOCl, and chloramines are a significant product. Chloramines eventually decompose to irreversibly damage proteins. In the present study, we demonstrate that SCN− reacts efficiently with chloramines in small molecules, in proteins, and in Escherichia coli cells to give OSCN− and the parent amine. Remarkably, OSCN− reacts faster than SCN− with chloramines. These reactions of SCN− and OSCN− with chloramines may repair some of the damage that is inflicted on protein amines by HOCl. Our observations are further evidence for the importance of secondary reactions during the redox cascades that are associated with oxidative stress by hypohalous acids. PMID:20085320

Poly-l-Lactic Acid Nanofiber-Polyamidoamine Hydrogel Composites: Preparation, Properties, and Preliminary Evaluation as Scaffolds for Human Pluripotent Stem Cell Culturing.

PubMed

Gualandi, Chiara; Bloise, Nora; Mauro, Nicolò; Ferruti, Paolo; Manfredi, Amedea; Sampaolesi, Maurilio; Liguori, Anna; Laurita, Romolo; Gherardi, Matteo; Colombo, Vittorio; Visai, Livia; Focarete, Maria Letizia; Ranucci, Elisabetta

2016-10-01

Electrospun poly-l-lactic acid (PLLA) nanofiber mats carrying surface amine groups, previously introduced by nitrogen atmospheric pressure nonequilibrium plasma, are embedded into aqueous solutions of oligomeric acrylamide-end capped AGMA1, a biocompatible polyamidoamine with arg-gly-asp (RGD)-reminiscent repeating units. The resultant mixture is finally cured giving PLLA-AGMA1 hydrogel composites that absorb large amounts of water and, in the swollen state, are translucent, soft, and pliable, yet as strong as the parent PLLA mat. They do not split apart from each other when swollen in water and remain highly flexible and resistant, since the hydrogel portion is covalently grafted onto the PLLA nanofibers via the addition reaction of the surface amine groups to a part of the terminal acrylic double bonds of AGMA1 oligomers. Preliminary tested as scaffolds, the composites prove capable of maintaining short-term undifferentiated cultures of human pluripotent stem cells in feeder-free conditions. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Important roles of enthalpic and entropic contributions to CO2 capture from simulated flue gas and ambient air using mesoporous silica grafted amines.

PubMed

Alkhabbaz, Mustafa A; Bollini, Praveen; Foo, Guo Shiou; Sievers, Carsten; Jones, Christopher W

2014-09-24

The measurement of isosteric heats of adsorption of silica supported amine materials in the low pressure range (0-0.1 bar) is critical for understanding the interactions between CO2 and amine sites at low coverage and hence to the development of efficient amine adsorbents for CO2 capture from flue gas and ambient air. Heats of adsorption for an array of silica-supported amine materials are experimentally measured at low coverage using a Calvet calorimeter equipped with a customized dosing manifold. In a series of 3-aminopropyl-functionalized silica materials, higher amine densities resulted in higher isosteric heats of adsorption, clearly showing that the density/proximity of amine sites can influence the amine efficiency of adsorbents. In a series of materials with fixed amine loading but different amine types, strongly basic primary and secondary amine materials are shown to have essentially identical heats of adsorption near 90 kJ/mol. However, the adsorption uptakes vary substantially as a function of CO2 partial pressure for different primary and secondary amines, demonstrating that entropic contributions to adsorption may play a key role in adsorption at secondary amine sites, making adsorption at these sites less efficient at the low coverages that are important to the direct capture of CO2 from ambient air. Thus, while primary amines are confirmed to be the most effective amine types for CO2 capture from ambient air, this is not due to enhanced enthalpic contributions associated with primary amines over secondary amines, but may be due to unfavorable entropic factors associated with organization of the second alkyl chain on the secondary amine during CO2 adsorption. Given this hypothesis, favorable entropic factors may be the main reason primary amine based adsorbents are more effective under air capture conditions.

Exercise Training, NADPH Oxidase p22phox Gene Polymorphisms, and Hypertension

PubMed Central

FEAIRHELLER, DEBORAH L.; BROWN, MICHAEL D.; PARK, JOON-YOUNG; BRINKLEY, TINA E.; BASU, SAMAR; HAGBERG, JAMES M.; FERRELL, ROBERT E.; FENTY-STEWART, NICOLA M.

2010-01-01

Introduction Oxidative stress that is mediated through NADPH oxidase activity plays a role in the pathology of hypertension, and aerobic exercise training reduces NADPH oxidase activity. The involvement of genetic variation in the p22phox (CYBA) subunit genes in individual oxidative stress responses to aerobic exercise training has yet to be examined in Pre and Stage 1 hypertensives. Methods Ninety-four sedentary Pre and Stage 1 hypertensive adults underwent 6 months of aerobic exercise training at a level of 70% V̇O2max to determine whether the CYBA polymorphisms, C242T and A640G, were associated with changes in urinary 8-iso-prostaglandin F2α (8-iso-PGF2α), urinary nitric oxide metabolites (NOx), and plasma total antioxidant capacity (TAC). Results Demographic and subject characteristics were similar among genotype groups for both polymorphisms. At baseline, a significant (P = 0.03) difference among the C2424T genotype groups in 8-iso-PGF2α levels was detected, with the TT homozygotes having the lowest levels and the CC homozygotes having the highest levels. However, no differences were found at baseline between the A640G genotype groups. After 6 months of aerobic exercise training, there was a significant increase in V̇O2max (P < 0.0001) in the entire study population. In addition, there were significant increases in both urinary 8-iso-PGF2α (P = 0.002) and plasma TAC (P = 0.03) levels and a significant decrease in endogenous urinary NOx (P < 0.0001). Overall, aerobic exercise training elicited no significant differences among genotype groups in either CYBA variant for any of the oxidative stress variables. Conclusions We found that compared with CYBA polymorphisms C242T and A640G, it was aerobic exercise training that had the greatest influence on the selected biomarkers; furthermore, our results suggest that the C242T CYBA variant influences baseline levels of urinary 8-iso-PGF2α but not the aerobic exercise-induced responses. PMID:19516159

Serotonin modulates substance P-induced plasma extravasation and vasodilatation in rat skin by an action through capsaicin-sensitive primary afferent nerves.

PubMed

Khalil, Z; Helme, R D

1990-09-17

Using a blister model of inflammation in the rat hind footpad, the present study was undertaken to examine the ability of serotonin (5-HT) to modulate an inflammatory reaction manifested as plasma extravasation and vasodilatation induced by the neuropeptide substance P (SP). In addition, the role of primary afferent sensory nerve fibres in these modulatory effects was studied in capsaicin pretreated rats. Using a protocol of simultaneous perfusion of amine and peptide over the blister base, no major modulatory effect was observed. On the other hand, using a protocol of sequential perfusion, 5-HT was found to extend the plasma extravasation and vasodilatation responses to SP. 5-HT maintained the plasma extravasation response to SP after cessation of stimulation (during the post-stimulation period). On the other hand, it extended the vasodilatation response to SP during the actual stimulation period by preventing the occurrence of tachyphylaxis. These modulatory effects were absent in capsacin-pretreated rats. The present study provides evidence for the first time in vivo to suggest that serotonin can modulate an inflammatory response to SP via a mechanism that involves capsaicin-sensitive sensory fibres.

Antibacterial performance on plasma polymerized heptylamine films loaded with silver nanoparticles

NASA Astrophysics Data System (ADS)

Lin, Yu-Chun; Lin, Chia-Chun; Lin, Chih-Hao; Wang, Meng-Jiy

2017-01-01

The antibacterial performance of the plasma-polymerized (pp) heptylamine thin films loaded with silver nanoparticles was evaluated against the colonization of Escherichia coli and Staphylococcus aureus. The properties including the thickness and chemical composition of the as deposited HApp films were modulated by adjusting plasma parameters. The acquired results showed that the film thickness was controlled in the range of 20 to 400 nm by adjusting deposition time. The subsequent immersion of the HApp thin films in silver nitrate solutions result in the formation of amine-metal complexes, in which the silver nanoparticles were reduced directly on the matrices to form Ag@HApp. The reduction reaction of silver was facilitated by applying NaBH4 as a reducing agent. The results of physicochemical analyses including morphological analysis and ellipsometry revealed that the silver nanoparticles were successfully reduced on the HApp films, and the amount of reduced silver was closely associated which the thickness of the plasma-polymerized films, the concentration of applied metal ions solutions, and the time of immobilization. Regarding the antibacterial performance, the Ag@HApp films reduced by NaBH4 showed antibacterial abilities of 70.1 and 68.2% against E. coli and S. aureus, respectively.

Analgesic effects of β-phenylethylamine and various methylated derivatives in mice.

PubMed

Mosnaim, Aron D; Hudzik, Thomas; Wolf, Marion E

2014-09-01

Administration of β-phenylethylamine (PEA), the simplest endogenous neuroamine, and various methylated PEA derivatives including α-methyl PEA (amphetamine, AMP) elicits analgesia in mice. Five or 20 min after intraperitoneal PEA injection of as little as 6 mg/kg resulted in an increased latency response time (from 2.4 ± 0.4 to 8.5 ± 2.3 or 7.0 ± 3.0 s, respectively) to the thermal stimulus (hot-plate test), which reached statistical significance at the 15 mg/kg (20 min; 13.1 ± 0.4 s) or 25 mg/kg dose (5 min; 15.3 ± 4.1 s). This PEA effect, was dose-dependent (albeit non-linear: 6, 12, 15, 25, 50 and 100 mg/kg), reached the cut-off time of 45 s at the upper PEA dose (5 min), and it was consistently enhanced by pretreatment with the monoamine oxidase inhibitor pargyline (P). Methylated PEA derivatives (15 and 100 mg/kg dose) produced various degrees of analgesia (in decreasing order p-Me PEA > PEA > N,N-diMe PEA > N-Me PEA) which, likewise to PEA itself, were consistently increased by P and declined over time (mice tested 5, 20 and 60 min after amine injection); small but statistically significant o- and β-Me PEA antinociceptive effects (5 min) were observed only at the higher dose (in the presence of P for β-Me PEA). A small analgesic effect was observed after the administration of AMP (5 or 10 mg/kg) which failed, even after P, to reach statistically significance. Independent of the amine and concentration tested, individual compound's antinociceptive properties were reliably increased by P (exception of AMP), decreased by reserpine (R) or haloperidol (H), and remained essentially unchanged after naloxone (N) administration suggesting the involvement of catecholamines, but not opioid peptides, in their observed analgesic effects. Injection of P + N produced results similar to those seen after P alone. Under the experimental conditions described neither P, R, H or N had any effects by themselves. These findings suggest additional understanding of the mechanism of action responsible for the analgesic effects of these amines would be of interest, leading further to controlled studies on their alleged usefulness as weight reducing agents and sport performance enhancers.

Hemin potentiates nitric oxide-mediated nitrosation of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) to 2-nitrosoamino-3-methylimidazo[4,5-f]quinoline.

PubMed

Lakshmi, Vijaya M; Clapper, Margie L; Chang, Wen-Chi; Zenser, Terry V

2005-03-01

Heme has been reported to be an important contributor to endogenous N-nitrosation within the colon and to the enhanced incidence of colon cancer observed with increased intake of red meat. This study uses the heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) as a target to evaluate hemin potentiation of nitric oxide (NO)-mediated nitrosation. Formation of 14C-2-nitrosoamino-3-methylimidazo[4,5-f]quinoline (N-NO-IQ) was monitored by HPLC following incubation of 10 microM IQ with the NO donor spermine NONOate (1.2 microM NO/min) at pH 7.4 in the presence or absence of hemin. N-NO-IQ formation due to autoxidation of NO was at the limit of detection (0.1 microM) and increased 22-fold in the presence of 10 microM hemin and an in situ system for generating H2O2 (glucose oxidase/glucose). A linear increase in N-NO-IQ formation was observed from 1 to 10 microM hemin. Significant nitrosamine formation occurred at fluxes of NO and H2O2 as low as 0.024 and 0.25 microM/min, respectively. Potentiation by hemin was not affected by a 400-fold excess flux of H2O2 over NO or a 4.8-fold excess flux of NO over H2O2. Reactive nitrogen species produced by hemin potentiation had a 46-fold greater affinity for IQ than those produced by autoxidation. Azide inhibited autoxidation, suggesting involvement of the nitrosonium ion, NO+. Hemin potentiation was inhibited by NADH, but not azide, suggesting oxidative nitrosylation with NO2* or a NO2*-like species. IQ and 2,3-diaminonaphthylene were much better targets for nitrosation than the secondary amine morpholine. Apc(min) mice with dextran sulfate sodium-induced colitis demonstrated increased levels of urinary nitrite and nitrate consistent with increased expression of iNOS and NO synthesis. As reported previously, identical conditions increased fecal N-nitroso compounds. Thus, hemin potentiation of NO-mediated nitrosation of heterocyclic amines provides a testable mechanism by which red meat consumption can generate N-nitroso compounds and initiate colon cancer under inflammatory conditions, such as colitis.

Trimethylaluminum and borane complexes of primary amines.

PubMed

Németh, Balázs; Guégan, Jean-Paul; Veszprémi, Tamás; Guillemin, Jean-Claude

2013-01-07

Trimethylaluminum (TMA) complexes of methyl-, n-propyl-, cyclopropyl-, allyl-, and propargylamine were synthesized and their experimental properties and theoretical characteristics were compared with the respective amine-borane analogues. The amine ligand of an amine-TMA Lewis acid-base complex can be easily changed by another amine through a 2:1 amine-TMA intermediate in pentane at room temperature. The exchange of the same ligands in the case of amine-boranes requires remarkably more time in line with the calculated relative energy of the respective transition state. The (1)H and (13)C NMR experiments examining the addition of one or more equivalent of amine to the respective Lewis acid-base complex conclude in the fast exchange of the amine ligand in the NMR time scale only in the cases of amine-TMA complexes, which could also be caused by similar 2:1 complexes. However, in gas phase, only 1:1 amine-TMA complexes are present as evidenced by ultraviolet photoelectron spectroscopy (UPS). The observed UP spectra, which are the first recorded photoelectron spectra of primary amine-TMA compounds, indicate that the stabilization effect of the lone electron pair of nitrogen atom in amines during the borane complexation is stronger than that of the TMA complexation. In line with this observation, the destabilization of the σ(Al-C) orbitals is lower than that of σ(B-H) orbitals during the formation of amine-TMA and amine-borane complexes, respectively. As showed by theoretical calculations, the CH(4) elimination of the studied amine-TMA complexes is exothermic, indicating the possibility of using these compounds in metal organic chemical vapor deposition techniques (MOCVD). On the other hand, our experimental conditions avoid this methane elimination and constitutes the first procedure employing distillation to isolate primary amine-TMA complexes.

Selective cytotoxicity of an oxygen-radical-generating enzyme conjugated to a monoclonal antibody.

PubMed

Battelli, M G; Abbondanza, A; Tazzari, P L; Dinota, A; Rizzi, S; Grassi, G; Gobbi, M; Stirpe, F

1988-07-01

The monoclonal antibody 8A, which recognizes a human plasma cell-associated antigen, was covalently linked to xanthine oxidase in a conjugate maintaining both immunological and enzymatic properties. A significant degree of target cell lysis was obtained at an enzyme concentration that was ineffective on non-target cells and on myeloid staminal cells (CFU-GM). The cytotoxic activity was abolished by an excess of antibody, by allopurinol and by superoxide dismutase and catalase. A possible use of the conjugate for bone marrow purging in multiple myeloma patients is suggested.

Impaired Monoamine and Organic Cation Uptake in Choroid Plexus in Mice with Targeted Disruption of the Plasma Membrane Monoamine Transporter (Slc29a4) Gene*

PubMed Central

Duan, Haichuan; Wang, Joanne

2013-01-01

The choroid plexus (CP) forms the blood-cerebrospinal fluid (CSF) barrier and protects the brain from circulating metabolites, drugs, and toxins. The plasma membrane monoamine transporter (PMAT, SLC29A4) is a new polyspecific organic cation transporter that transports a wide variety of organic cations including biogenic amines, cationic drugs, and neurotoxins. PMAT is known to be expressed in the CP, but its specific role in CP transport of organic cations has not been clearly defined. Here we showed that PMAT transcript is highly expressed in human and mouse CPs, whereas transcripts of other functionally related transporters are minimally expressed in the CPs. Immunofluorescence staining further revealed that PMAT protein is localized to the apical (CSF-facing) membrane of the CP epithelium, consistent with a role of transporting organic cations from the CSF into CP epithelial cells. To further evaluate the role of PMAT in the CP, mice with targeted deletion of the Slc29a4 gene were generated and validated. Although Pmat−/− mice showed no overt abnormalities, the uptake of monoamines and the neurotoxin 1-methyl-4-phenylpyridinium was significantly reduced in CP tissues isolated from the knock-out mice. Together, our data demonstrated that PMAT is a major transporter for CP uptake of bioactive amines and xenobiotic cations. By removing its substrates from the CSF, PMAT may play an important role in protecting the brain from cationic neurotoxins and other potentially toxic organic cations. PMID:23255610

Oxidative stress induces protein and DNA radical formation in follicular dendritic cells (FDCs) of the germinal center and modulates its cell death patterns in late sepsis

PubMed Central

Chatterjee, Saurabh; Lardinois, Olivier; Bhattacharjee, Suchandra; Tucker, Jeff; Corbett, Jean; Deterding, Leesa; Ehrenshaft, Marilyn; Bonini, Marcelo; Mason, Ronald P.

2011-01-01

Profound depletion of follicular dendritic cells (FDCs) is a hallmark of sepsis-like syndrome, but the exact causes for the ensuing cell death are unknown. The cell death-driven depletion contributes to immunoparalysis and is responsible for most of the morbidity and mortality in sepsis. Here we have utilized immuno-spin trapping, a method for detection of free radical formation, to detect oxidative stress-induced protein and DNA radical adducts in FDCs isolated from the spleen of septic mice and human tonsil-derived HK cells, a subtype of germinal center FDCs, to study their role in FDC depletion. At 24 h post-LPS administration, protein radical formation and oxidation was significantly elevated in vivo and in HK cells as shown by ELISA and confocal microscopy. The xanthine oxidase inhibitor allopurinol and the iron chelator desferrioxamine significantly decreased the formation of protein radicals, suggesting the role of xanthine oxidase and Fenton-like chemistry in radical formation. Protein and DNA radical formation correlated mostly with apoptotic features at 24 h and necrotic morphology of all the cell types studied at 48 h with concomitant inhibition of caspase-3. The cytotoxity of FDCs resulted in decreased CD45R/CD138+ve plasma cell numbers, indicating a possible defect in B cell differentiation. In one such mechanism, radical formation initiated by xanthine oxidase formed protein and DNA radicals which may lead to cell death of germinal center FDCs. PMID:21215311

Selective inhibition of extracellular oxidants liberated from human neutrophils--A new mechanism potentially involved in the anti-inflammatory activity of hydroxychloroquine.

PubMed

Jančinová, Viera; Pažoureková, Silvia; Lucová, Marianna; Perečko, Tomáš; Mihalová, Danica; Bauerová, Katarína; Nosáľ, Radomír; Drábiková, Katarína

2015-09-01

Hydroxychloroquine is used in the therapy of rheumatoid arthritis or lupus erythematosus. Although these diseases are often accompanied by activation of neutrophils, there are still few data relating to the impact of hydroxychloroquine on these cells. We investigated the effect of orally administered hydroxychloroquine on neutrophil oxidative burst in rats with adjuvant arthritis. In human neutrophils, extra- and intracellular formation of oxidants, mobilisation of intracellular calcium and the phosphorylation of proteins regulating NADPH oxidase assembly were analysed. Administration of hydroxychloroquine decreased the concentration of oxidants in blood of arthritic rats. The inhibition was comparable with the reference drug methotrexate, yet it was not accompanied by a reduction in neutrophil count. When both drugs were co-applied, the effect became more pronounced. In isolated human neutrophils, treatment with hydroxychloroquine resulted in reduced mobilisation of intracellular calcium, diminished concentration of external oxidants and in decreased phosphorylation of Ca(2+)-dependent protein kinase C isoforms PKCα and PKCβII, which regulate activation of NADPH oxidase on plasma membrane. On the other hand, no reduction was observed in intracellular oxidants or in the phosphorylation of p40(phox) and PKCδ, two proteins directing the oxidase assembly to intracellular membranes. Hydroxychloroquine reduced neutrophil-derived oxidants potentially involved in tissue damage and protected those capable to suppress inflammation. The observed effects may represent a new mechanism involved in the anti-inflammatory activity of this drug. Copyright © 2015 Elsevier B.V. All rights reserved.

A site-directed mutagenesis analysis of tNOX functional domains

NASA Technical Reports Server (NTRS)

Chueh, Pin-Ju; Morre, Dorothy M.; Morre, D. James

2002-01-01

Constitutive NADH oxidase proteins of the mammalian cell surface exhibit two different activities, oxidation of hydroquinones (or NADH) and protein disulfide-thiol interchange which alternate to yield oscillatory patterns with period lengths of 24 min. A drug-responsive tNOX (tumor-associated NADH oxidase) has a period length of about 22 min. The tNOX cDNA has been cloned and expressed. These two proteins are representative of cycling oxidase proteins of the plant and animal cell surface. In this report, we describe a series of eight amino acid replacements in tNOX which, when expressed in Escherichia coli, were analyzed for enzymatic activity, drug response and period length. Replacement sites selected include six cysteines that lie within the processed plasma membrane (34 kDa) form of the protein, and amino acids located in putative drug and adenine nucleotide (NADH) binding domains. The latter, plus two of the cysteine replacements, resulted in a loss of enzymatic activity. The recombinant tNOX with the modified drug binding site retained activity but the activity was no longer drug-responsive. The four remaining cysteine replacements were of interest in that both activity and drug response were retained but the period length for both NADH oxidation and protein disulfide-thiol interchange was increased from 22 min to 36 or 42 min. The findings confirm the correctness of the drug and adenine nucleotide binding motifs within the tNOX protein and imply a potential critical role of cysteine residues in determining the period length.

Effects of NADPH oxidase inhibitor on diabetic nephropathy in OLETF rats: the role of reducing oxidative stress in its protective property.

PubMed

Nam, Soo Min; Lee, Mi Young; Koh, Jang Hyun; Park, Jun Ho; Shin, Jang Yel; Shin, Young Goo; Koh, Sang Baek; Lee, Eun Young; Chung, Choon Hee

2009-02-01

Diabetic nephropathy is the most serious complication in diabetes mellitus. Oxidative stress via nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and vascular endothelial growth factor (VEGF) pathway play critical roles in the development of diabetic nephropathy. We evaluated the effects of apocynin, NADPH oxidase inhibitor on diabetic nephropathy in a type 2 diabetic rat model. Sixteen Otsuka Long Evans Tokushima Fatty (OLETF) rats and 9 Long Evans Tokushima Otsuka (LETO) were divided into the following three groups: LETO rats (n=9), control OLETF rats (n=7) and apocynin-treated OLETF rats (n=9). We examined body weights, plasma glucose levels, urinary albumin-creatinine ratio (ACR) and protein-creatinine ratio (PCR). At 50 weeks, experimental rats were sacrificed and their kidneys were extracted for hematoxylin eosin stain, immunohistochemical VEGF stain and VEGF mRNA real-time RT-PCR. To examine oxidative stress, we checked 24h urinary 8-OHdG (8-hydroxy-2'-deoxyguanosine) and MDA (malondialdehyde). Urinary protein and albumin excretions were reduced after apocynin treatment, though apocynin could not significantly decrease serum glucose levels. There were improvements of glomerular and mesangial expansion in the apocynin-treated OLETF rats. Apocynin significantly decreased optical density of glomerular VEGF expression in immunohistochemical stain and reduced the concentration of 24h urinary 8-OHdG and MDA. From these results, it was suggested that apocynin may have the potential to protect against diabetic nephropathy via amelioration of oxidative stress.

Smoking induces long-lasting effects through a monoamine-oxidase epigenetic regulation.

PubMed

Launay, Jean-Marie; Del Pino, Muriel; Chironi, Gilles; Callebert, Jacques; Peoc'h, Katell; Mégnien, Jean-Louis; Mallet, Jacques; Simon, Alain; Rendu, Francine

2009-11-23

Postulating that serotonin (5-HT), released from smoking-activated platelets could be involved in smoking-induced vascular modifications, we studied its catabolism in a series of 115 men distributed as current smokers (S), never smokers (NS) and former smokers (FS) who had stopped smoking for a mean of 13 years. 5-HT, monoamine oxidase (MAO-B) activities and amounts were measured in platelets, and 5-hydroxyindolacetic acid (5-HIAA)--the 5-HT/MAO catabolite--in plasma samples. Both platelet 5-HT and plasma 5-HIAA levels were correlated with the 10-year cardiovascular Framingham relative risk (P<0.01), but these correlations became non-significant after adjustment for smoking status, underlining that the determining risk factor among those taken into account in the Framingham risk calculation was smoking. Surprisingly, the platelet 5-HT content was similar in S and NS but lower in FS with a parallel higher plasma level of 5-HIAA in FS. This was unforeseen since MAO-B activity was inhibited during smoking (P<0.00001). It was, however, consistent with a higher enzyme protein concentration found in S and FS than in NS (P<0.001). It thus appears that MAO inhibition during smoking was compensated by a higher synthesis. To investigate the persistent increase in MAO-B protein concentration, a study of the methylation of its gene promoter was undertaken in a small supplementary cohort of similar subjects. We found that the methylation frequency of the MAOB gene promoter was markedly lower (P<0.0001) for S and FS vs. NS due to cigarette smoke-induced increase of nucleic acid demethylase activity. This is one of the first reports that smoking induces an epigenetic modification. A better understanding of the epigenome may help to further elucidate the physiopathology and the development of new therapeutic approaches to tobacco addiction. The results could have a larger impact than cardiovascular damage, considering that MAO-dependent 5-HT catabolism is also involved in addiction, predisposition to cancer, behaviour and mental health.

Atmospheric Amines and Ammonia Measured with a Chemical Ionization Mass Spectrometer (CIMS)

DOE Office of Scientific and Technical Information (OSTI.GOV)

You, Y.; Kanawade, V. P.; de Gouw, J. A.

We report ambient measurements of amines and ammonia with a fast response chemical ionization mass spectrometer (CIMS) in a Southeastern U.S. forest in Alabama and a moderately polluted Midwestern site during the summer. In the Alabama forest, mostly C3-amines (from pptv to tens of pptv) and ammonia (up to 2 ppbv) were detected on a daily basis. C3-amines and ammonia showed similar diurnal trends and temperature and wind direction dependences, and were not associated with transported CO and SO2 plumes. Consistent with temperature dependences, amine and ammonia in the gas and aerosol phases showed opposite diurnal trends, indicating gas-to-particle partitioningmore » of amines and ammonia. Temperature dependences also imply reversible processes of amines and ammonia evaporation from soil surfaces in daytime and deposition of amines and ammonia to soil surfaces at nighttime. Various amines (C1-C6) at the pptv level were observed in the transported biomass burning plumes, showing that biomass burning can be a substantial source of amines in the Southeast U.S. At the moderately polluted Kent site, higher concentrations of amines (C1-C6, from pptv to tens of pptv) and ammonia (up to 6 ppbv) were detected. Diurnal variations of C1- to C3-amines and ammonia were correlated with the ambient temperature. C4- to C6-amines showed abrupt increases during the nighttime, suggesting that they were emitted from local sources. These abundant amines and ammonia may in part explain the frequent new particle formation events reported from Kent. Lower amine concentrations at the rural forested site highlight the importance of constraining anthropogenic sources of amines.« less

Development of a general non-noble metal catalyst for the benign amination of alcohols with amines and ammonia.

PubMed

Cui, Xinjiang; Dai, Xingchao; Deng, Youquan; Shi, Feng

2013-03-11

The N-alkylation of amines or ammonia with alcohols is a valuable route for the synthesis of N-alkyl amines. However, as a potentially clean and economic choice for N-alkyl amine synthesis, non-noble metal catalysts with high activity and good selectivity are rarely reported. Normally, they are severely limited due to low activity and poor generality. Herein, a simple NiCuFeOx catalyst was designed and prepared for the N-alkylation of ammonia or amines with alcohol or primary amines. N-alkyl amines with various structures were successfully synthesized in moderate to excellent yields in the absence of organic ligands and bases. Typically, primary amines could be efficiently transformed into secondary amines and N-heterocyclic compounds, and secondary amines could be N-alkylated to synthesize tertiary amines. Note that primary and secondary amines could be produced through a one-pot reaction of ammonia and alcohols. In addition to excellent catalytic performance, the catalyst itself possesses outstanding superiority, that is, it is air and moisture stable. Moreover, the magnetic property of this catalyst makes it easily separable from the reaction mixture and it could be recovered and reused for several runs without obvious deactivation. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Requirement for Coenzyme Q in Plasma Membrane Electron Transport

NASA Astrophysics Data System (ADS)

Sun, I. L.; Sun, E. E.; Crane, F. L.; Morre, D. J.; Lindgren, A.; Low, H.

1992-12-01

Coenzyme Q is required in the electron transport system of rat hepatocyte and human erythrocyte plasma membranes. Extraction of coenzyme Q from the membrane decreases NADH dehydrogenase and NADH:oxygen oxidoreductase activity. Addition of coenzyme Q to the extracted membrane restores the activity. Partial restoration of activity is also found with α-tocopherylquinone, but not with vitamin K_1. Analogs of coenzyme Q inhibit NADH dehydrogenase and oxidase activity and the inhibition is reversed by added coenzyme Q. Ferricyanide reduction by transmembrane electron transport from HeLa cells is inhibited by coenzyme Q analogs and restored with added coenzyme Q10. Reduction of external ferricyanide and diferric transferrin by HeLa cells is accompanied by proton release from the cells. Inhibition of the reduction by coenzyme Q analogs also inhibits the proton release, and coenzyme Q10 restores the proton release activity. Trans-plasma membrane electron transport stimulates growth of serum-deficient cells, and added coenzyme Q10 increases growth of HeLa (human adenocarcinoma) and BALB/3T3 (mouse fibroblast) cells. The evidence is consistent with a function for coenzyme Q in a trans-plasma membrane electron transport system which influences cell growth.

Dissecting blue light signal transduction pathway in leaf epidermis using a pharmacological approach.

PubMed

Živanović, Branka D; Shabala, Lana I; Elzenga, Theo J M; Shabala, Sergey N

2015-10-01

Blue light signalling pathway in broad bean leaf epidermal cells includes key membrane transporters: plasma- and endomembrane channels and pumps of H (+) , Ca (2+) and K (+) ions, and plasma membrane redox system. Blue light signalling pathway in epidermal tissue isolated from the abaxial side of fully developed Vicia faba leaves was dissected by measuring the effect of inhibitors of second messengers on net K(+), Ca(2+) and H(+) fluxes using non-invasive ion-selective microelectrodes (the MIFE system). Switching the blue light on-off caused transient changes of the ion fluxes. The effects of seven groups of inhibitors were tested in this study: CaM antagonists, ATPase inhibitors, Ca(2+) anatagonists or chelators, agents affecting IP3 formation, redox system inhibitors, inhibitors of endomembrane Ca(2+) transport systems and an inhibitor of plasma membrane Ca(2+)-permeable channels. Most of the inhibitors had a significant effect on steady-state (basal) net fluxes, as well as on the magnitude of the transient ion flux responses to blue light fluctuations. The data presented in this study suggest that redox signalling and, specifically, plasma membrane NADPH oxidase and coupled Ca(2+) and K(+) fluxes play an essential role in blue light signal transduction.

Allylic aminations with hindered secondary amine nucleophiles catalyzed by heterobimetallic Pd-Ti complexes.

PubMed

Walker, Whitney K; Anderson, Diana L; Stokes, Ryjul W; Smith, Stacey J; Michaelis, David J

2015-02-06

Phosphinoamide-scaffolded heterobimetallic palladium-titanium complexes are highly effective catalysts for allylic aminations of allylic chlorides with hindered secondary amine nucleophiles. Three titanium-containing ligands are shown to assemble active catalysts in situ and enable catalysis at room temperature. A variety of sterically bulky secondary amines are efficiently allylated in high yields with as little as 1 mol % palladium catalyst. Piperidine and pyrrolidine products are also efficiently generated via intramolecular aminations with hindered amine nucleophiles.

Amine dehydrogenases: efficient biocatalysts for the reductive amination of carbonyl compounds.

PubMed

Knaus, Tanja; Böhmer, Wesley; Mutti, Francesco G

2017-01-21

Amines constitute the major targets for the production of a plethora of chemical compounds that have applications in the pharmaceutical, agrochemical and bulk chemical industries. However, the asymmetric synthesis of α-chiral amines with elevated catalytic efficiency and atom economy is still a very challenging synthetic problem. Here, we investigated the biocatalytic reductive amination of carbonyl compounds employing a rising class of enzymes for amine synthesis: amine dehydrogenases (AmDHs). The three AmDHs from this study - operating in tandem with a formate dehydrogenase from Candida boidinii (Cb-FDH) for the recycling of the nicotinamide coenzyme - performed the efficient amination of a range of diverse aromatic and aliphatic ketones and aldehydes with up to quantitative conversion and elevated turnover numbers (TONs). Moreover, the reductive amination of prochiral ketones proceeded with perfect stereoselectivity, always affording the ( R )-configured amines with more than 99% enantiomeric excess. The most suitable amine dehydrogenase, the optimised catalyst loading and the required reaction time were determined for each substrate. The biocatalytic reductive amination with this dual-enzyme system (AmDH-Cb-FDH) possesses elevated atom efficiency as it utilizes the ammonium formate buffer as the source of both nitrogen and reducing equivalents. Inorganic carbonate is the sole by-product.

A Biochemical Approach to Study the Role of the Terminal Oxidases in Aerobic Respiration in Shewanella oneidensis MR-1

PubMed Central

Le Laz, Sébastien; Kpebe, Arlette; Bauzan, Marielle; Lignon, Sabrina; Rousset, Marc; Brugna, Myriam

2014-01-01

The genome of the facultative anaerobic γ-proteobacterium Shewanella oneidensis MR-1 encodes for three terminal oxidases: a bd-type quinol oxidase and two heme-copper oxidases, a A-type cytochrome c oxidase and a cbb 3-type oxidase. In this study, we used a biochemical approach and directly measured oxidase activities coupled to mass-spectrometry analysis to investigate the physiological role of the three terminal oxidases under aerobic and microaerobic conditions. Our data revealed that the cbb 3-type oxidase is the major terminal oxidase under aerobic conditions while both cbb 3-type and bd-type oxidases are involved in respiration at low-O2 tensions. On the contrary, the low O2-affinity A-type cytochrome c oxidase was not detected in our experimental conditions even under aerobic conditions and would therefore not be required for aerobic respiration in S. oneidensis MR-1. In addition, the deduced amino acid sequence suggests that the A-type cytochrome c oxidase is a ccaa 3-type oxidase since an uncommon extra-C terminal domain contains two c-type heme binding motifs. The particularity of the aerobic respiratory pathway and the physiological implication of the presence of a ccaa 3-type oxidase in S. oneidensis MR-1 are discussed. PMID:24466040

Reducing peanut allergens by high pressure combined with polyphenol oxidase

NASA Astrophysics Data System (ADS)

Chung, Si-Yin; Houska, Milan; Reed, Shawndrika

2013-12-01

Polyphenol oxidase (PPO) has been shown to reduce major peanut allergens. Since high pressure (HP) can increase enzyme activity, we postulated that further reduction of peanut allergens can be achieved through HP combined with PPO. Peanut extracts containing caffeic acid were treated with each of the following: (1) HP; (2) HP+PPO; (3) PPO; and (4) none. HP was conducted at 300 and 500 MPa, each for 3 and 10 min, 37 °C. After treatment, SDS-PAGE was performed and allergenic capacity (IgE binding) was determined colorimetrically in inhibition enzyme-linked immunosorbent assay and Western blots, using a pooled plasma from peanut-allergic patients. Data showed that HP alone had no effect on major peanut allergens. However, HP at 500 MPa combined with PPO (HP500/PPO) induced a higher (approximately twofold) reduction of major peanut allergens and IgE binding than PPO alone or HP300/PPO. There was no difference between treatment times. We concluded that HP500/PPO at 3-min enhanced a twofold reduction of the allergenic capacity of peanut extracts, as compared to PPO itself.

Metal and base free synthesis of primary amines via ipso amination of organoboronic acids mediated by [bis(trifluoroacetoxy)iodo]benzene (PIFA).

PubMed

Chatterjee, Nachiketa; Goswami, Avijit

2015-08-07

A metal and base free synthesis of primary amines has been developed at ambient temperature through ipso amination of diversely functionalized organoboronic acids, employing a combination of [bis(trifluoroacetoxy)iodo]benzene (PIFA)-N-bromosuccinimide (NBS) and methoxyamine hydrochloride as the aminating reagent. The amines were primarily obtained as their trifluoroacetate salts which on subsequent aqueous alkaline work up provided the corresponding free amines. The combination of PIFA-NBS is found to be the mildest choice compared to the commonly used strong bases (e.g. n-BuLi, Cs2CO3) for activating the aminating agent. The reaction is expected to proceed via activation of the aminating reagent followed by B-N 1,2-aryl migration.

Generation of reactive oxygen species (ROS) is a key factor for stimulation of macrophage proliferation by ceramide 1-phosphate

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arana, Lide; Gangoiti, Patricia; Ouro, Alberto

2012-02-15

We previously demonstrated that ceramide 1-phosphate (C1P) is mitogenic for fibroblasts and macrophages. However, the mechanisms involved in this action were only partially described. Here, we demonstrate that C1P stimulates reactive oxygen species (ROS) formation in primary bone marrow-derived macrophages, and that ROS are required for the mitogenic effect of C1P. ROS production was dependent upon prior activation of NADPH oxidase by C1P, which was determined by measuring phosphorylation of the p40phox subunit and translocation of p47phox from the cytosol to the plasma membrane. In addition, C1P activated cytosolic calcium-dependent phospholipase A{sub 2} and protein kinase C-{alpha}, and NADPH oxidasemore » activation was blocked by selective inhibitors of these enzymes. These inhibitors, and inhibitors of ROS production, blocked the mitogenic effect of C1P. By using BHNB-C1P (a photolabile caged-C1P analog), we demonstrate that all of these C1P actions are caused by intracellular C1P. It can be concluded that the enzyme responsible for C1P-stimulated ROS generation in bone marrow-derived macrophages is NADPH oxidase, and that this enzyme is downstream of PKC-{alpha} and cPLA{sub 2}-{alpha} in this pathway. -- Highlights: Black-Right-Pointing-Pointer Ceramide 1-phosphate (C1P) stimulates reactive oxygen species (ROS) formation. Black-Right-Pointing-Pointer The enzyme responsible for ROS generation by C1P in macrophages is NADPH oxidase. Black-Right-Pointing-Pointer NADPH oxidase lies downstream of cPLA{sub 2}-{alpha} and PKC-{alpha} in this pathway. Black-Right-Pointing-Pointer ROS generation is essential for the stimulation of macrophage proliferation by C1P.« less

Low-Dose Dextromethorphan, a NADPH Oxidase Inhibitor, Reduces Blood Pressure and Enhances Vascular Protection in Experimental Hypertension

PubMed Central

Wu, Tao-Cheng; Chao, Chih-Yu; Lin, Shing-Jong; Chen, Jaw-Wen

2012-01-01

Background Vascular oxidative stress may be increased with age and aggravate endothelial dysfunction and vascular injury in hypertension. This study aimed to investigate the effects of dextromethorphan (DM), a NADPH oxidase inhibitor, either alone or in combination treatment, on blood pressure (BP) and vascular protection in aged spontaneous hypertensive rats (SHRs). Methodology/Principal Findings Eighteen-week-old WKY rats and SHRs were housed for 2 weeks. SHRs were randomly assigned to one of the 12 groups: untreated; DM monotherapy with 1, 5 or 25 mg/kg/day; amlodipine (AM, a calcium channel blocker) monotherapy with 1 or 5 mg/kg/day; and combination therapy of DM 1, 5 or 25 mg/kg/day with AM 1 or 5 mg/kg/day individually for 4 weeks. The in vitro effects of DM were also examined. In SHRs, AM monotherapy dose-dependently reduced arterial systolic BP. DM in various doses significantly and similarly reduced arterial systolic BP. Combination of DM with AM gave additive effects on BP reduction. DM, either alone or in combination with AM, improved aortic endothelial function indicated by ex vivo acetylcholine-induced relaxation. The combination of low-dose DM with AM gave most significant inhibition on aortic wall thickness in SHRs. Plasma total antioxidant status was significantly increased by all the therapies except for the combination of high-dose DM with high-dose AM. Serum nitrite and nitrate level was significantly reduced by AM but not by DM or the combination of DM with AM. Furthermore, in vitro treatment with DM reduced angiotensin II-induced reactive oxygen species and NADPH oxidase activation in human aortic endothelial cells. Conclusions/Significance Treatment of DM reduced BP and enhanced vascular protection probably by inhibiting vascular NADPH oxidase in aged hypertensive animals with or without AM treatment. It provides the potential rationale to a novel combination treatment with low-dose DM and AM in clinical hypertension. PMID:23049937

Interference studies with two hospital-grade and two home-grade glucose meters.

PubMed

Lyon, Martha E; Baskin, Leland B; Braakman, Sandy; Presti, Steven; Dubois, Jeffrey; Shirey, Terry

2009-10-01

Interference studies of four glucose meters (Nova Biomedical [Waltham, MA] StatStrip [hospital grade], Roche Diagnostics [Indianapolis, IN] Accu-Chek Aviva [home grade], Abbott Diabetes Care [Alameda, CA] Precision FreeStyle Freedom [home grade], and LifeScan [Milpitas, CA] SureStep Flexx [hospital grade]) were evaluated and compared to the clinical laboratory plasma hexokinase reference method (Roche Hitachi 912 chemistry analyzer). These meters were chosen to reflect the continuum of care from hospital to home grade meters commonly seen in North America. Within-run precision was determined using a freshly prepared whole blood sample spiked with concentrated glucose to give three glucose concentrations. Day-to-day precision was evaluated using aqueous control materials supplied by each vendor. Common interferences, including hematocrit, maltose, and ascorbate, were tested alone and in combination with one another on each of the four glucose testing devices at three blood glucose concentrations. Within-run precision for all glucose meters was <5% except for the FreeStyle (up to 7.6%). Between-day precision was <6% for all glucose meters. Ascorbate caused differences (percentage change from a sample without added interfering substances) of >5% with pyrroloquinolinequinone (PQQ)-glucose dehydrogenase-based technologies (Aviva and Freestyle) and the glucose oxidase-based Flexx meter. Maltose strongly affected the PQQ-glucose dehydrogenase-based meter systems. When combinations of interferences (ascorbate, maltose, and hematocrit mixtures) were tested, the extent of the interference was up to 193% (Aviva), 179% (FreeStyle), 25.1% (Flexx), and 5.9% (StatStrip). The interference was most pronounced at low glucose (3.9-4.4 mmol/L). All evaluated glucose meter systems demonstrated varying degrees of interference by hematocrit, ascorbate, and maltose mixtures. PQQ-glucose dehydrogenase-based technologies showed greater susceptibility than glucose oxidase-based systems. However, the modified glucose oxidase-based amperometric method (Nova StatStrip) was less affected in comparison with the glucose oxidase-based photometric method (LifeScan SureStep Flexx).

Reactive oxygen species derived from NAD(P)H oxidase play a role on ethanol-induced hypertension and endothelial dysfunction in rat resistance arteries.

PubMed

Simplicio, Janaina A; do Vale, Gabriel T; Gonzaga, Natália A; Leite, Letícia N; Hipólito, Ulisses V; Pereira, Camila A; Tostes, Rita C; Tirapelli, Carlos R

2017-02-01

Chronic ethanol consumption is a risk factor for cardiovascular diseases. We studied whether NAD(P)H oxidase-derived reactive oxygen species (ROS) play a role in ethanol-induced hypertension, vascular dysfunction, and protein expression in resistance arteries. Male Wistar rats were treated with ethanol (20 % v/v) for 6 weeks. Ethanol treatment increased blood pressure and decreased acetylcholine-induced relaxation in the rat mesenteric arterial bed (MAB). These responses were attenuated by apocynin (30 mg/kg/day; p.o. gavage). Ethanol consumption increased superoxide anion (O 2 - ) generation and decreased nitrate/nitrite (NO x ) concentration in the rat MAB and apocynin prevented these responses. Conversely, ethanol did not affect the concentration of hydrogen peroxide (H 2 O 2 ) and reduced glutathione (GSH) or the activity of superoxide dismutase (SOD) and catalase (CAT) in the rat MAB. Ethanol increased interleukin (IL)-10 levels in the rat MAB but did not affect the levels of tumor necrosis factor (TNF)-α, IL-6, or IL-1β. Ethanol increased the expression of Nox2 and the phosphorylation of SAPK/JNK, but reduced eNOS expression in the rat MAB. Apocynin prevented these responses. However, ethanol treatment did not affect the expression of Nox1, Nox4, p38MAPK, ERK1/2, or SAPK/JNK in the rat MAB. Ethanol increased plasma levels of TBARS, TNF-α, IL-6, IL-1β, and IL-10, whereas it decreased NO x levels. The major finding of our study is that NAD(P)H oxidase-derived ROS play a role on ethanol-induced hypertension and endothelial dysfunction in resistance arteries. Moreover, ethanol consumption affects the expression and phosphorylation of proteins that regulate vascular function and NAD(P)H oxidase-derived ROS play a role in such responses.

Role of amine structure on carbon dioxide adsorption from ultradilute gas streams such as ambient air.

PubMed

Didas, Stephanie A; Kulkarni, Ambarish R; Sholl, David S; Jones, Christopher W

2012-10-01

A fundamental study on the adsorption properties of primary, secondary, and tertiary amine materials is used to evaluate what amine type(s) are best suited for ultradilute CO(2) capture applications. A series of comparable materials comprised of primary, secondary, or tertiary amines ligated to a mesoporous silica support via a propyl linker are used to systematically assess the role of amine type. Both CO(2) and water adsorption isotherms are presented for these materials in the range relevant to CO(2) capture from ambient air and it is demonstrated that primary amines are the best candidates for CO(2) capture from air. Primary amines possess both the highest amine efficiency for CO(2) adsorption as well as enhanced water affinity compared to other amine types or the bare silica support. The results suggest that the rational design of amine adsorbents for the extraction of CO(2) from ambient air should focus on adsorbents rich in primary amines. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The direct reductive amination of electron-deficient amines with aldehydes: the unique reactivity of the Re2O7 catalyst.

PubMed

Das, Braja Gopal; Ghorai, Prasanta

2012-08-25

An unprecedented direct reductive amination of electron-deficient amines such as Cbz-, Boc-, EtOCO-, Fmoc-, Bz-, ArSO(2)-, Ar(2)PO-, etc. protected amines with aldehydes is achieved using the Re(2)O(7) catalyst and silanes as the hydride source. Excellent regioselective mono-alkylation and chemoselective reductive-amination were observed.

Concentration, size distribution and dry deposition of amines in atmospheric particles of urban Guangzhou, China

NASA Astrophysics Data System (ADS)

Liu, Fengxian; Bi, Xinhui; Zhang, Guohua; Peng, Long; Lian, Xiufeng; Lu, Huiying; Fu, Yuzhen; Wang, Xinming; Peng, Ping'an; Sheng, Guoying

2017-12-01

Size-segregated PM10 samples were collected in Guangzhou, China during autumn of 2014. Nine amines, including seven aliphatic amines and two heterocyclic amines, were detected using a gas chromatography-mass spectrometer after derivatization by benzenesulfonyl chloride. The total concentration of the nine amines (Ʃamines) was 79.6-140.9 ng m-3 in PM10. The most abundant species was methylamine (MA), which had a concentration of 29.2-70.1 ng m-3. MA, dimethylamine (DMA), diethylamine (DEA) and dibutylamine (DBA) were the predominant amines in the samples and accounted for approximately 80% of Ʃamines in each size segment. Two heterocyclic amines, pyrrolidine (PYR) and morpholine (MOR), were detected in all samples and had average concentrations of 1.14 ± 0.37 and 1.89 ± 0.64 ng m-3, respectively, in particles with aerodynamic diameters < 3 μm. More than 80% of Ʃamines were found in particles with diameters <1.5 μm, indicating that amines are mainly enriched in fine particles. All amines exhibited a bimodal distribution with a fine mode at 0.49-1.5 μm and a coarse mode at 7.2-10 μm. The maximum contributions of amines to particles (0.21%) and amines-N to water-soluble organic nitrogen (WSON) (3.1%) were found at the sizes < 0.49 μm. The maximum contribution of amines-C to water-soluble organic carbon (WSOC) was 1.6% over the size range of 0.95-1.5 μm. The molar ratio of Ʃamines to ammonium ranged from 0.0068 to 0.0107 in particles with diameters <1.5 μm, and the maximum ratio occurred in the smallest particles (diameter< 0.49 μm). The average dry deposition flux and velocity of Ʃamines in PM10 were 7.9 ± 1.6 μg m-2 d-1 and 0.084 ± 0.0021 cm s-1, respectively. The results of this study provide essential information on the contribution of amines to secondary organic aerosols and dry removal mechanisms in urban areas.

Utility of Hantzsch reaction for development of highly sensitive spectrofluorimetric method for determination of alfuzosin and terazosin in bulk, dosage forms and human plasma.

PubMed

Hammad, Mohamed A; Omar, Mahmoud A; Salman, Baher I

2017-09-01

A highly sensitive, cheap, simple and accurate spectrofluorimetric method has been developed and validated for the determination of alfuzosin hydrochloride and terazosin hydrochloride in their pharmaceutical dosage forms and in human plasma. The developed method is based on the reaction of the primary amine moiety in the studied drugs with acetylacetone and formaldehyde according to the Hantzsch reaction, producing yellow fluorescent products that can be measured spectrofluorimetrically at 480 nm after excitation at 415 nm. Different experimental parameters affecting the development and stability of the reaction products were carefully studied and optimized. The fluorescence-concentration plots of alfuzosin and terazosin were rectilinear over a concentration range of 70-900 ng ml -1 , with quantitation limits 27.1 and 32.2 ng ml -1 for alfuzosin and terazosin, respectively. The proposed method was validated according to ICH guidelines and successfully applied to the analysis of the investigated drugs in dosage forms, content uniformity test and spiked human plasma with high accuracy. Copyright © 2017 John Wiley & Sons, Ltd.

Biochemical diagnosis of phaeochromocytoma: two instructive case reports.

PubMed Central

Stewart, M F; Reed, P; Weinkove, C; Moriarty, K J; Ralston, A J

1993-01-01

The biochemical features of two patients with phaeochromocytomas illustrate the inadvisability of depending on a single group of analytes for the diagnosis. The first case presented as a surgical emergency with retroperitoneal haemorrhage. Biochemical diagnosis was difficult since total 24 hour urinary free catecholamine excretion was within normal limits in two out of three samples, and only marginally raised in the third with an atypical preponderance of adrenaline. Plasma catecholamine concentrations were also normal. But urinary excretion of the catecholamine metabolites, metadrenaline and 4-hydroxy-3-methoxy mandelic acid (HMMA), was consistently raised. In contrast, the second patient presenting with headache and labile hypertension showed normal metabolite excretion in the face of grossly increased free noradrenaline excretion and raised plasma noradrenaline concentrations. It is therefore recommend that, as well as urinary free catecholamines, one group of their main metabolites, the 3-methoxy amines (normetadrenaline and metadrenaline) or HMMA, should routinely be measured whenever a phaeochromocytoma is suspected. PMID:8463426

N-alkylated aminopyrazines for use as hydrophilic optical agents

NASA Astrophysics Data System (ADS)

Poreddy, Amruta R.; Asmelash, Bethel; Galen, Karen P.; Fitch, Richard M.; Shieh, Jeng-Jong; Wilcox, James M.; Schoenstein, Tasha M.; Wojdyla, Jolette K.; Gaston, Kimberly R.; Freskos, John N.; Neumann, William L.; Rajagopalan, Raghavan; Ahn, Hyo-Yang; Kostelc, James G.; Debreczeny, Martin P.; Belfield, Kevin D.; Dorshow, Richard B.

2009-02-01

Rapid assessment of glomerular filtration rate (GFR), which measures the amount of plasma filtered through the kidney within a given time, would greatly facilitate monitoring of renal function for patients at the bedside in the clinic. In our pursuit to develop exogenous fluorescent tracers for real-time monitoring of renal function by optical methods, N-alkylated aminopyrazine dyes and their hydrophilic conjugates based on poly (ethylene glycol) (PEG) were synthesized via reductive amination as the key step. Photophysical properties indicated a bathochromic shift on the order of 50 nm in both absorption and emission compared to naked aminopyrazines which could be very useful in enhancing both tissue penetration as well as easier detection methods. Structure-activity relationship (SAR) and pharmacokinetic (PK) studies, and the correlation of in vivo optical data with plasma PK for measurement of clearance (and hence GFR) are focus of the current investigation.

Effect of Functional Groups on Biodegradation and Pre-osteoblastic Cell Response on the Plasma-Polymerized Magnesium Surface

NASA Astrophysics Data System (ADS)

Ko, Yeong-Mu; Lee, Kang; Kim, Byung-Hoon

2013-01-01

Magnesium (Mg) is light, has biocompatibility, and has mechanical properties close to those of natural bone. However, pure Mg severely corrodes in a physiological environment, which may result in fracture prior to substantial tissue healing. In this study, the Mg surface was modified by depositing a thin polymeric film containing COOH, NH2, and OH groups through plasma polymerization of acrylic acid, allyl amine, and allyl alcohol in order to improve its anticorrosion and bioactive properties. The -COOH group had a significant effect on bonelike apatite formation compared with -NH2 and -OH. It was also concluded that a bonelike-apatite formed COOH/Mg surface was more effective for reducing biodegradation rate than the other surfaces. The results of in vitro cell test revealed significantly enhanced cell proliferation and differentiation on the COOH/Mg and NH2/Mg surfaces compared with other surfaces.

Is Xanthine oxidase activity in polycystic ovary syndrome associated with inflammatory and cardiovascular risk factors?

PubMed

Isık, Hatice; Aynıoglu, Oner; Tımur, Hakan; Sahbaz, Ahmet; Harma, Muge; Can, Murat; Guven, Berrak; Alptekin, Husnu; Kokturk, Furuzan

2016-08-01

The aim of this study is to examine women with polycystic ovary syndrome (PCOS) to determine the relationship between xanthine oxidase (XO) and oxidative stress, inflammatory status, and various clinical and biochemical parameters. In this cross-sectional study a total of 83 women including 45 PCOS patients and 38 healthy women were enrolled. We collected blood samples for XO and superoxide dismutase (SOD) activity, hormone levels, cholesterol values, and inflammatory markers. Body mass index (BMI) , waist-to-hip ratio (WHR), and blood pressure were assessed. Blood samples were taken for hormonal levels, cholesterol levels, fasting plasma glucose (FPG), fasting plasma insulin (FPI), homeostatic model assessment-insulin resistance (HOMA-IR) index, quantitative insulin sensitivity check index (QUICKI), C-reactive protein (CRP), white blood cell and neutrophil counts, XO and SOD activities. The basal hormone levels, triglyceride (TG) levels, TG/HDL-C (high density lipoprotein-cholesterol) ratios FPG, FPI and HOMA-IR levels were higher in PCOS patients compared to controls (p<0.05). Platelet and plateletcrit (PCT) values, CRP, and XO activity were significantly increased, however SOD activity was decreased in PCOS patients (p<0.001). XO activity was positively correlated with LH/FSH and TG/HDL ratios, CRP, PCT, FPG, FPI, and HOMA-IR, and negatively correlated with QUICKI levels. In conclusion, XO is a useful marker to assess oxidative stress in PCOS patients. Positive correlations between XO and inflammatory markers and cardiovascular disease risk factors suggest that XO plays an important role in the pathogenesis of PCOS and its metabolic complications. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Metabolic mechanisms of drug-nutrient interactions.

PubMed

Hathcock, J N

1985-01-01

Metabolic mechanisms of nutrition and drug interactions include 1) the effects of diet on drug metabolism and action and 2) the effects of drugs on nutritional processes. The type, amount, and timing of foods consumed influence drug dissolution, absorption, distribution, metabolism, and excretion. High-fat meals enhance the absorption of griseofulvin and some other drugs. Milk and other sources of calcium inhibit absorption of tetracycline. High-fat meals increase plasma concentrations of free fatty acids and thereby displace many drugs from binding sites on plasma albumin. High-protein diets increase the activity of the mixed-function oxidase system and enhance the metabolism of numerous drugs. High-electrolyte intakes increase excretion of lithium and also diminish the action of diuretic agents. Bile acid sequestrants and some laxatives decrease lipid digestion and absorption, as well as absorption of the fat-soluble vitamins. Numerous drugs, including tetracycline and cholestyramine, bind iron and decrease its absorption. Coumarins inhibit the function of vitamin K. Phenobarbital and other anticonvulsants are inducers of cytochrome P-450 and the mixed-function oxidase system. Long-term treatment with these inducers can cause excessive metabolism and deficiency of vitamin D. Prooxidant drugs such as chloroquine, drugs detoxified by conjugation with glutathione, and alcohol can deplete reduced glutathione with consequent effects on amino acid transport and the redox status of cells. Acid-forming foods acidify the urine and increase the loss of alkaline drugs such as the amphetamines. Base-forming drugs increase the loss of acidic drugs such as barbiturates. The range of metabolic interactions of drugs and nutrients includes the full scope of physiological processes to which drugs and nutrients are subject.

Mechanisms of the antihypertensive effects of Nigella sativa oil in L-NAME-induced hypertensive rats

PubMed Central

Jaarin, Kamsiah; Foong, Wai Dic; Yeoh, Min Hui; Kamarul, Zaman Yusoff Nik; Qodriyah, Haji Mohd Saad; Azman, Abdullah; Zuhair, Japar Sidik Fadhlullah; Juliana, Abdul Hamid; Kamisah, Yusof

2015-01-01

OBJECTIVES This study was conducted to determine whether the blood pressure-lowering effect of Nigella sativa might be mediated by its effects on nitric oxide, angiotensin-converting enzyme, heme oxygenase and oxidative stress markers. METHODS: Twenty-four adult male Sprague-Dawley rats were divided equally into 4 groups. One group served as the control (group 1), whereas the other three groups (groups 2-4) were administered L-NAME (25 mg/kg, intraperitoneally). Groups 3 and 4 were given oral nicardipine daily at a dose of 3 mg/kg and Nigella sativa oil at a dose of 2.5 mg/kg for 8 weeks, respectively, concomitantly with L-NAME administration. RESULTS Nigella sativa oil prevented the increase in systolic blood pressure in the L-NAME-treated rats. The blood pressure reduction was associated with a reduction in cardiac lipid peroxidation product, NADPH oxidase, angiotensin-converting enzyme activity and plasma nitric oxide, as well as with an increase in heme oxygenase-1 activity in the heart. The effects of Nigella sativa on blood pressure, lipid peroxidation product, nicotinamide adenine dinucleotide phosphate oxidase and angiotensin-converting enzyme were similar to those of nicardipine. In contrast, L-NAME had opposite effects on lipid peroxidation, angiotensin-converting enzyme and NO. CONCLUSION: The antihypertensive effect of Nigella sativa oil appears to be mediated by a reduction in cardiac oxidative stress and angiotensin-converting enzyme activity, an increase in cardiac heme oxygenase-1 activity and a prevention of plasma nitric oxide loss. Thus, Nigella sativa oil might be beneficial for controlling hypertension. PMID:26602523

Surface Charge, Electroosmotic Flow and DNA Extension in Chemically Modified Thermoplastic Nanoslits and Nanochannels

PubMed Central

Uba, Franklin I.; Pullagurla, Swathi R.; Sirasunthorn, Nichanun; Wu, Jiahao; Park, Sunggook; Chantiwas, Rattikan; Cho, Yoonkyoung; Shin, Heungjoo; Soper, Steven A.

2014-01-01

Thermoplastics have become attractive alternatives to glass/quartz for microfluidics, but the realization of thermoplastic nanofluidic devices has been slow in spite of the rather simple fabrication techniques that can be used to produce these devices. This slow transition has in part been attributed to insufficient understanding of surface charge effects on the transport properties of single molecules through thermoplastic nanochannels. We report the surface modification of thermoplastic nanochannels and an assessment of the associated surface charge density, zeta potential and electroosmotic flow (EOF). Mixed-scale fluidic networks were fabricated in poly(methylmethacrylate), PMMA. Oxygen plasma was used to generate surface-confined carboxylic acids with devices assembled using low temperature fusion bonding. Amination of the carboxylated surfaces using ethylenediamine (EDA) was accomplished via EDC coupling. XPS and ATR-FTIR revealed the presence of carboxyl and amine groups on the appropriately prepared surfaces. A modified conductance equation for nanochannels was developed to determine their surface conductance and was found to be in good agreement with our experimental results. The measured surface charge density and zeta potential of these devices were lower than glass nanofluidic devices and dependent on the surface modification adopted, as well as the size of the channel. This property, coupled to an apparent increase in fluid viscosity due to nanoconfinement, contributed to the suppression of the EOF in PMMA nanofluidic devices by an order of magnitude compared to the micro-scale devices. Carboxylated PMMA nanochannels were efficient for the transport and elongation of λ-DNA while these same DNA molecules were unable to translocate through aminated nanochannels. PMID:25369728

Neoglycolipid analogues of ganglioside G sub M1 as functional receptors of cholera toxin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pacuszka, T.; Bradley, R.M.; Fishman, P.H.

1991-03-12

The authors synthesized several lipid analogues of ganglioside G{sub M1} by attaching its oligosaccharide moiety (G{sub M1}OS) to aminophospholipids, aliphatic amines, and cholesteryl hemisuccinate. They incubated G{sub M1}-deficient rat glioma C6 cells with each of the derivatives as well as native G{sub M1} and assayed the cells for their ability to bind and respond to cholera toxin. On the basis of the observed increase in binding of {sup 125}I-labeled cholera toxin, it was apparent that the cells took up and initially incorporated most of the derivatives into the plasma membrane. In the case of the aliphatic amine derivatives, the abilitymore » to generate new toxin binding sites was dependent on chain length; whereas the C{sub 10} derivative was ineffective, C{sub 12} and higher analogues were effective. Increased binding was dependent on both the concentration of the neoglycolipid in the medium and the time of exposure. Cells pretreated with the various derivatives accumulated cyclic AMP in response to cholera toxin, but there were differences in their effectiveness. The cholesterol and long-chain aliphatic amine derivatives were more effective than native G{sub M1}, whereas the phospholipid derivatives were less effective. The distance between G{sub M1}OS and the phospholipid also appeared to influence its functional activity. The results indicate that although G{sub M1}OS provides the recognition site for the binding of cholera toxin, the nature of the lipid moiety plays an important role in the action of the toxin.« less

Impacts of crystal metal on secondary aliphatic amine aerosol formation during dust storm episodes in Beijing

NASA Astrophysics Data System (ADS)

Liu, Qingyang; Bei, Yiling

2016-03-01

Trimethylamine (TMA) enters the atmosphere from a variety of sources and is a ubiquitous atmospheric organic base. The atmospheric reaction mechanism of TMA with key atmospheric oxidants is important to predict its distribution and environmental behavior in the particle phase. While previous studies have extensively focused on the production of particle amine salts (i.e. trimethylamine-N-oxide (TMAO)) using chamber experiments, the atmospheric behavior of TMAO in the environment is still poorly understood. Ambient fine particulate matter (PM2.5) was collected at two sampling sites in Beijing from March 10 to May 10, 2012. We analyzed the samples for water-soluble ions, crystal metals, TMA, and TMAO. Water-soluble ions (e.g. SO42-, NO3- , NH4+), TMA, and TMAO were measured using ion chromatography, while crystal metal (e.g. Al, Fe, Mn) in PM2.5 was quantified by inductively coupled plasma mass spectrometry (ICP-MS). Two dust storms (DS) occurred during the sampling period on March 28 and April 28. Mineral dust impacted PM2.5 mass and composition greatly during dust storm days, as it contributed approximately 1.2-4.0 times greater on dust storm days versus non-dust storm days. We found TMAO concentrations were highly associated with aluminum in PM2.5. Further, we applied the density functional theory (DFT) method to confirm that aluminum plays a catalytic effect in the reaction of TMA with ozone (O3). Our work improves understanding of the effect of crystal metals on secondary aliphatic amine aerosol formation in the atmosphere.

Surface charge, electroosmotic flow and DNA extension in chemically modified thermoplastic nanoslits and nanochannels.

PubMed

Uba, Franklin I; Pullagurla, Swathi R; Sirasunthorn, Nichanun; Wu, Jiahao; Park, Sunggook; Chantiwas, Rattikan; Cho, Yoon-Kyoung; Shin, Heungjoo; Soper, Steven A

2015-01-07

Thermoplastics have become attractive alternatives to glass/quartz for microfluidics, but the realization of thermoplastic nanofluidic devices has been slow in spite of the rather simple fabrication techniques that can be used to produce these devices. This slow transition has in part been attributed to insufficient understanding of surface charge effects on the transport properties of single molecules through thermoplastic nanochannels. We report the surface modification of thermoplastic nanochannels and an assessment of the associated surface charge density, zeta potential and electroosmotic flow (EOF). Mixed-scale fluidic networks were fabricated in poly(methylmethacrylate), PMMA. Oxygen plasma was used to generate surface-confined carboxylic acids with devices assembled using low temperature fusion bonding. Amination of the carboxylated surfaces using ethylenediamine (EDA) was accomplished via EDC coupling. XPS and ATR-FTIR revealed the presence of carboxyl and amine groups on the appropriately prepared surfaces. A modified conductance equation for nanochannels was developed to determine their surface conductance and was found to be in good agreement with our experimental results. The measured surface charge density and zeta potential of these devices were lower than glass nanofluidic devices and dependent on the surface modification adopted, as well as the size of the channel. This property, coupled to an apparent increase in fluid viscosity due to nanoconfinement, contributed to the suppression of the EOF in PMMA nanofluidic devices by an order of magnitude compared to the micro-scale devices. Carboxylated PMMA nanochannels were efficient for the transport and elongation of λ-DNA while these same DNA molecules were unable to translocate through aminated nanochannels.

Quantitative plasma biomarker analysis in HDI exposure assessment.

PubMed

Flack, Sheila L; Fent, Kenneth W; Trelles Gaines, Linda G; Thomasen, Jennifer M; Whittaker, Steve; Ball, Louise M; Nylander-French, Leena A

2010-01-01

Quantification of amines in biological samples is important for evaluating occupational exposure to diisocyanates. In this study, we describe the quantification of 1,6-hexamethylene diamine (HDA) levels in hydrolyzed plasma of 46 spray painters applying 1,6-hexamethylene diisocyanate (HDI)-containing paint in vehicle repair shops collected during repeated visits to their workplace and their relationship with dermal and inhalation exposure to HDI monomer. HDA was detected in 76% of plasma samples, as heptafluorobutyryl derivatives, and the range of HDA concentrations was < or =0.02-0.92 microg l(-1). After log-transformation of the data, the correlation between plasma HDA levels and HDI inhalation exposure measured on the same workday was low (N = 108, r = 0.22, P = 0.026) compared with the correlation between plasma HDA levels and inhalation exposure occurring approximately 20 to 60 days before blood collection (N = 29, r = 0.57, P = 0.0014). The correlation between plasma HDA levels and HDI dermal exposure measured on the same workday, although statistically significant, was low (N = 108, r = 0.22, P = 0.040) while the correlation between HDA and dermal exposure occurring approximately 20 to 60 days before blood collection was slightly improved (N = 29, r = 0.36, P = 0.053). We evaluated various workplace factors and controls (i.e. location, personal protective equipment use and paint booth type) as modifiers of plasma HDA levels. Workers using a downdraft-ventilated booth had significantly lower plasma HDA levels relative to semi-downdraft and crossdraft booth types (P = 0.0108); this trend was comparable to HDI inhalation and dermal exposure levels stratified by booth type. These findings indicate that HDA concentration in hydrolyzed plasma may be used as a biomarker of cumulative inhalation and dermal exposure to HDI and for investigating the effectiveness of exposure controls in the workplace.

Ruthenium-catalyzed regioselective allylic amination of 2,3,3-trifluoroallylic carbonates.

PubMed

Isobe, Shin-Ichi; Terasaki, Shou; Hanakawa, Taisyun; Mizuno, Shota; Kawatsura, Motoi

2017-04-05

We demonstrated the ruthenium-catalyzed allylic amination of 2,3,3-trifluoroallylic carbonates with several types of amines. The reactions proceeded with several types of amines, and succeeded in obtaining polyfluorinated terminal alkenes possessing branched allylic amines as a single regioisomer.

A one-pot parallel reductive amination of aldehydes with heteroaromatic amines.

PubMed

Bogolubsky, Andrey V; Moroz, Yurii S; Mykhailiuk, Pavel K; Panov, Dmitriy M; Pipko, Sergey E; Konovets, Anzhelika I; Tolmachev, Andrey

2014-08-11

A parallel reductive amination of heteroaromatic amines has been performed using a combination of ZnCl2-TMSOAc (activating agents) and NaBH(OAc)3 (reducing agent). A library of diverse secondary amines was easily prepared on a 50-300 mg scale.

Amine dehydrogenases: efficient biocatalysts for the reductive amination of carbonyl compounds

PubMed Central

Mutti, Francesco G.

2017-01-01

Amines constitute the major targets for the production of a plethora of chemical compounds that have applications in the pharmaceutical, agrochemical and bulk chemical industries. However, the asymmetric synthesis of α-chiral amines with elevated catalytic efficiency and atom economy is still a very challenging synthetic problem. Here, we investigated the biocatalytic reductive amination of carbonyl compounds employing a rising class of enzymes for amine synthesis: amine dehydrogenases (AmDHs). The three AmDHs from this study – operating in tandem with a formate dehydrogenase from Candida boidinii (Cb-FDH) for the recycling of the nicotinamide coenzyme – performed the efficient amination of a range of diverse aromatic and aliphatic ketones and aldehydes with up to quantitative conversion and elevated turnover numbers (TONs). Moreover, the reductive amination of prochiral ketones proceeded with perfect stereoselectivity, always affording the (R)-configured amines with more than 99% enantiomeric excess. The most suitable amine dehydrogenase, the optimised catalyst loading and the required reaction time were determined for each substrate. The biocatalytic reductive amination with this dual-enzyme system (AmDH–Cb-FDH) possesses elevated atom efficiency as it utilizes the ammonium formate buffer as the source of both nitrogen and reducing equivalents. Inorganic carbonate is the sole by-product. PMID:28663713

A Catalyst-Free Amination of Functional Organolithium Reagents by Flow Chemistry.

PubMed

Kim, Heejin; Yonekura, Yuya; Yoshida, Jun-Ichi

2018-04-03

Reported is the electrophilic amination of functional organolithium intermediates with well-designed aminating reagents under mild reaction conditions using flow microreactors. The aminating reagents were optimized to achieve efficient C-N bond formation without using any catalyst. The electrophilic amination reactions of functionalized aryllithiums were successfully conducted under mild reaction conditions, within 1 minute, by using flow microreactors. The aminating reagent was also prepared by the flow method. Based on stopped-flow NMR analysis, the reaction time for the preparation of the aminating reagent was quickly optimized without the necessity of work-up. Integrated one-flow synthesis consisting of the generation of an aryllithium, the preparation of an aminating reagent, and their combined reaction was successfully achieved to give the desired amine within 5 minutes of total reaction time. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Thermometric titrations of amines with nitrosyl perchlorate in acetonitrile solvent.

PubMed

Gündüz, T; Kiliç, E; Cakirer, O

1996-05-01

Thirteen aliphatic and four aromatic amines, namely diethylamine, triethylamine, n-propylamine, di-n-propylamine, tri-n-butylamine, isopropylamine, di-isopropylamine, n-butylamine, di-n-butylamine, tri-n-butylamine, isobutylamine, sec-butylamine, tert-butylamine, aniline, N,N-dimethylaniline, 2-nitroaniline and 4-nitroaniline were titrated thermometrically with nitrosyl perchlorate in acetonitrile solvent. All the aliphatic amines gave very well-shaped thermometric titration curves. The calculated recovery values of the amines were very good. In comparison, the aromatic amines, aniline and N,N-dimethylaniline gave rather well-shaped titration curves, but the recovery values were fairly low. 2-Nitro- and 4-nitro anilines gave no thermometric response at all. The heats of reaction of the amines with nitrosyl perchlorate are rather high. However, the average heat of reaction of the aromatic amines is approximately two-thirds that of the average heat of the aliphatic amines. To support this method all the amines were also titrated potentiometrically and very similar results to those obtained with the thermometric method are seen. The nitrosyl ion is a Lewis acid, strong enough to titrate quantitatively aliphatic amines in acetonitrile solvent, but not strong enough to titrate aromatic amines at the required level in the same solvent.

The human plasma-metabolome: Reference values in 800 French healthy volunteers; impact of cholesterol, gender and age

PubMed Central

Al-Salameh, Abdallah; Croixmarie, Vincent; Masson, Perrine; Corruble, Emmanuelle; Fève, Bruno; Colle, Romain; Ripoll, Laurent; Walther, Bernard; Boursier-Neyret, Claire; Werner, Erwan; Becquemont, Laurent; Chanson, Philippe

2017-01-01

Metabolomic approaches are increasingly used to identify new disease biomarkers, yet normal values of many plasma metabolites remain poorly defined. The aim of this study was to define the “normal” metabolome in healthy volunteers. We included 800 French volunteers aged between 18 and 86, equally distributed according to sex, free of any medication and considered healthy on the basis of their medical history, clinical examination and standard laboratory tests. We quantified 185 plasma metabolites, including amino acids, biogenic amines, acylcarnitines, phosphatidylcholines, sphingomyelins and hexose, using tandem mass spectrometry with the Biocrates AbsoluteIDQ p180 kit. Principal components analysis was applied to identify the main factors responsible for metabolome variability and orthogonal projection to latent structures analysis was employed to confirm the observed patterns and identify pattern-related metabolites. We established a plasma metabolite reference dataset for 144/185 metabolites. Total blood cholesterol, gender and age were identified as the principal factors explaining metabolome variability. High total blood cholesterol levels were associated with higher plasma sphingomyelins and phosphatidylcholines concentrations. Compared to women, men had higher concentrations of creatinine, branched-chain amino acids and lysophosphatidylcholines, and lower concentrations of sphingomyelins and phosphatidylcholines. Elderly healthy subjects had higher sphingomyelins and phosphatidylcholines plasma levels than young subjects. We established reference human metabolome values in a large and well-defined population of French healthy volunteers. This study provides an essential baseline for defining the “normal” metabolome and its main sources of variation. PMID:28278231

The human plasma-metabolome: Reference values in 800 French healthy volunteers; impact of cholesterol, gender and age.

PubMed

Trabado, Séverine; Al-Salameh, Abdallah; Croixmarie, Vincent; Masson, Perrine; Corruble, Emmanuelle; Fève, Bruno; Colle, Romain; Ripoll, Laurent; Walther, Bernard; Boursier-Neyret, Claire; Werner, Erwan; Becquemont, Laurent; Chanson, Philippe

2017-01-01

Metabolomic approaches are increasingly used to identify new disease biomarkers, yet normal values of many plasma metabolites remain poorly defined. The aim of this study was to define the "normal" metabolome in healthy volunteers. We included 800 French volunteers aged between 18 and 86, equally distributed according to sex, free of any medication and considered healthy on the basis of their medical history, clinical examination and standard laboratory tests. We quantified 185 plasma metabolites, including amino acids, biogenic amines, acylcarnitines, phosphatidylcholines, sphingomyelins and hexose, using tandem mass spectrometry with the Biocrates AbsoluteIDQ p180 kit. Principal components analysis was applied to identify the main factors responsible for metabolome variability and orthogonal projection to latent structures analysis was employed to confirm the observed patterns and identify pattern-related metabolites. We established a plasma metabolite reference dataset for 144/185 metabolites. Total blood cholesterol, gender and age were identified as the principal factors explaining metabolome variability. High total blood cholesterol levels were associated with higher plasma sphingomyelins and phosphatidylcholines concentrations. Compared to women, men had higher concentrations of creatinine, branched-chain amino acids and lysophosphatidylcholines, and lower concentrations of sphingomyelins and phosphatidylcholines. Elderly healthy subjects had higher sphingomyelins and phosphatidylcholines plasma levels than young subjects. We established reference human metabolome values in a large and well-defined population of French healthy volunteers. This study provides an essential baseline for defining the "normal" metabolome and its main sources of variation.

Partial hydrophilic modification of biaxially oriented polypropylene film by an atmospheric pressure plasma jet with the allylamine monomer

NASA Astrophysics Data System (ADS)

Chen, W. X.; Yu, J. S.; Hu, W.; Chen, G. L.

2016-11-01

In this paper, the partial modification of the biaxially oriented polypropylene (BOPP) film for potential biological and packaging applications was achieved via hydrophilic modification using atmospheric pressure plasma jet (APPJ). In the APPJ system, the allylamine (ALA) monomer was polymerized on the BOPP surface by either the Ar/O2 or the He/O2 plasma. The results showed that plasmatic modification created many micro/nano sized holes on the BOPP film, which increased the surface roughness dramatically and the increased roughness enhanced the combining intensity between the BOPP film and the ALA polymer. However, such a plasmatic modification increased the water vapor permeability. The FTIR and XPS characterizations showed that the amine groups were grafted onto the BOPP film, and the contact angle of the BOPP film decreases from 98.5° to 8°. Compared with the BOPP films treated by the Ar or He plasma, the barrier property of the modified BOPP film increased significantly when the ALA polymer was incorporated. The bio-affinity/toxicity of ALA polymer was illustrated by the attachment of the cultured SMMC-7721 hepatoma cells on the modified BOPP film. The significant enhancement in the cell density indicated that modified BOPP film was highly bio-compatible and non-toxic, especially treated with the Ar/O2/ALA plasma.

Polymer-silica hybrids for separation of CO2 and catalysis of organic reactions

NASA Astrophysics Data System (ADS)

Silva Mojica, Ernesto

Porous materials comprising polymeric and inorganic segments have attracted interest from the scientific community due to their unique properties and functionalities. The physical and chemical characteristics of these materials can be effectively exploited for adsorption applications. This dissertation covers the experimental techniques for fabrication of poly(vinyl alcohol) (PVA) and silica (SiO2) porous supports, and their functionalization with polyamines for developing adsorbents with potential applications in separation of CO2 and catalysis of organic reactions. The supports were synthesized by processes involving (i) covalent cross-linking of PVA, (ii) hydrolysis and poly-condensation of silica precursors (i,e,. sol-gel synthesis), and formation of porous structures via (iii) direct templating and (iv) phase inversion techniques. Their physical structure was controlled by the proper combination of the preparation procedures, which resulted in micro-structured porous materials in the form of micro-particles, membranes, and pellets. Their adsorption characteristics were tailored by functionalization with polyethyleneimine (PEI), and their physicochemical properties were characterized by vibrational spectroscopy (FTIR, UV-vis), microscopy (SEM), calorimetry (TGA, DSC), and adsorption techniques (BET, step-switch adsorption). Spectroscopic investigations of the interfacial cross-linking reactions of PEI and PVA with glutaraldehyde (GA) revealed that PEI catalyzes the cross-linking reactions of PVA in absence of external acid catalysts. In-situ IR spectroscopy coupled with a focal plane array (FPA) image detector allowed the characterization of a gradient interface on a PEI/PVA composite membrane and the investigation of the cross-linking reactions as a function of time and position. The results served as a basis to postulate possible intermediates, and propose the reaction mechanisms. The formulation of amine-functionalized CO2 capture sorbents was based on the spectroscopic investigation of the interactions of CO 2 with amine molecules under simulated CO2 capture conditions. Industrial CO2 capture processes involve fluidization and require degradation-resistant sorbents in the form of pellets. Agglomeration of silica-based CO2 capture sorbents involved the formulation of a polymer binder solution and the design of a scalable pelletization process. The characterization of these pellets revealed the formation of a CO 2-permeable polymer-silica network, which is resistant to attrition, and exhibits similar CO2 capture and degradation performance as the non-pelletized sorbents. The performance of these sorbents and pellets was tested in lab-scale and bench-scale adsorption units, using in-house fabricated fixed-bed and fluidized-bed reactors. A compartmental modeling technique was used to simulate the CO2 adsorption process and to elucidate the kinetic and thermodynamic parameters that impact the commercial viability of emerging CO2 capture technologies. The fundamental concepts and experimental techniques developed for the preparation of CO2 capture sorbents served as a basis for fabricating amine-functionalized polymer-silica hybrids for applications in catalysis of organic reactions. (i) Basic catalysts for carbon-carbon addition reactions were prepared by immobilization of amine molecules on silica supports. The activity of these catalysts and the mechanisms of base-catalyzed organic condensation reactions were investigated by an in-situ FTIR micro-scale reactor. (ii) Particle-loaded PVA composite membranes were selected for immobilization of glucose oxidase (GOx). GOx was immobilized by adsorption at pH values between 3.5 and 7.1. The results showed that adsorption was primarily achieved via hydrophobic interactions, and that PVA membranes loaded with amine-functionalized particles could help retain the activity of immobilized GOx by providing a proper hydrophilic/hydrophobic balance to the immobilized enzymes micro-environment.

Identification of amines in wintertime ambient particulate material using high resolution aerosol mass spectrometry

NASA Astrophysics Data System (ADS)

Bottenus, Courtney L. H.; Massoli, Paola; Sueper, Donna; Canagaratna, Manjula R.; VanderSchelden, Graham; Jobson, B. Thomas; VanReken, Timothy M.

2018-05-01

Significant amounts of amines were detected in fine particulate matter (PM) during ambient wintertime conditions in Yakima, WA, using a high resolution time-of-flight aerosol mass spectrometer (HR-ToF-AMS). Positive matrix factorization (PMF) of the organic aerosol (OA) signal resulted in a six-factor solution that included two previously unreported amine OA factors. The contributions of the amine factors were strongly episodic, but the concentration of the combined amine factors was as high as 10-15 μg m-3 (2-min average) during those episodes. In one occasion, the Amine-II component was 45% of total OA signal. The Amine-I factor was dominated by spectral peaks at m/z 86 (C5H12N+) and m/z 100 (C6H14N+), while the Amine-II factor was dominated by spectral peaks at m/z 58 (C3H8N+ and C2H6N2+) and m/z 72 (C4H10N+ and C3H8N2+). The ions dominating each amine factor showed distinct time traces, suggesting different sources or formation processes. Investigation into the chemistry of the amine factors suggests a correlation with inorganic anions for Amine-I, but no evidence that the Amine-II was being neutralized by the same inorganic ions. We also excluded the presence of organonitrates (ON) in the OA. The presence of C2H4O2+ at m/z 60 (a levoglucosan fragment) in the Amine-I spectrum suggests some influence of biomass burning emissions (more specifically residential wood combustion) in this PMF factor, but wind direction suggested that the most likely sources of these amines were agricultural activities and feedlots to the S-SW of the site.

A Simple Secondary Amine Synthesis: Reductive Amination Using Sodium Triacetoxyborohydride

NASA Astrophysics Data System (ADS)

Carlson, Merle W.; Ciszewski, James T.; Bhatti, Micah M.; Swanson, Wesley F.; Wilson, Anne M.

2000-02-01

We present a reductive amination experiment for a second-semester organic chemistry class. It utilizes an imine intermediate and sodium triacetoxyborohydride, a mild reducing agent. The progress of the reaction is followed by TLC as the starting materials (the aldehyde and primary amine), the imine intermediate, and the secondary amine product are visible under ultraviolet light. This experiment provides an introduction to the observation of intermediates, the synthesis of amines, and the concept of mild reducing agents.

Role of Amine Functionality for CO2 Chemisorption on Silica.

PubMed

Hahn, Maximilian W; Jelic, Jelena; Berger, Edith; Reuter, Karsten; Jentys, Andreas; Lercher, Johannes A

2016-03-03

The mechanism of CO2 adsorption on primary, secondary, and bibasic aminosilanes synthetically functionalized in porous SiO2 was qualitatively and quantitatively investigated by a combination of IR spectroscopy, thermogravimetry, and quantum mechanical modeling. The mode of CO2 adsorption depends particularly on the nature of the amine group and the spacing between the aminosilanes. Primary amines bonded CO2 preferentially through the formation of intermolecular ammonium carbamates, whereas CO2 was predominantly stabilized as carbamic acid, when interacting with secondary amines. Ammonium carbamate formation requires the transfer of the carbamic acid proton to a second primary amine group to form the ammonium ion and hence two (primary) amine groups are required to bind one CO2 molecule. The higher base strength of secondary amines enables the stabilization of carbamic acid, which is thereby hindered to interact further with nearby amine functions, because their association with Si-OH groups (either protonation or hydrogen bonding) does not allow further stabilization of carbamic acid as carbamate. Steric hindrance of the formation of intermolecular ammonium carbamates leads to higher uptake capacities for secondary amines functionalized in porous SiO2 at higher amine densities. In aminosilanes possessing a primary and a secondary amine group, the secondary amine group tends to be protonated by Si-OH groups and therefore does not substantially interact with CO2.

Rapid enzymatic analysis of plasma for tyrosine.

PubMed

Shimizu, H; Taniguchi, K; Sugiyama, M; Kanno, T

1990-01-01

In this rapid, simple, and convenient enzymatic method for measurement of tyrosine in plasma, tyrosine is converted to tyramine by action of tyrosine decarboxylase (EC 4.1.1.25) and the tyramine produced is oxidized to p-hydroxybenzyl aldehyde and hydrogen peroxide by action of tyramine oxidase (EC 1.4.3.9). The hydrogen peroxide is reacted with 4-aminoantipyrine and N-ethyl-N-(2-hydroxy-3-sulfopropyl)-m-toluidine in the presence of peroxidase (EC 1.11.1.7) to obtain quinoneimine dye, the absorbance of which is measured at 570 nm. Thus tyrosine is measured in the visible range. The CV was 4.6% or less, and the measurement was unaffected by other amino acids, except for phenylalanine. The values obtained (y) correlated well with those obtained with an amino acid analyzer (x): y = 0.902x + 3.92 mumol/L (Syx = 12.3; r = 0.985; n = 54).

Hemolysis in a patient with alkaptonuria and chronic kidney failure.

PubMed

Heng, Anne-Elisabeth; Courbebaisse, Marie; Kemeny, Jean Louis; Matesan, Raluca; Bonniol, Claude; Deteix, Patrice; Souweine, Bertrand

2010-07-01

In alkaptonuria, the absence of homogentisic acid oxidase results in the accumulation of homogentisic acid (HGA) in the body. Fatal disease cases are infrequent, and death often results from kidney or cardiac complications. We report a 24-year-old alkaptonuric man with severe decreased kidney function who developed fatal metabolic acidosis and intravascular hemolysis. Hemolysis may have been caused by rapid and extensive accumulation of HGA and subsequent accumulation of plasma soluble melanins. Toxic effects of plasma soluble melanins, their intermediates, and reactive oxygen side products are increased when antioxidant mechanisms are overwhelmed. A decrease in serum antioxidative activity has been reported in patients with chronic decreased kidney function. However, despite administration of large doses of an antioxidant agent and ascorbic acid and intensive kidney support, hemolysis and acidosis could not be brought under control and hemolysis led to the death of the patient.

Azobisisobutyronitrile initiated aerobic oxidative transformation of amines: coupling of primary amines and cyanation of tertiary amines.

PubMed

Liu, Lianghui; Wang, Zikuan; Fu, Xuefeng; Yan, Chun-Hua

2012-11-16

In the presence of a catalytic amount of radical initiator AIBN, primary amines are oxidatively coupled to imines and tertiary amines are cyanated to α-aminonitriles. These "metal-free" aerobic oxidative coupling reactions may find applications in a wide range of "green" oxidation chemistry.

Non-amine-based dopamine transporter (reuptake) inhibitors retain properties of amine-based progenitors.

PubMed

Madras, Bertha K; Fahey, Michele A; Miller, Gregory M; De La Garza, Richard; Goulet, Martin; Spealman, Roger D; Meltzer, Peter C; George, Susan R; O'Dowd, Brian F; Bonab, Ali A; Livni, Eli; Fischman, Alan J

2003-10-31

Without exception, therapeutic and addictive drugs that produce their primary effects by blocking monoamine transporters in brain contain an amine nitrogen in their structure. This fundamental canon of drug design was based on a prevailing premise that an amine nitrogen is required to mimic the structures of monoamine neurotransmitters and other natural products. Non-amines, a novel class of compounds that contain no amine nitrogen, block monoamine transporters in the nM range and display markedly high selectivity for monoamine transporters, but not for receptors. Non-amines retain the spectrum of biochemical and pharmacological properties characteristic of amine-bearing counterparts. These novel drugs compel a revision of current concepts of drug-monoamine transporter complex formation and open avenues for discovery of a new generation of therapeutic drugs.

Readily Available Chiral Benzimidazoles-Derived Guanidines as Organocatalysts in the Asymmetric α-Amination of 1,3-Dicarbonyl Compounds.

PubMed

Benavent, Llorenç; Puccetti, Francesco; Baeza, Alejandro; Gómez-Martínez, Melania

2017-08-11

The synthesis and the evaluation as organocatalysts of new chiral guanidines derived from benzimidazoles in the enantioselective α-amination of 1,3-dicarbonyl compounds using di- t -butylazodicarboxylate as aminating agent is herein disclosed. The catalysts are readily synthesized through the reaction of 2-chlorobezimidazole and a chiral amine in moderate-to-good yields. Among all of them, those derived from ( R )-1-phenylethan-1-amine ( 1 ) and ( S )-1-(2-naphthyl)ethan-1-amine ( 3 ) turned out to be the most efficient for such asymmetric transformation, rendering good-to-high yields and moderate-to-good enantioselectivities for the amination products.

Sequential Metabolism of Secondary Alkyl Amines to Metabolic-Intermediate Complexes: Opposing Roles for the Secondary Hydroxylamine and Primary Amine Metabolites of Desipramine, (S)-Fluoxetine, and N-Desmethyldiltiazem

PubMed Central

Hanson, Kelsey L.; VandenBrink, Brooke M.; Babu, Kantipudi N.; Allen, Kyle E.; Nelson, Wendel L.

2010-01-01

Three secondary amines desipramine (DES), (S)-fluoxetine [(S)-FLX], and N-desmethyldiltiazem (MA) undergo N-hydroxylation to the corresponding secondary hydroxylamines [N-hydroxydesipramine, (S)-N-hydroxyfluoxetine, and N-hydroxy-N-desmethyldiltiazem] by cytochromes P450 2C11, 2C19, and 3A4, respectively. The expected primary amine products, N-desmethyldesipramine, (S)-norfluoxetine, and N,N-didesmethyldiltiazem, are also observed. The formation of metabolic-intermediate (MI) complexes from these substrates and metabolites was examined. In each example, the initial rates of MI complex accumulation followed the order secondary hydroxylamine > secondary amine ≫ primary amine, suggesting that the primary amine metabolites do not contribute to formation of MI complexes from these secondary amines. Furthermore, the primary amine metabolites, which accumulate in incubations of the secondary amines, inhibit MI complex formation. Mass balance studies provided estimates of the product ratios of N-dealkylation to N-hydroxylation. The ratios were 2.9 (DES-CYP2C11), 3.6 [(S)-FLX-CYP2C19], and 0.8 (MA-CYP3A4), indicating that secondary hydroxylamines are significant metabolites of the P450-mediated metabolism of secondary alkyl amines. Parallel studies with N-methyl-d3-desipramine and CYP2C11 demonstrated significant isotopically sensitive switching from N-demethylation to N-hydroxylation. These findings demonstrate that the major pathway to MI complex formation from these secondary amines arises from N-hydroxylation rather than N-dealkylation and that the primary amines are significant competitive inhibitors of MI complex formation. PMID:20200233

Sequential metabolism of secondary alkyl amines to metabolic-intermediate complexes: opposing roles for the secondary hydroxylamine and primary amine metabolites of desipramine, (s)-fluoxetine, and N-desmethyldiltiazem.

PubMed

Hanson, Kelsey L; VandenBrink, Brooke M; Babu, Kantipudi N; Allen, Kyle E; Nelson, Wendel L; Kunze, Kent L

2010-06-01

Three secondary amines desipramine (DES), (S)-fluoxetine [(S)-FLX], and N-desmethyldiltiazem (MA) undergo N-hydroxylation to the corresponding secondary hydroxylamines [N-hydroxydesipramine, (S)-N-hydroxyfluoxetine, and N-hydroxy-N-desmethyldiltiazem] by cytochromes P450 2C11, 2C19, and 3A4, respectively. The expected primary amine products, N-desmethyldesipramine, (S)-norfluoxetine, and N,N-didesmethyldiltiazem, are also observed. The formation of metabolic-intermediate (MI) complexes from these substrates and metabolites was examined. In each example, the initial rates of MI complex accumulation followed the order secondary hydroxylamine > secondary amine > primary amine, suggesting that the primary amine metabolites do not contribute to formation of MI complexes from these secondary amines. Furthermore, the primary amine metabolites, which accumulate in incubations of the secondary amines, inhibit MI complex formation. Mass balance studies provided estimates of the product ratios of N-dealkylation to N-hydroxylation. The ratios were 2.9 (DES-CYP2C11), 3.6 [(S)-FLX-CYP2C19], and 0.8 (MA-CYP3A4), indicating that secondary hydroxylamines are significant metabolites of the P450-mediated metabolism of secondary alkyl amines. Parallel studies with N-methyl-d(3)-desipramine and CYP2C11 demonstrated significant isotopically sensitive switching from N-demethylation to N-hydroxylation. These findings demonstrate that the major pathway to MI complex formation from these secondary amines arises from N-hydroxylation rather than N-dealkylation and that the primary amines are significant competitive inhibitors of MI complex formation.

[Plasma levels of mediator amino acids in patients with Parkinson disease].

PubMed

Vitreshchak, T V; Poleshchuk, V V; Piradov, M A

2004-01-01

Content of neurotransmitter amino acids before and after treatment with He-Ne-laser was measured in blood of two groups of the Parkinson's disease patients distinguished by low (first group) and high (second group) activity of monoamine oxidase B and Cu/Zn-superoxide dismutase. An increase in taurine level at the early stage of the disease (first group of patients) suggests that taurine may be a marker of compensatory abilities of the organism. The violation of the glutamate/taurine balance at the later stages of the disease and its normalization following the laserotherapy accompanied improvement of neurological symptoms.

Selective cytotoxicity of an oxygen-radical-generating enzyme conjugated to a monoclonal antibody.

PubMed Central

Battelli, M G; Abbondanza, A; Tazzari, P L; Dinota, A; Rizzi, S; Grassi, G; Gobbi, M; Stirpe, F

1988-01-01

The monoclonal antibody 8A, which recognizes a human plasma cell-associated antigen, was covalently linked to xanthine oxidase in a conjugate maintaining both immunological and enzymatic properties. A significant degree of target cell lysis was obtained at an enzyme concentration that was ineffective on non-target cells and on myeloid staminal cells (CFU-GM). The cytotoxic activity was abolished by an excess of antibody, by allopurinol and by superoxide dismutase and catalase. A possible use of the conjugate for bone marrow purging in multiple myeloma patients is suggested. PMID:3262464

Formation of N-alkylpyrroles via intermolecular redox amination.

PubMed

Pahadi, Nirmal K; Paley, Miranda; Jana, Ranjan; Waetzig, Shelli R; Tunge, Jon A

2009-11-25

A wide variety of aldehydes, ketones, and lactols undergo redox amination when allowed to react with 3-pyrroline in the presence of a mild Brønsted acid catalyst. This reaction utilizes the inherent reducing power of 3-pyrroline to perform the equivalent of a reductive amination to form alkyl pyrroles. In doing so, the reaction avoids stoichiometric reducing agents that are typically associated with reductive aminations. Moreover, the redox amination protocol allows access to alkyl pyrroles that cannot be made via standard reductive amination.

Effect of salt-tolerant yeast of Candida versatilis and Zygosaccharomyces rouxii on the production of biogenic amines during soy sauce fermentation.

PubMed

Qi, Wei; Hou, Li-Hua; Guo, Hong-Lian; Wang, Chun-Ling; Fan, Zhen-Chuan; Liu, Jin-Fu; Cao, Xiao-Hong

2014-06-01

This study aimed to enhance and improve the quality and safety of soy sauce. In the present work, the change of biogenic amines, such as histamine, tyramine, cadaverine, spermidine, was examined by the treatment of Candida versatilis and Zygosaccharomyces rouxii, and the influence of salt-tolerant yeast on biogenic amines was analysed during the whole fermentation process. The results showed that the content of biogenic amines was elevated after yeast treatment and the content of biogenic amines was influenced by using yeast. The dominating biogenic amine in soy sauce was tyramine. At the end of fermentation, the concentrations of biogenic amines produced by Zygosaccharomyces rouxii and Candida versatilis in the soy mash were 122.71 mg kg(-1) and 69.96 mg kg(-1) . The changes of biogenic amines in high-salt liquid soy mash during fermentation process indicated that a variety of biogenic amines were increased in the fermentation ageing period, which may be due to amino acid decarboxylation to form biogenic amines by yeast decarboxylase. The fermentation period of soy sauce should be longer than 5 months because biogenic amines began to decline after this time period. © 2013 Society of Chemical Industry.

Angiotensin II receptor blocker improves the lowered exercise capacity and impaired mitochondrial function of the skeletal muscle in type 2 diabetic mice.

PubMed

Takada, Shingo; Kinugawa, Shintaro; Hirabayashi, Kagami; Suga, Tadashi; Yokota, Takashi; Takahashi, Masashige; Fukushima, Arata; Homma, Tsuneaki; Ono, Taisuke; Sobirin, Mochamad A; Masaki, Yoshihiro; Mizushima, Wataru; Kadoguchi, Tomoyasu; Okita, Koichi; Tsutsui, Hiroyuki

2013-04-01

NAD(P)H oxidase-induced oxidative stress is at least in part involved with lowered exercise capacity and impaired mitochondrial function in high-fat diet (HFD)-induced diabetic mice. NAD(P)H oxidase can be activated by activation of the renin-angiotensin system. We investigated whether ANG II receptor blocker can improve exercise capacity in diabetic mice. C57BL/6J mice were fed a normal diet (ND) or HFD, and each group of mice was divided into two groups: treatment with or without olmesartan (OLM; 3 mg·kg(-1)·day(-1) in the drinking water). The following groups of mice were studied: ND, ND+OLM, HFD, and HFD+OLM (n = 10 for each group). After 8 wk, HFD significantly increased body weight, plasma glucose, and insulin compared with ND, and OLM did not affect these parameters in either group. Exercise capacity, as determined by treadmill tests, was significantly reduced in HFD, and this reduction was ameliorated in HFD+OLM. ADP-dependent mitochondrial respiration was significantly decreased, and NAD(P)H oxidase activity and superoxide production by lucigenin chemiluminescence were significantly increased in skeletal muscle from HFD, which were attenuated by OLM. There were no such effects by OLM in ND. We concluded that OLM ameliorated the decrease in exercise capacity in diabetic mice via improvement in mitochondrial function and attenuation of oxidative stress in skeletal muscle. These data may have a clinical impact on exercise capacity in the medical treatment of diabetes mellitus.

Pharmacodynamics of norepinephrine reuptake inhibition: Modeling the peripheral and central effects of atomoxetine, duloxetine, and edivoxetine on the biomarker 3,4-dihydroxyphenylglycol in humans.

PubMed

Kielbasa, William; Lobo, Evelyn

2015-12-01

Norepinephrine, a neurotransmitter in the autonomic sympathetic nervous system, is deaminated by monoamine oxidase to 3,4-dihydroxyphenylglycol (DHPG). Inhibition of the NE transporter (NET) using DHPG as a biomarker was evaluated using atomoxetine, duloxetine, and edivoxetine as probe NET inhibitors. Pharmacokinetic and pharmacodynamic data were obtained from healthy subjects (n = 160) from 5 clinical trials. An indirect response model was used to describe the relationship between drug plasma concentration and DHPG concentration in plasma and cerebrospinal fluid (CSF). The baseline plasma DHPG concentration (1130-1240 ng/mL) and Imax (33%-37%) were similar for the 3 drugs. The unbound plasma drug IC50 (IC50U ) based on plasma DHPG was 0.973 nM for duloxetine, 0.136 nM for atomoxetine, and 0.041 nM for edivoxetine. The baseline CSF DHPG concentration (1850-2260 ng/mL) was similar for the 3 drugs, but unlike plasma DHPG, the Imax for DHPG was 38% for duloxetine, 53% for atomoxetine, and75% for edivoxetine. The IC50U based on CSF DHPG was 2.72 nM for atomoxetine, 1.22 nM for duloxetine, and 0.794 nM for edivoxetine. These modeling results provide insights into the pharmacology of NET inhibitors and the use of DHPG as a biomarker. © 2015, The American College of Clinical Pharmacology.

Viability and Functionality of Cryopreserved Peripheral Blood Mononuclear Cells in Pediatric Dengue

PubMed Central

Perdomo-Celis, Federico; Salgado, Doris M.; Castañeda, Diana M.

2016-01-01

Cryopreserved peripheral blood mononuclear cells (PBMCs) are widely used in studies of dengue. In this disease, elevated frequency of apoptotic PBMCs has been described, and molecules such as soluble tumor necrosis factor (TNF)-related apoptosis-inducing ligands (sTRAIL) are involved. This effect of dengue may affect the efficiency of PBMC cryopreservation. Here, we evaluate the viability (trypan blue dye exclusion and amine-reactive dye staining) and functionality (frequency of gamma interferon [IFN-γ]-producing T cells after polyclonal stimulation) of fresh and cryopreserved PBMCs from children with dengue (in acute and convalescence phases), children with other febrile illnesses, and healthy children as controls. Plasma sTRAIL levels were also evaluated. The frequencies of nonviable PBMCs detected by the two viability assays were positively correlated (r = 0.74; P < 0.0001). Cryopreservation particularly affected the PBMCs of children with dengue, who had a higher frequency of nonviable cells than healthy children and children with other febrile illnesses (P ≤ 0.02), and PBMC viability levels were restored in the convalescent phase. In the acute phase, an increased frequency of CD3+ CD8+ amine-positive cells was found before cryopreservation (P = 0.01). Except for B cells in the acute phase, cryopreservation usually did not affect the relative frequencies of viable PBMC subpopulations. Dengue infection reduced the frequency of IFN-γ-producing CD3+ cells after stimulation compared with healthy controls and convalescent-phase patients (P ≤ 0.003), and plasma sTRAIL correlated with this decreased frequency in dengue (rho = −0.56; P = 0.01). Natural dengue infection in children can affect the viability and functionality of cryopreserved PBMCs. PMID:26961858

Carbohydrate-based electrochemical biosensor for detection of a cancer biomarker in human plasma.

PubMed

Devillers, Marion; Ahmad, Lama; Korri-Youssoufi, Hafsa; Salmon, Laurent

2017-10-15

Autocrine motility factor (AMF) is a tumor-secreted cytokine that stimulates tumor cell motility in vitro and metastasis in vivo. AMF could be detected in serum or urine of cancer patients with worse prognosis. Reported as a cancer biomarker, AMF secretion into body fluids might be closely related to metastases formation. In this study, a sensitive and specific carbohydrate-based electrochemical biosensor was designed for the detection and quantification of a protein model of AMF, namely phosphoglucose isomerase from rabbit muscle (RmPGI). Indeed, RmPGI displays high homology with AMF and has been shown to have AMF activity. The biosensor was constructed by covalent binding of the enzyme substrate d-fructose 6-phosphate (F6P). Immobilization was achieved on a gold surface electrode following a bottom-up approach through an aminated surface obtained by electrochemical patterning of ethylene diamine and terminal amine polyethylene glycol chain to prevent non-specific interactions. Carbohydrate-protein interactions were quantified in a range of 10 fM to 100nM. Complex formation was analyzed through monitoring of the redox couple Fe 2+ /Fe 3+ by electrochemical impedance spectroscopy and square wave voltammetry. The F6P-biosensor demonstrates a detection limit of 6.6 fM and high selectivity when compared to other non-specific glycolytic proteins such as d-glucose-6-phosphate dehydrogenase. Detection of protein in spiked plasma was demonstrated and accuracy of 95% is obtained compared to result obtained in PBS (phosphate buffered saline). F6P-biosensor is a very promising proof of concept required for the design of a carbohydrate-based electrochemical biosensor using the enzyme substrate as bioreceptor. Such biosensor could be generalized to detect other protein biomarkers of interest. Copyright © 2017 Elsevier B.V. All rights reserved.

Metabolic profiles of male meat eaters, fish eaters, vegetarians, and vegans from the EPIC-Oxford cohort12

PubMed Central

Schmidt, Julie A; Rinaldi, Sabina; Ferrari, Pietro; Carayol, Marion; Achaintre, David; Scalbert, Augustin; Cross, Amanda J; Gunter, Marc J; Fensom, Georgina K; Appleby, Paul N; Key, Timothy J; Travis, Ruth C

2015-01-01

Background: Human metabolism is influenced by dietary factors and lifestyle, environmental, and genetic factors; thus, men who exclude some or all animal products from their diet might have different metabolic profiles than meat eaters. Objective: We aimed to investigate differences in concentrations of 118 circulating metabolites, including acylcarnitines, amino acids, biogenic amines, glycerophospholipids, hexose, and sphingolipids related to lipid, protein, and carbohydrate metabolism between male meat eaters, fish eaters, vegetarians, and vegans from the Oxford arm of the European Prospective Investigation into Cancer and Nutrition. Design: In this cross-sectional study, concentrations of metabolites were measured by mass spectrometry in plasma from 379 men categorized according to their diet group. Differences in mean metabolite concentrations across diet groups were tested by using ANOVA, and a false discovery rate–controlling procedure was used to account for multiple testing. Principal component analysis was used to investigate patterns in metabolic profiles. Results: Concentrations of 79% of metabolites differed significantly by diet group. In the vast majority of these cases, vegans had the lowest concentration, whereas meat eaters most often had the highest concentrations of the acylcarnitines, glycerophospholipids, and sphingolipids, and fish eaters or vegetarians most often had the highest concentrations of the amino acids and a biogenic amine. A clear separation between patterns in the metabolic profiles of the 4 diet groups was seen, with vegans being noticeably different from the other groups because of lower concentrations of some glycerophospholipids and sphingolipids. Conclusions: Metabolic profiles in plasma could effectively differentiate between men from different habitual diet groups, especially vegan men compared with men who consume animal products. The difference in metabolic profiles was mainly explained by the lower concentrations of glycerophospholipids and sphingolipids in vegans. PMID:26511225

Metabolic profiles of male meat eaters, fish eaters, vegetarians, and vegans from the EPIC-Oxford cohort.

PubMed

Schmidt, Julie A; Rinaldi, Sabina; Ferrari, Pietro; Carayol, Marion; Achaintre, David; Scalbert, Augustin; Cross, Amanda J; Gunter, Marc J; Fensom, Georgina K; Appleby, Paul N; Key, Timothy J; Travis, Ruth C

2015-12-01

Human metabolism is influenced by dietary factors and lifestyle, environmental, and genetic factors; thus, men who exclude some or all animal products from their diet might have different metabolic profiles than meat eaters. We aimed to investigate differences in concentrations of 118 circulating metabolites, including acylcarnitines, amino acids, biogenic amines, glycerophospholipids, hexose, and sphingolipids related to lipid, protein, and carbohydrate metabolism between male meat eaters, fish eaters, vegetarians, and vegans from the Oxford arm of the European Prospective Investigation into Cancer and Nutrition. In this cross-sectional study, concentrations of metabolites were measured by mass spectrometry in plasma from 379 men categorized according to their diet group. Differences in mean metabolite concentrations across diet groups were tested by using ANOVA, and a false discovery rate-controlling procedure was used to account for multiple testing. Principal component analysis was used to investigate patterns in metabolic profiles. Concentrations of 79% of metabolites differed significantly by diet group. In the vast majority of these cases, vegans had the lowest concentration, whereas meat eaters most often had the highest concentrations of the acylcarnitines, glycerophospholipids, and sphingolipids, and fish eaters or vegetarians most often had the highest concentrations of the amino acids and a biogenic amine. A clear separation between patterns in the metabolic profiles of the 4 diet groups was seen, with vegans being noticeably different from the other groups because of lower concentrations of some glycerophospholipids and sphingolipids. Metabolic profiles in plasma could effectively differentiate between men from different habitual diet groups, especially vegan men compared with men who consume animal products. The difference in metabolic profiles was mainly explained by the lower concentrations of glycerophospholipids and sphingolipids in vegans.

Nitrosamine formation in amine scrubbing at desorber temperatures.

PubMed

Fine, Nathan A; Goldman, Mark J; Rochelle, Gary T

2014-01-01

Amine scrubbing is a thermodynamically efficient and industrially proven method for carbon capture, but amine solvents can nitrosate in the desorber, forming potentially carcinogenic nitrosamines. The kinetics of reactions involving nitrite and monoethanolamine (MEA), diethanolamine (DEA), methylethanolamine (MMEA), and methyldiethanolamine (MDEA) were determined under desorber conditions. The nitrosations of MEA, DEA, and MMEA are first order in nitrite, carbamate species, and hydronium ion. Nitrosation of MDEA, a tertiary amine, is not catalyzed by the addition of CO2 since it cannot form a stable carbamate. Concentrated and CO2 loaded MEA was blended with low concentrations of N-(2-hydroxyethyl) glycine (HeGly), hydroxyethyl-ethylenediamine (HEEDA), and DEA, secondary amines common in MEA degradation. Nitrosamine yield was proportional to the concentration of secondary amine and was a function of CO2 loading and temperature. Blends of tertiary amines with piperazine (PZ) showed n-nitrosopiperazine (MNPZ) yields close to unity, validating the slow nitrosation rates hypothesized for tertiary amines. These results provide a useful tool for estimating nitrosamine accumulation over a range of amine solvents.

Bioavailability of the ferulic acid-derived phenolic compounds of a rice bran enzymatic extract and their activity against superoxide production.

PubMed

Perez-Ternero, Cristina; Macià, Alba; de Sotomayor, Maria Alvarez; Parrado, Juan; Motilva, Maria-Jose; Herrera, Maria-Dolores

2017-06-21

Rice bran is an exceptional source of such antioxidant molecules as γ-oryzanol and ferulic acid, but their bioavailability and metabolism within this matrix remain unknown. The aims of this work were to describe the oral bioavailability and metabolic pathways of the ferulic acid-derived phenolic compounds contained in a rice bran enzymatic extract (RBEE), and to determine its effect on NADPH oxidase activity. Wistar rats were administered with RBEE and sacrificed at different times over a period of 24 h to obtain plasma. An additional group was used for collection of urine and faeces over a period of 48 h. The phenolic metabolites were determined by ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS), and plasma pharmacokinetic parameters were calculated. In parallel, aortic rings were incubated in the plasma of rats sacrificed 30 min after RBEE gavage, or in the presence of RBEE, ferulic acid or γ-oryzanol. Endothelin-1-induced superoxide production was recorded by lucigenin-enhanced luminescence. Twenty-five ferulic acid metabolites showing biphasic behaviour were found in the plasma, most of which were found in the urine as well, while in the faeces, colonic metabolism led to simpler phenolic compounds. Superoxide production was abrogated by phenolic compound-enriched plasma and by RBEE and ferulic acid, thus showing the biological potential of RBEE as a nutraceutical ingredient.

Adsorption of Wine Constituents on Functionalized Surfaces.

PubMed

Mierczynska-Vasilev, Agnieszka; Smith, Paul A

2016-10-18

The adsorption of macromolecules on solid surfaces is of great importance in the field of nanotechnology, biomaterials, biotechnological, and food processes. In the field of oenology adsorption of wine macromolecules such as polyphenols, polysaccharides, and proteins is much less desirable on membrane materials because of fouling and reduced filtering performance. On the other hand, adsorption of these molecules on processing aids is very beneficial for achieving wine clarity and stability. In this article, the effect of surface chemical functionalities on the adsorption of white, rosé, and red wine constituents was evaluated. Allylamine, acrylic acid, and ethanol were selected as precursors for plasma polymerization in order to generate coatings rich in amine, carboxyl, and hydroxyl chemical groups, respectively. The surface chemical functionalities were characterized by X-ray photoelectron spectroscopy (XPS) and the ability of different surface chemical functionalities to adsorb wine constituents were characterized by quartz crystal microbalance with dissipation (QCM-D) and atomic force microscopy (AFM). The results demonstrated that the amine and carboxyl modified surfaces encourage adsorption of constituents from white wine. The hydroxyl modified surfaces have the ability to preferentially adsorb rosé wine constituents, whereas red wine adsorbed to the highest extent on acrylic acid surface.

2-(Maleimidomethyl)-1,3-Dioxanes (MD): a Serum-Stable Self-hydrolysable Hydrophilic Alternative to Classical Maleimide Conjugation

NASA Astrophysics Data System (ADS)

Dovgan, Igor; Kolodych, Sergii; Koniev, Oleksandr; Wagner, Alain

2016-08-01

The vast majority of antibody-drug conjugates (ADC) are prepared through amine-to-thiol conjugation. To date, N-Succinimidyl-4-(maleimidomethyl) cyclohexanecarboxylate (SMCC) has been one of the most frequently applied reagents for the preparation of ADC and other functional conjugates. However, SMCC-based conjugates suffer from limited stability in blood circulation and from a hydrophobic character of the linker, which may give rise to major pharmacokinetic implications. To address this issue, we have developed a heterobifunctional analogue of a SMCC reagent, i.e., sodium 4-(maleimidomethyl)-1,3-dioxane-5-carbonyl)oxy)-2,3,5,6- tetrafluorobenzenesulfonate (MDTF) for amine-to-thiol conjugation. By replacing the cyclohexyl ring in the SMCC structure with the 1,3-dioxane, we increased the hydrophilicity of the linker. A FRET probe based on MD linker was prepared and showed superior stability compared to the MCC linker in human plasma, as well as in a variety of aqueous buffers. A detailed investigation demonstrated an accelerated succinimide ring opening for MD linker, resulting in stabilized conjugates. Finally, the MDTF reagent was applied for the preparation of serum stable antibody-dye conjugate.

Poly(amido)amine (PAMAM) dendrimer-cisplatin complexes for chemotherapy of cisplatin-resistant ovarian cancer cells

NASA Astrophysics Data System (ADS)

Yellepeddi, Venkata Kashyap; Vangara, Kiran Kumar; Palakurthi, Srinath

2013-09-01

Dendrimer-cisplatin complexes were prepared using PAMAM dendrimers with terminal -NH2 and -COOH groups as well as biotin-conjugated dendrimers. Preformulation parameters of dendrimer-cisplatin complexes were studied using differential scanning calorimetry (DSC) and inductively coupled plasma-mass spectrometry (ICP-MS). Cytotoxicity and mechanism of cytotoxicity of dendrimer-cisplatin complexes was investigated in OVCAR-3, SKOV, A2780 and cisplatin-resistant CP70 human ovarian cancer cell lines. The loading of cisplatin in dendrimers was 11 % (w/w). PAMAM G4 dendrimers with amine surface groups (biotinylated and native) have shown 2.5- to 3.0-fold reduction in IC50 values in ovarian cancer cells when compared with carboxylate surface dendrimers ( p < 0.05). A correlation was observed among cytotoxicity of the complexes, cellular uptake, and platinum-DNA adduct formation. Treatment with dendrimer-cisplatin complexes resulted in a 7.0-fold increase ( p < 0.05) in expression of apoptotic genes ( Bcl2, Bax, p53) and 13.2- to 27.1-fold increase ( p < 0.05) in the activity of caspases 3, 8, and 9 in vitro. Results suggest that PAMAM dendrimers can be used as potential carrier for cisplatin chemotherapy of ovarian cancer.

Striking Confinement Effect: AuCl[subscript 4][superscript -] Binding to Amines in a Nanocage Cavity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Henao, Juan D.; Suh, Young-Woong; Lee, Jeong-Kyu

2009-02-23

Binding of AuCl{sub 4}{sup -} to amine groups tethered to the interior of a 2 nm siloxane nanocage was determined in solutions containing various concentrations of acid. The mode of binding was inferred from EXAFS and UV-vis spectra to be by ligand exchange of amine for chloride, which implies that the amines remain unprotonated. Cyclic voltammetry confirmed that the Au complexes bind to the nanocage interior and established a 1:1 relationship between bound Au complex and amine groups. The results suggested a 5-7 pH unit shift in the protonation constant of the interior amines relative to free amines in solution.

Transition Metal Free C-N Bond Forming Dearomatizations and Aryl C-H Aminations by in Situ Release of a Hydroxylamine-Based Aminating Agent.

PubMed

Farndon, Joshua J; Ma, Xiaofeng; Bower, John F

2017-10-11

We outline a simple protocol that accesses directly unprotected secondary amines by intramolecular C-N bond forming dearomatization or aryl C-H amination. The method is dependent on the generation of a potent electrophilic aminating agent released by in situ deprotection of O-Ts activated N-Boc hydroxylamines.

Bioinspired organocatalytic aerobic C-H oxidation of amines with an ortho-quinone catalyst.

PubMed

Qin, Yan; Zhang, Long; Lv, Jian; Luo, Sanzhong; Cheng, Jin-Pei

2015-03-20

A simple bioinspired ortho-quinone catalyst for the aerobic oxidative dehydrogenation of amines to imines is reported. Without any metal cocatalysts, the identified optimal ortho-quinone catalyst enables the oxidations of α-branched primary amines and cyclic secondary amines. Mechanistic studies have disclosed the origins of different performances of ortho-quinone vs para-quinone in biomimetic amine oxidations.

Experimental investigation and DFT calculation of different amine/ammonium salts adsorption on kaolinite

NASA Astrophysics Data System (ADS)

Chen, Jun; Min, Fan-fei; Liu, Lingyun; Liu, Chunfu; Lu, Fangqin

2017-10-01

The adsorption of four different amine/ammonium salts of DDA (Dodecyl amine), MDA (N-methyldodecyl amine), DMDA (N,N-dimethyldodecyl amine) and DTAC (Dodecyl trimethyl ammonium chloride) on kaolinite particles was investigated in the study through the measurement of contact angles, zeta potentials, aggregation observation, adsorption and sedimentation. The results show that different amine/ammonium salts can adsorb on the kaolinite surface to enhance the hydrophobicity and reduce the electronegativity of kaolinite particle surface, and thus induce a strong hydrophobic aggregation of kaolinite particles which promotes the settlement of kaolinite. To explore the adsorption mechanism of these four amine/ammonium salts on kaolinite surfaces, the adsorptions of DDA+, MDA+, DMDA+ and DTAC+ on kaolinite (001) surface and (00 1 bar) surface are calculated with DFT (Density functional theory). The DFT calculation results indicate that different amine/ammonium cations can strongly adsorbed on kaolinite (001) surface and (00 1 bar) surface by forming Nsbnd H⋯O strong hydrogen bonds or Csbnd H⋯O weak hydrogen bonds, and there are strongly electrostatic attractions between different amine/ammonium cations and kaolinite surfaces. The main adsorption mechanism of amine/ammonium cations on kaolinite is hydrogen-bond interaction and electrostatic attraction.

Oxidations of N-(3-indoleethyl) cyclic aliphatic amines by horseradish peroxidase: the indole ring binds to the enzyme and mediates electron-transfer amine oxidation.

PubMed

Ling, Ke-Qing; Li, Wen-Shan; Sayre, Lawrence M

2008-01-23

Although oxidations of aromatic amines by horseradish peroxidase (HRP) are well-known, typical aliphatic amines are not substrates of HRP. In this study, the reactions of N-benzyl and N-methyl cyclic amines with HRP were found to be slow, but reactions of N-(3-indoleethyl) cyclic amines were 2-3 orders of magnitude faster. Analyses of pH-rate profiles revealed a dominant contribution to reaction by the amine-free base forms, the only species found to bind to the enzyme. A metabolic study on a family of congeneric N-(3-indoleethyl) cyclic amines indicated competition between amine and indole oxidation pathways. Amine oxidation dominated for the seven- and eight-membered azacycles, where ring size supports the change in hybridization from sp3 to sp2 that occurs upon one-electron amine nitrogen oxidation, whereas only indole oxidation was observed for the six-membered ring congener. Optical difference spectroscopic binding data and computational docking simulations suggest that all the arylalkylamine substrates bind to the enzyme through their aromatic termini with similar binding modes and binding affinities. Kinetic saturation was observed for a particularly soluble substrate, consistent with an obligatory role of an enzyme-substrate complexation preceding electron transfer. The significant rate enhancements seen for the indoleethylamine substrates suggest the ability of the bound indole ring to mediate what amounts to medium long-range electron-transfer oxidation of the tertiary amine center by the HRP oxidants. This is the first systematic investigation to document aliphatic amine oxidation by HRP at rates consistent with normal metabolic turnover, and the demonstration that this is facilitated by an auxiliary electron-rich aromatic ring.

A point mutation of valine-311 to methionine in Bacillus subtilis protoporphyrinogen oxidase does not greatly increase resistance to the diphenyl ether herbicide oxyfluorfen.

PubMed

Jeong, Eunjoo; Houn, Thavrak; Kuk, Yongin; Kim, Eun-Seon; Chandru, Hema Kumar; Baik, Myunggi; Back, Kyoungwhan; Guh, Ja-Ock; Han, Oksoo

2003-10-01

In an effort to asses the effect of Val311Met point mutation of Bacillus subtilis protoporphyrinogen oxidase on the resistance to diphenyl ether herbicides, a Val311Met point mutant of B. subtilis protoporphyrinogen oxidase was prepared, heterologously expressed in Escherichia coli, and the purified recombinant Val311Met mutant protoporphyrinogen oxidase was kinetically characterized. The mutant protoporphyrinogen oxidase showed very similar kinetic patterns to wild type protoporphyrinogen oxidase, with slightly decreased activity dependent on pH and the concentrations of NaCl, Tween 20, and imidazole. When oxyfluorfen was used as a competitive inhibitor, the Val311Met mutant protoporphyrinogen oxidase showed an increased inhibition constant about 1.5 times that of wild type protoporphyrinogen oxidase. The marginal increase of the inhibition constant indicates that the Val311Met point mutation in B. subtilis protoporphyrinogen oxidase may not be an important determinant in the mechanism that protects protoporphyrinogen oxidase against diphenyl ether herbicides.

RECOVERY OF URANIUM FROM AQUEOUS PHOSPHATE-CONTAINING SOLUTIONS

DOEpatents

Igelsrud, I.; Stephen, E.F.

1959-08-11

ABS>A method is presented for recovering hexavalent uranium from an acidic phosphaie solution. A high molecular weight amine, such as a mixture of cccoanut oil amines, is added to the solution in such amount as to give a ratio of about 2000 parts by weight of amine to 1 part by weight of uranium. The uranium is precipitated with the amines and the whole filtered from the solution. The uranium is leached from the amine mass by washing with aqueous sodium carbonate solution; and the amine mixture is available for reuse.

Controlling Nitrosamines, Nitramines, and Amines in Amine-Based CO₂ Capture Systems with Continuous Ultraviolet and Ozone Treatment of Washwater.

PubMed

Dai, Ning; Mitch, William A

2015-07-21

Formation of nitrosamines and nitramines from reactions between flue gas NOx and the amines used in CO2 capture units has arisen as a significant concern. Washwater scrubbers can capture nitrosamines and nitramines. They can also capture amines, preventing formation of nitrosamines and nitramines downwind by amine reactions with ambient NOx. The continuous application of UV alone, or a combination of UV and ozone to the return line of a washwater treatment unit was evaluated to control the accumulation of nitrosamines, nitramines and amines in a laboratory-scale washwater unit. With model secondary amine solvents ranging from nonvolatile diethanolamine to volatile morpholine, application of 272-537 mJ/cm(2) UV incident fluence alone reduced the accumulation of nitrosamines and nitramines by approximately an order of magnitude. Modeling indicated that the gains achieved by UV treatment should increase over time, because UV treatment converts the time dependence of nitrosamine accumulation from a quadratic to a linear function. Ozone (21 mg/L) maintained low steady-state concentrations of amines in the washwater. While modeling indicated that more than 80% of nitrosamine accumulation in the washwater was associated with reaction of washwater amines with residual NOx, a reduction in nitrosamine accumulation rates due to ozone oxidation of amines was not fully realized because the ozonation products of amines reduced nitrosamine photolysis rates by competing for photons.

SO 2-Resistant Immobilized Amine Sorbents for CO 2 Capture

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tumuluri, Uma

2014-01-01

The solid amine sorbent for CO 2 capture process has advantages of simplicity and low operating cost compared to the MEA (monoethanolamine) process. Solid amine sorbents reported so far suffered from either low CO 2 capture capacity or low stability in the flue gas environment. This project is aimed at developing a SO 2-resistant solid amine sorbent for capturing CO 2 from coal–fired power plants with SCR/FGD which emits SO 2ranging from 15 to 30 ppm and NO ranging from 5 to 10 ppm. The amine sorbent we developed in a previous project degraded rapidly with 65% decrease in themore » initial capture capacity in presence of 1% SO 2. This amine sorbent was further modified by coating with polyethyleneglycol (PEG) to increase the SO 2-resistance. Polyethylene glycol (PEG) was found to decrease the SO 2-amine interaction, resulting in the decrease in the maximum SO desorption temperature (Tmax ) of amine sorbent. The PEG-coated amine sorbent exhibited higher stability with only 40% decrease in the initial capture capacity compared to un-coated amine sorbents. The cost of the solid amine sorbent developed in this project is estimated to be less than $7.00/lb; the sorbent exhibited CO 2 capture capacity more than 2.3 mmol/g. The results of this study provided the scientific basis for further development of SO 2-resistant sorbents.« less

Multipotent MAO and cholinesterase inhibitors for the treatment of Alzheimer's disease: synthesis, pharmacological analysis and molecular modeling of heterocyclic substituted alkyl and cycloalkyl propargyl amine.

PubMed

Samadi, Abdelouahid; de los Ríos, Cristóbal; Bolea, Irene; Chioua, Mourad; Iriepa, Isabel; Moraleda, Ignacio; Bartolini, Manuela; Andrisano, Vincenza; Gálvez, Enrique; Valderas, Carolina; Unzeta, Mercedes; Marco-Contelles, José

2012-06-01

The synthesis, pharmacological evaluation and molecular modeling of heterocyclic substituted alkyl and cycloalkyl propargyl amines 1-7 of type I, and 9-12 of type II, designed as multipotent inhibitors able to simultaneously inhibit monoamine oxidases (MAO-A/B) as well as cholinesterase (AChE/BuChE) enzymes, as potential drugs for the treatment of Alzheimer's disease, are described. Indole derivatives 1-7 of type I are well known MAO inhibitors whose capacity to inhibit AChE and BuChE was here investigated for the first time. As a result, compound 7 was identified as a MAO-B inhibitor (IC(50) = 31 ± 2 nM) and a moderately selective eqBuChE inhibitor (IC(50) = 4.7 ± 0.2 μM). Conversely, the new and readily available 5-amino-7-(prop-2-yn-1-yl)-6,7,8,9-tetrahydropyrido[2,3-b][1,6]naphthyridine derivatives 9-13 of type II are poor MAO inhibitors, but showed AChE selective inhibition, compound 12 being the most attractive as it acts as a non-competitive inhibitor on EeAChE (IC(50) = 25 ± 3 nM, K(i) = 65 nM). The ability of this compound to interact with the AChE peripheral binding site was confirmed by kinetic studies and by molecular modeling investigation. Studies on human ChEs confirmed that 12 is a selective AChE inhibitor with inhibitory potency in the submicromolar range. Moreover, in agreement with its mode of action, 12 was shown to be able to inhibit Aβ aggregation induced by hAChE by 30.6%. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Factors influencing the regioselectivity of the oxidation of asymmetric secondary amines with singlet oxygen.

PubMed

Ushakov, Dmitry B; Plutschack, Matthew B; Gilmore, Kerry; Seeberger, Peter H

2015-04-20

Aerobic amine oxidation is an attractive and elegant process for the α functionalization of amines. However, there are still several mechanistic uncertainties, particularly the factors governing the regioselectivity of the oxidation of asymmetric secondary amines and the oxidation rates of mixed primary amines. Herein, it is reported that singlet-oxygen-mediated oxidation of 1° and 2° amines is sensitive to the strength of the α-C-H bond and steric factors. Estimation of the relative bond dissociation energy by natural bond order analysis or by means of one-bond C-H coupling constants allowed the regioselectivity of secondary amine oxidations to be explained and predicted. In addition, the findings were utilized to synthesize highly regioselective substrates and perform selective amine cross-couplings to produce imines. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of pitavastatin in different SLCO1B1 backgrounds on repaglinide pharmacokinetics and pharmacodynamics in healthy Chinese males.

PubMed

Zhu, Jie; Song, Min; Tan, Hong-Yi; Huang, Li-Hua; Huang, Zhi-Jun; Liu, Chang; Fu, Zhi-Min; Huang, Yuan-Yuan; Tan, Zhi-Rong; Chen, Xiao-Ping; Yuan, Hong; Yang, Guo-Ping

2013-05-01

The effect of pitavastatin and SLCO1B1 genetic background on the pharmacokinetic and pharmacodynamic properties of repaglinide was investigated. In this randomized, placebo-controlled, crossover study, twelve healthy Chinese males were administered with pitavastatin 4 mg/d or the placebo for 5 d followed by repaglinide 4 mg given orally on d 5. Plasma repaglinide and glucose levels were measured by liquid chromatography-tandem mass spectrometry (LC/MS/MS) and the glucose oxidase method, respectively. Treatment with pitavastatin significantly increased the peak plasma concentration (Cmax) of repaglinide (P=0.003) in SLCO1B1*1b homozygotes (P=0.015) and SLCO1B1*15 carriers (P=0.031). Treatment with pitavastatin led to a marginal increase in the area under plasma concentration-time curve from 0 h to infinity (AUC0⇒∞) of repaglinide (P=0.091). There was no significant difference in pharmacokinetic parameters or hypoglycemic effects of repaglinide among SLCO1B1 genotypes in either the pitavastatin or control group. Pitavastatin increased the Cmax of the plasma concentration of repaglinide in an SLCO1B1 genotype dependent manner, but had no apparent effect on the pharmacodynamics of repaglinide in healthy volunteers. The p values for this statement were not reported.

Stereospecificity of phenylalanine plasma kinetics and hydroxylation in man following oral application of a stable isotope-labelled pseudo-racemic mixture of L- and D-phenylalanine.

PubMed

Lehmann, W D; Theobald, N; Fischer, R; Heinrich, H C

1983-03-14

L-[15N]Phenylalanine and D-[2H5]phenylalanine have been administered orally to two healthy adult volunteers as a pseudo-racemic mixture at a dose of 25 mg/kg each. After oral application, the plasma kinetics of phenylalanine and tyrosine have been followed by the combined use of high pressure liquid chromatography and field desorption mass spectrometry. Additional incubation with D-amino acid oxidase was used to determine the enantiomeric composition of the differently labelled species of phenylalanine and tyrosine. D-Phenylalanine plasma levels show a faster rise to higher maximum values compared to L-phenylalanine (D/L ratio at maximum 3.19, 3.26). L-Phenylalanine is efficiently hydroxylated to L-tyrosine. In contrast, conversion of D-phenylalanine to the L-form with subsequent hydroxylation to L-tyrosine was observed. From the plasma kinetics it is estimated that about 1/3 of the applied dose of 25 mg/kg of D-phenylalanine is converted to the L-isomer. Of the administered dose of L-phenylalanine only very small amounts are excreted into urine as such (0.25%, 0.8%), whereas a substantial amount of the D-phenylalanine dose is found in urine (27.4%, 38.0%).

Knölker's iron complex: an efficient in situ generated catalyst for reductive amination of alkyl aldehydes and amines.

PubMed

Pagnoux-Ozherelyeva, Anastassiya; Pannetier, Nicolas; Mbaye, Mbaye Diagne; Gaillard, Sylvain; Renaud, Jean-Luc

2012-05-14

An aminated series: a well-defined iron-catalyzed reductive amination reaction of aldehydes and ketones with aliphatic amines using molecular hydrogen is presented. Under mild conditions, good yields for a broad range of alkyl ketones as well as aldehydes were achieved. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of Seed Treatment by Cold Plasma on the Resistance of Tomato to Ralstonia solanacearum (Bacterial Wilt)

PubMed Central

Jiang, Jiafeng; Lu, Yufang; Li, Jiangang; Li, Ling; He, Xin; Shao, Hanliang; Dong, Yuanhua

2014-01-01

This study investigated the effect of cold plasma seed treatment on tomato bacterial wilt, caused by Ralstonia solanacearum (R. solanacearum), and the regulation of resistance mechanisms. The effect of cold plasma of 80W on seed germination, plant growth, nutrient uptake, disease severity, hydrogen peroxide (H2O2) concentration and activities of peroxidase (POD; EC 1.11.1.7), polyphenol oxidase (PPO; EC 1.10.3.2) and phenylalanine ammonia lyase (PAL; EC 4.3.1.5) were examined in tomato plants. Plasma treatment increased tomato resistance to R. solanacearum with an efficacy of 25.0%. Plasma treatment significantly increased both germination and plant growth in comparison with the control treatment, and plasma-treated plants absorbed more calcium and boron than the controls. In addition, H2O2 levels in treated plants rose faster and reached a higher peak, at 2.579 µM gFW−1, 140% greater than that of the control. Activities of POD (421.3 U gFW−1), PPO (508.8 U gFW−1) and PAL (707.3 U gFW−1) were also greater in the treated plants than in the controls (103.0 U gFW−1, 166.0 U gFW−1 and 309.4 U gFW−1, respectively). These results suggest that plasma treatment affects the regulation of plant growth, H2O2 concentration, and POD, PPO and PAL activity in tomato, resulting in an improved resistance to R. solanacearum. Consequently, cold plasma seed treatment has the potential to control tomato bacterial wilt caused by R. solanacearum. PMID:24840508

KDM1 Class Flavin-Dependent Protein Lysine Demethylases

PubMed Central

Burg, Jonathan M.; Link, Jennifer E.; Morgan, Brittany S.; Heller, Frederick J.; Hargrove, Amanda E.; McCafferty, Dewey G.

2015-01-01

Flavin-dependent, lysine-specific protein demethylases (KDM1s) are a subfamily of amine oxidases that catalyze the selective posttranslational oxidative demethylation of methyllysine side chains within protein and peptide substrates. KDM1s participate in the widespread epigenetic regulation of both normal and disease state transcriptional programs. Their activities are central to various cellular functions, such as hematopoietic and neuronal differentiation, cancer proliferation and metastasis, and viral lytic replication and establishment of latency. Interestingly, KDM1s function as catalytic subunits within complexes with coregulatory molecules that modulate enzymatic activity of the demethylases and coordinate their access to specific substrates at distinct sites within the cell and chromatin. Although several classes of KDM1 -selective small molecule inhibitors have been recently developed, these pan-active site inhibition strategies lack the ability to selectively discriminate between KDM1 activity in specific, and occasionally opposing, functional contexts within these complexes. Here we review the discovery of this class of demethylases, their structures, chemical mechanisms, and specificity. Additionally, we review inhibition of this class of enzymes as well as emerging interactions with coregulatory molecules that regulate demethylase activity in highly specific functional contexts of biological and potential therapeutic importance. PMID:25787087

Transrepressive function of TLX requires the histone demethylase LSD1.

PubMed

Yokoyama, Atsushi; Takezawa, Shinichiro; Schüle, Roland; Kitagawa, Hirochika; Kato, Shigeaki

2008-06-01

TLX is an orphan nuclear receptor (also called NR2E1) that regulates the expression of target genes by functioning as a constitutive transrepressor. The physiological significance of TLX in the cytodifferentiation of neural cells in the brain is known. However, the corepressors supporting the transrepressive function of TLX have yet to be identified. In this report, Y79 retinoblastoma cells were subjected to biochemical techniques to purify proteins that interact with TLX, and we identified LSD1 (also called KDM1), which appears to form a complex with CoREST and histone deacetylase 1. LSD1 interacted with TLX directly through its SWIRM and amine oxidase domains. LSD1 potentiated the transrepressive function of TLX through its histone demethylase activity as determined by a luciferase assay using a genomically integrated reporter gene. LSD1 and TLX were recruited to a TLX-binding site in the PTEN gene promoter, accompanied by the demethylation of H3K4me2 and deacetylation of H3. Knockdown of either TLX or LSD1 derepressed expression of the endogenous PTEN gene and inhibited cell proliferation of Y79 cells. Thus, the present study suggests that LSD1 is a prime corepressor for TLX.

Ontogenesis of uptake and deamination of 5-hydroxytryptamine, dopamine and beta-phenylethylamine in isolated perfused lung and lung homogenates from rats.

PubMed Central

Ben-Harari, R. R.; Youdim, M. B.

1981-01-01

1. Uptake of 5-hydroxytryptamine (5-HT) and beta-phenylethylamine (PEA) was studied in perfused lung from male rats between 10 and 70 days old. 2. Monoamine oxidase (MAO) activity towards 5-HT, PEA and dopamine was studied in homogenate preparations of lung from rats aged between 5 and 80 days. 3. Uptake of 5-HT (10 microM) decreased throughout the age range studied but uptake of PEA (50 microM) increased for the first 30 days and beyond this age it decreased. Metabolites formed for both amines reflected the changes in uptake. 4. MAO activity deaminating 5-HT is well developed by day 10 and reaches its maximum by day 40. For dopamine and PEA, MAO activity remained low until day 20, and the developed rapidly, reaching a maximum by day 40 for dopamine; activity towards PEA did not reach a maximum by day 80. 5. These results show that uptake and MAO activity changes with age and thus the lung responds like other tissues. 6. These results also demonstrate the independent development of uptake and MAO activity towards 5-HT, PEA and dopamine. PMID:7284689

The metabolism of 5-hydroxytryptamine and beta-phenylethylamine in perfused rat lung and in vitro.

PubMed Central

Bakhle, Y S; Youdim, M B

1979-01-01

1 Metabolism of 5-hydroxytryptamine (5-HT) and beta-phenylethylamine (PHE) by monoamine oxidase (MAO) was investigated in rat isolated lungs and in mitochondrial preparations from rat lung. 2. In perfused lungs 5-HT metabolism had an apparent Km of 2 microgram and PHE metaoblism a Km of 54 microgram, whereas in vitro the Km values were 330 microgram and 28 microgram respectively. 3 In vitro, MAO activity had substrate and inhibitor specificities compatible with the presence of A and B types of MAO. 4 In perfused lung, metabolism of 5-HT but not that of PHE was inhibited by desmethylimipramine. 5 These results show that PHE metabolism in perfused lung, unlike that of other metabolized amines, is not limited by transport and the transport process for PHE is unlike that of 5-HT or noradrenaline. 6 These results also show that the kinetic parameters obtained for MAO activity in vitro do not generally apply to the isolated lung where transport of substrate can be the deciding factor. This discrepancy emphasizes that the enzymic properties of the whole organ cannot relaibly be deduced from its enzymic content. PMID:32944

Systems biology approach in Chlamydomonas reveals connections between copper nutrition and multiple metabolic steps.

PubMed

Castruita, Madeli; Casero, David; Karpowicz, Steven J; Kropat, Janette; Vieler, Astrid; Hsieh, Scott I; Yan, Weihong; Cokus, Shawn; Loo, Joseph A; Benning, Christoph; Pellegrini, Matteo; Merchant, Sabeeha S

2011-04-01

In this work, we query the Chlamydomonas reinhardtii copper regulon at a whole-genome level. Our RNA-Seq data simulation and analysis pipeline validated a 2-fold cutoff and 10 RPKM (reads per kilobase of mappable length per million mapped reads) (~1 mRNA per cell) to reveal 63 CRR1 targets plus another 86 copper-responsive genes. Proteomic and immunoblot analyses captured 25% of the corresponding proteins, whose abundance was also dependent on copper nutrition, validating transcriptional regulation as a major control mechanism for copper signaling in Chlamydomonas. The impact of copper deficiency on the expression of several O₂-dependent enzymes included steps in lipid modification pathways. Quantitative lipid profiles indicated increased polyunsaturation of fatty acids on thylakoid membrane digalactosyldiglycerides, indicating a global impact of copper deficiency on the photosynthetic apparatus. Discovery of a putative plastid copper chaperone and a membrane protease in the thylakoid suggest a mechanism for blocking copper utilization in the chloroplast. We also found an example of copper sparing in the N assimilation pathway: the replacement of copper amine oxidase by a flavin-dependent backup enzyme. Forty percent of the targets are previously uncharacterized proteins, indicating considerable potential for new discovery in the biology of copper.

Cerebral cortical blood flow maps are reorganized in MAOB-deficient mice

PubMed Central

Scremin, Oscar U.; Holschneider, Daniel P.; Chen, Kevin; Li, Mingen G.; Shih, Jean C.

2014-01-01

Cerebral cortical blood flow (CBF) was measured autoradiographically in conscious mice without the monoamine oxidase B (MAOB) gene (KO, n = 11) and the corresponding wild-type animals (WILD, n = 11). Subgroups of animals of each genotype received a continuous intravenous infusion over 30 min of phenylethylamine (PEA), an endogenous substrate of MAOB, (8 nmol g−1 min−1 in normal saline at a volume rate of 0.11 μl g−1 min−1) or saline at the same volume rate. Maps of relative CBF distribution showed predominance of midline motor and sensory area CBF in KO mice over WILD mice that received saline. PEA enhanced CBF in lateral frontal and piriform cortex in both KO and WILD mice. These changes may reflect a differential activation due to chronic and acute PEA elevations on motor and olfactory function, as well as on the anxiogenic effects of this amine. In addition to its effects on regional CBF distribution, PEA decreased CBF globally in KO mice (range −31% to −41% decrease from control levels) with a lesser effect in WILD mice. It is concluded that MAOB may normally regulate CBF distribution and its response to blood PEA. PMID:10095040

An integrated scheme for the simultaneous determination of biogenic amines, precursor amino acids, and related metabolites by liquid chromatography with electrochemical detection.

PubMed

Oka, K; Kojima, K; Togari, A; Nagatsu, T; Kiss, B

1984-06-08

A new method using high-performance liquid chromatography with electrochemical detection (HPLC-ED) for the simultaneous determination of monoamines, their precursor amino acids, and related major metabolites in small samples of brain tissue weighing from 0.5 to 50 mg is described. The method is based on the preliminary isolation of monoamines (dopamine, norepinephrine, epinephrine, and serotonin), their precursor amino acids (tyrosine, 3,4-dihydroxyphenylalanine, tryptophan and 5-hydroxytryptophan), and their major metabolites (3-methoxytyramine, normetanephrine, 3,4-dihydroxyphenylacetic acid, homovanillic acid, vanillylmandelic acid, 3-methoxy-4-hydroxyphenylethyleneglycol, and 5-hydroxyindoleacetic acid) by chromatography on small columns of Amberlite CG-50 and Dowex 50W, and by ethyl acetate extraction. All the compounds in the four isolated fractions were measured by HPLC-ED on a reversed-phase column under four different conditions. The sensitivity was from 0.1 to 40 pmol, depending on the substances analysed. This newly established method was applied to the study of the effects of an aromatic L-amino acid decarboxylase inhibitor (NSD-1015) and a monoamine oxidase inhibitor (pargyline) on the levels of monoamines, their precursor amino acids and their major metabolites in brain regions of mice.

Laccase: microbial sources, production, purification, and potential biotechnological applications.

PubMed

Shraddha; Shekher, Ravi; Sehgal, Simran; Kamthania, Mohit; Kumar, Ajay

2011-01-01

Laccase belongs to the blue multicopper oxidases and participates in cross-linking of monomers, degradation of polymers, and ring cleavage of aromatic compounds. It is widely distributed in higher plants and fungi. It is present in Ascomycetes, Deuteromycetes and Basidiomycetes and abundant in lignin-degrading white-rot fungi. It is also used in the synthesis of organic substance, where typical substrates are amines and phenols, the reaction products are dimers and oligomers derived from the coupling of reactive radical intermediates. In the recent years, these enzymes have gained application in the field of textile, pulp and paper, and food industry. Recently, it is also used in the design of biosensors, biofuel cells, as a medical diagnostics tool and bioremediation agent to clean up herbicides, pesticides and certain explosives in soil. Laccases have received attention of researchers in the last few decades due to their ability to oxidize both phenolic and nonphenolic lignin-related compounds as well as highly recalcitrant environmental pollutants. It has been identified as the principal enzyme associated with cuticular hardening in insects. Two main forms have been found: laccase-1 and laccase-2. This paper reviews the occurrence, mode of action, general properties, production, applications, and immobilization of laccases within different industrial fields.

Iron-catalyzed synthesis of secondary amines: on the way to green reductive aminations.

PubMed

Stemmler, Tobias; Surkus, Annette-Enrika; Pohl, Marga-Martina; Junge, Kathrin; Beller, Matthias

2014-11-01

Amines represent important intermediates in chemical and biological processes. Herein, we describe the use of a nanostructured iron-based catalyst for the tandem reductive amination between nitroarenes and aldehydes using hydrogen as reductant. The nanostructured iron-catalyst is prepared by immobilization of an iron-phenanthroline complex onto a commercially available carbon support. In the reaction sequence a primary amine is formed in situ from the corresponding nitro compound. Reversible condensation with aldehydes forms the respective imines, which are finally reduced to the desired secondary amine. This synthesis of secondary amines is atom-economical and environmentally attractive using cheap and readily available organic compounds as starting materials. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mechanistic and Structural Analyses of the Roles of Active Site Residues in the Yeast Polyamine Oxidase Fms1: Characterization of the N195A and D94N Enzymes†

PubMed Central

Adachi, Mariya S.; Taylor, Alexander B.; Hart, P. John; Fitzpatrick, Paul F.

2012-01-01

The flavoprotein Fms1 from Saccharomyces cerevisiae catalyzes the oxidation of spermine in the biosynthetic pathway for pantothenic acid. The same reaction is catalyzed by the mammalian polyamine and spermine oxidases. The active site of Fms1 contains three amino acid residues positioned to interact with the polyamine substrate, His67, Asn195, and Asp94. These three residues form a hydrogen-bonding triad with Asn195 the central residue. Previous studies of the effects of mutating His67 are consistent with that residue being important both for interacting with the substrate and for maintaining the hydrogen bonds in the triad (Adachi, M. S., Taylor, A. B., Hart, P. J., and Fitzpatrick, P. F. (2012) Biochemistry 51, 4888-4897). The N195A and D94N enzymes have now been characterized to evaluate their roles in catalysis. Both mutations primarily affect the reductive half-reaction. With N1-acetylspermine as substrate, the rate constant for flavin reduction decreases ~450-fold for both mutations; the effects with spermine as substrate are smaller, 20- to 40-fold. The kcat/Kamine and kcat pH profiles with N1acetylspermine are only slightly changed from the profiles for the wild-type enzyme, consistent with the pKa values arising from the amine substrate or product and not from active site residues. The structure of the N195A enzyme was determined at a resolution of 2.0 Å. The structure shows a molecule of tetraethylene glycol in the active site and establishes that the mutation has no effect on the protein structure. Overall, the results are consistent with the role of Asn195 and Asp94 being to properly position the polyamine substrate for oxidation. PMID:23034052

Molecular insights into human monoamine oxidase (MAO) inhibition by 1,4-naphthoquinone: evidences for menadione (vitamin K3) acting as a competitive and reversible inhibitor of MAO.

PubMed

Coelho Cerqueira, Eduardo; Netz, Paulo Augusto; Diniz, Cristiane; Petry do Canto, Vanessa; Follmer, Cristian

2011-12-15

Monoamine oxidase (MAO) catalyzes the oxidative deamination of biogenic and exogenous amines and its inhibitors have therapeutic value for several conditions including affective disorders, stroke, neurodegenerative diseases and aging. The discovery of 2,3,6-trimethyl-1,4-naphthoquinone (TMN) as a nonselective and reversible inhibitor of MAO, has suggested 1,4-naphthoquinone (1,4-NQ) as a potential scaffold for designing new MAO inhibitors. Combining molecular modeling tools and biochemical assays we evaluate the kinetic and molecular details of the inhibition of human MAO by 1,4-NQ, comparing it with TMN and menadione. Menadione (2-methyl-1,4-naphthoquinone) is a multitarget drug that acts as a precursor of vitamin K and an inducer of mitochondrial permeability transition. Herein we show that MAO-B was inhibited competitively by 1,4-NQ (K(i)=1.4 μM) whereas MAO-A was inhibited by non-competitive mechanism (K(i)=7.7 μM). Contrasting with TMN and 1,4-NQ, menadione exhibited a 60-fold selectivity for MAO-B (K(i)=0.4 μM) in comparison with MAO-A (K(i)=26 μM), which makes it as selective as rasagiline. Fluorescence and molecular modeling data indicated that these inhibitors interact with the flavin moiety at the active site of the enzyme. Additionally, docking studies suggest the phenyl side groups of Tyr407 and Tyr444 (for MAO-A) or Tyr398 and Tyr435 (for MAO-B) play an important role in the interaction of the enzyme with 1,4-NQ scaffold through forces of dispersion as verified for menadione, TMN and 1,4-NQ. Taken together, our findings reveal the molecular details of MAO inhibition by 1,4-NQ scaffold and show for the first time that menadione acts as a competitive and reversible inhibitor of human MAO. Copyright © 2011 Elsevier Ltd. All rights reserved.

Specific binding of [(18)F]fluoroethyl-harmol to monoamine oxidase A in rat brain cryostat sections, and compartmental analysis of binding in living brain.

PubMed

Maschauer, Simone; Haller, Adelina; Riss, Patrick J; Kuwert, Torsten; Prante, Olaf; Cumming, Paul

2015-12-01

We investigated [(18)F]fluoroethyl-harmol ([(18)F]FEH) as a reversible and selective ligand for positron emission tomography (PET) studies of monoamine oxidase A (MAO-A). Binding of [(18)F]FEH in rat brain cryostat sections indicated high affinity (KD = 3 nM), and density (Bmax; 600 pmol/g). The plasma free fraction was 45%, and untransformed parent constituted only 13% of plasma radioactivity at 10 min after injection. Compartmental analysis of PET recordings in pargyline-treated rats showed high permeability to brain (K1; 0.32 mL/g/min) and slow washout (k2; 0.024/min), resulting in a uniformly high equilibrium distribution volume (VD; 20 mL/g). Using this VD to estimate unbound ligand in brain of untreated rats, the binding potential ranged from 4.2 in cerebellum to 7.2 in thalamus. We also calculated maps of rats receiving [(18)F]FEH at a range of specific activities, and then estimated saturation binding parameters in the living brain. In thalamus, striatum and frontal cortex KD was globally close to 300 nM and Bmax was close to 1600 pmol/g; the 100-fold discrepancy in affinity suggests a very low free fraction for [(18)F]FEH in the living brain. Based on a synthesis of findings, we calculate the endogenous dopamine concentration to be 0.4 μM in the striatal compartment containing MAO-A, thus unlikely to exert competition against [(18)F]FEH binding in vivo. In summary, [(18)F]FEH has good properties for the detection of MAO-A in the rat brain by PET, and may present logistic advantages for clinical research at centers lacking a medical cyclotron. We made a compartmental analysis of [(18)F]fluoroethylharmol ([(18)F]FEH) binding to monoamine oxidase A (MAO-A) in living rat brain and estimated the saturation binding parameters from the binding potential (BPND). The Bmax was of comparable magnitude to that in vitro, but with apparent affinity (300 nM), it was 100-fold lower in vivo. PET imaging with [(18) F]FEH is well suited for quantitation of MAO-A in living brain. © 2015 International Society for Neurochemistry.

Chemoselective Installation of Amine Bonds on Proteins through Aza-Michael Ligation.

PubMed

Freedy, Allyson M; Matos, Maria J; Boutureira, Omar; Corzana, Francisco; Guerreiro, Ana; Akkapeddi, Padma; Somovilla, Víctor J; Rodrigues, Tiago; Nicholls, Karl; Xie, Bangwen; Jiménez-Osés, Gonzalo; Brindle, Kevin M; Neves, André A; Bernardes, Gonçalo J L

2017-12-20

Chemical modification of proteins is essential for a variety of important diagnostic and therapeutic applications. Many strategies developed to date lack chemo- and regioselectivity as well as result in non-native linkages that may suffer from instability in vivo and adversely affect the protein's structure and function. We describe here the reaction of N-nucleophiles with the amino acid dehydroalanine (Dha) in a protein context. When Dha is chemically installed in proteins, the addition of a wide-range N-nucleophiles enables the rapid formation of amine linkages (secondary and tertiary) in a chemoselective manner under mild, biocompatible conditions. These new linkages are stable at a wide range of pH values (pH 2.8 to 12.8), under reducing conditions (biological thiols such as glutathione) and in human plasma. This method is demonstrated for three proteins and is shown to be fully compatible with disulfide bridges, as evidenced by the selective modification of recombinant albumin that displays 17 structurally relevant disulfides. The practicability and utility of our approach is further demonstrated by the construction of a chemically modified C2A domain of Synaptotagmin-I protein that retains its ability to preferentially bind to apoptotic cells at a level comparable to the native protein. Importantly, the method was useful for building a homogeneous antibody-drug conjugate with a precise drug-to-antibody ratio of 2. The kinase inhibitor crizotinib was directly conjugated to Dha through its piperidine motif, and its antibody-mediated intracellular delivery results in 10-fold improvement of its cancer cell-killing efficacy. The simplicity and exquisite site-selectivity of the aza-Michael ligation described herein allows the construction of stable secondary and tertiary amine-linked protein conjugates without affecting the structure and function of biologically relevant proteins.

Combined analytical approaches to define biodistribution and biological activity of semi-synthetic berberrubine, the active metabolite of natural berberine.

PubMed

Porru, Emanuele; Franco, Placido; Calabria, Donato; Spinozzi, Silvia; Roberti, Marinella; Caliceti, Cristiana; Roda, Aldo

2018-06-01

Berberine (BBR) is a natural alkaloid obtained from Berberis species plants, known for its protective effects against several diseases. Among the primary BBR metabolites, berberrubine (M1) showed the highest plasma concentration but few and conflicting data are available regarding its concentration in biological fluids related to its new potential activity on vascular cells. A combined analytical approach was applied to study biodistribution of M1 in comparison with BBR. The optimization of sample clean-up combined with a fully validated HPLC-ESI-MS/MS tailored for M1 allows sufficient detectability and accuracy to be reached in the different studied organs even when administered at low dose, comparable to that assumed by human. A predictive human vascular endothelial cell-based assay to measure intracellular xanthine oxidase has been developed and applied to study unexplored activities of M1 alongside other common activities. Results showed that oral M1 treatment exhibits higher plasma levels than BBR, reaching maximum concentration 400-fold higher than BBR (204 vs 0.5 ng/mL); moreover, M1 exhibits higher concentrations than BBR also in all the biological compartments analyzed. Noteworthy, the two compounds follow two different excretion routes: M1 through urine, while BBR through feces. In vitro studies demonstrated that M1 inhibited intracellular xanthine oxidase activity, one of the major sources of reactive oxygen species in vasculature, with an IC50 = 9.90 ± 0.01 μg/mL and reduced the expression of the inflammatory marker ICAM-1. These peculiar characteristics allow new perspectives to be opened up for the direct use of M1 instead of BBR in endothelial dysfunction treatment.

Unconjugated Bilirubin Inhibits Proteolytic Cleavage of von Willebrand Factor by ADAMTS13 Protease

PubMed Central

Lu, Rui-Nan; Yang, Shangbin; Wu, Haifeng M.; Zheng, X. Long

2015-01-01

Summary Background Bilirubin is a yellow breakdown product of heme catabolism. Increased serum levels of unconjugated bilirubin are conditions commonly seen in premature neonates and adults with acute hemolysis including thrombotic microangiopathy. Previous studies have shown that unconjugated bilirubin lowers plasma ADAMTS13 activity, but the mechanism is not fully understood. Objectives The study is to determine whether unconjugated bilirubin directly inhibits the cleavage of von Willebrand factor (VWF) and its analogs by ADAMTS13. Methods Fluorogenic, SELDI-TOF mass spectrometric assay, and Western blotting analyses were employed to address this question. Results Unconjugated bilirubin inhibits the cleavage of F485-rVWF73-H, D633-rVWF73-H, and GST-rVWF71-11K by ADAMTS13 in a concentration-dependent manner with a half-maximal inhibitory concentration (IC50) of ~13 μM, ~70 μM, and ~17 μM, respectively. Unconjugated bilirubin also dose-dependently inhibits the cleavage of multimeric VWF by ADAMTS13 under denaturing conditions. The inhibitory activity of bilirubin on the cleavage of D633-rVWF73-H and multimeric VWF, but not F485-rVWF73-H, was eliminated after incubation with bilirubin oxidase that converts bilirubin to biliverdin. Furthermore, plasma ADAMTS13 activity in patients with hyperbilirubinemia is lower prior to than after treatment with bilirubin oxidase. Conclusions unconjugated bilirubin directly inhibits ADAMTS13’s ability to cleave both peptidyl and native VWF substrates in addition to its interference with certain fluorogenic assays. Our findings may help proper interpretation of ADAMTS13 results under pathological conditions. Whether elevated serum unconjugated bilirubin has an adverse effect in vivo remains to be determined in our future study. PMID:25782102

Effect of Febuxostat, a Xanthine Oxidase Inhibitor, on Cardiovascular Risk in Hyperuricemic Patients with Hypertension: A Prospective, Open-label, Pilot Study.

PubMed

Tani, Shigemasa; Nagao, Ken; Hirayama, Atsushi

2015-12-01

There is growing evidence of an association between high uric acid (UA) levels and cardiovascular disease (CVD). We hypothesized that febuxostat, a xanthine oxidase inhibitor, may be associated with suppressing the renin-angiotensin-aldosterone system (RAAS) and improving renal function in hyperurecemic patients with hypertension. We conducted a 6-month prospective study in which we randomized hypertensive hyperuricemic patients to either a febuxostat group (n = 30) or a control group (n = 30). The dose of febuxostat was adjusted to maintain the serum UA level at <6.0 mg/dL. In the febuxostat group, the plasma renin activity (PRA), plasma aldosterone concentration (PAC), and serum UA level significantly decreased by 33 % (p = 0.0012), 14 % (p = 0.001), and 29 % (p < 0.0001), respectively. The estimated glomerular filtration rate (eGFR) significantly increased by 5.5 % (p = 0.001). Similar changes were not observed in the control group. Furthermore, a significant correlation was observed between the percent changes in the serum UA levels and the percent changes in the PRA (r = 0.277, p = 0.033), PAC (r = 0.310, p = 0.016), serum blood urea nitrogen levels (r = 0.434, p = 0.0005), serum creatinine levels (r = 0.413, p = 0.002), and eGFR (r = -0.474, p = 0.0001). These results support the hypothesis that febuxostat might not only reduce serum UA levels but also suppress RAAS and improve renal function in hyperuricemic patients with hypertension, possibly leading to prevention of CVD.

Activation Mobilizes the Cholesterol in the Late Endosomes-Lysosomes of Niemann Pick Type C Cells

PubMed Central

Lange, Yvonne; Ye, Jin; Steck, Theodore L.

2012-01-01

A variety of intercalating amphipaths increase the chemical activity of plasma membrane cholesterol. To test whether intracellular cholesterol can be similarly activated, we examined NPC1 and NPC2 fibroblasts, since they accumulate large amounts of cholesterol in their late endosomes and lysosomes (LE/L). We gauged the mobility of intracellular sterol from its appearance at the surface of the intact cells, as determined by its susceptibility to cholesterol oxidase and its isotope exchange with extracellular 2-(hydroxypropyl)-β-cyclodextrin-cholesterol. The entire cytoplasmic cholesterol pool in these cells was mobile, exchanging with the plasma membrane with an apparent half-time of ∼3–4 hours, ∼4–5 times slower than that for wild type human fibroblasts (half-time ∼0.75 hours). The mobility of the intracellular cholesterol was increased by the membrane-intercalating amphipaths chlorpromazine and 1-octanol. Chlorpromazine also promoted the net transfer of LE/L cholesterol to serum and cyclodextrin. Surprisingly, the mobility of LE/L cholesterol was greatly stimulated by treating intact NPC cells with glutaraldehyde or formaldehyde. Similar effects were seen with wild type fibroblasts in which the LE/L cholesterol pool had been expanded using U18666A. We also showed that the cholesterol in the intracellular membranes of fixed wild-type fibroblasts was mobile; it was rapidly oxidized by cholesterol oxidase and was rapidly replenished by exogenous sterol. We conclude that a) the cholesterol in NPC cells can exit the LE/L (and the extensive membranous inclusions therein) over a few hours; b) this mobility is stimulated by the activation of the cholesterol with intercalating amphipaths; c) intracellular cholesterol is even more mobile in fixed cells; and d) amphipaths that activate cholesterol might be useful in treating NPC disease. PMID:22276143

Atmospheric chemistry and environmental impact of the use of amines in carbon capture and storage (CCS).

PubMed

Nielsen, Claus J; Herrmann, Hartmut; Weller, Christian

2012-10-07

This critical review addresses the atmospheric gas phase and aqueous phase amine chemistry that is relevant to potential emissions from amine-based carbon capture and storage (CCS). The focus is on amine, nitrosamine and nitramine degradation, and nitrosamine and nitramine formation processes. A comparison between the relative importance of the various atmospheric sinks for amines, nitrosamines and nitramines is presented.

High capacity immobilized amine sorbents

DOEpatents

Gray, McMahan L [Pittsburgh, PA; Champagne, Kenneth J [Fredericktown, PA; Soong, Yee [Monroeville, PA; Filburn, Thomas [Granby, CT

2007-10-30

A method is provided for making low-cost CO.sub.2 sorbents that can be used in large-scale gas-solid processes. The improved method entails treating an amine to increase the number of secondary amine groups and impregnating the amine in a porous solid support. The method increases the CO.sub.2 capture capacity and decreases the cost of utilizing an amine-enriched solid sorbent in CO.sub.2 capture systems.

Photoredox-catalyzed Direct Reductive Amination of Aldehydes without an External Hydrogen/Hydride Source.

PubMed

Alam, Rauful; Molander, Gary A

2018-05-04

The direct reductive amination of aromatic aldehydes has been realized using a photocatalyst under visible light irradiation. The single electron oxidation of an in situ formed aminal species generates the putative α-amino radical that eventually delivers the reductive amination product. This method is operationally simple, highly selective, and functional group tolerant, which allows the direct synthesis of benzylic amines by a unique mechanistic pathway.

Potential occupational risk of amines in carbon capture for power generation.

PubMed

Gentry, P Robinan; House-Knight, Tamara; Harris, Angela; Greene, Tracy; Campleman, Sharan

2014-08-01

While CO2 capture and storage (CCS) technology has been well studied in terms of its efficacy and cost of implementation, there is limited available data concerning the potential for occupational exposure to amines, mixtures of amines, or degradation of by-products from the CCS process. This paper is a critical review of the available data concerning the potential effects of amines and CCS-degradation by-products. A comprehensive review of the occupational health and safety issues associated with exposure to amines and amine by-products at CCS facilities was performed, along with a review of the regulatory status and guidelines of amines, by-products, and CCS process vapor mixtures. There are no specific guidelines or regulations regarding permissible levels of exposure via air for amines and degradation products that could form atmospheric oxidation of amines released from post-combustion CO2 capture plants. While there has been a worldwide effort to develop legal and regulatory frameworks for CCS, none are directly related to occupational exposures. By-products of alkanolamine degradation may pose the most significant health hazard to workers in CCS facilities, with several aldehydes, amides, nitramines, and nitrosamines classified as either known or potential/possible human carcinogens. The absence of large-scale CCS facilities; absence and unreliability of reported data in the literature from pilot facilities; and proprietary amine blends make it difficult to estimate potential amine exposures and predict formation and exposure to degradation products.

Introducing inducible fluorescent split cholesterol oxidase to mammalian cells.

PubMed

Chernov, Konstantin G; Neuvonen, Maarit; Brock, Ivonne; Ikonen, Elina; Verkhusha, Vladislav V

2017-05-26

Cholesterol oxidase (COase) is a bacterial enzyme catalyzing the first step in the biodegradation of cholesterol. COase is an important biotechnological tool for clinical diagnostics and production of steroid drugs and insecticides. It is also used for tracking intracellular cholesterol; however, its utility is limited by the lack of an efficient temporal control of its activity. To overcome this we have developed a regulatable fragment complementation system for COase cloned from Chromobacterium sp. The enzyme was split into two moieties that were fused to FKBP (FK506-binding protein) and FRB (rapamycin-binding domain) pair and split GFP fragments. The addition of rapamycin reconstituted a fluorescent enzyme, termed split GFP-COase, the fluorescence level of which correlated with its oxidation activity. A rapid decrease of cellular cholesterol induced by intracellular expression of the split GFP-COase promoted the dissociation of a cholesterol biosensor D4H from the plasma membrane. The process was reversible as upon rapamycin removal, the split GFP-COase fluorescence was lost, and cellular cholesterol levels returned to normal. These data demonstrate that the split GFP-COase provides a novel tool to manipulate cholesterol in mammalian cells. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

40 CFR 721.7285 - Amines, N-cocoalkyltrimethylenedi-, citrates.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Amines, N-cocoalkyltrimethylenedi... Specific Chemical Substances § 721.7285 Amines, N-cocoalkyltrimethylenedi-, citrates. (a) Chemical substances and significant new uses subject to reporting. (1) The chemical substance identified as amines, N...

40 CFR 721.7286 - Amines, N-tallowalkyltripropylenetetra-, citrates.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Amines, N-tallowalkyltripropylenetetra... Specific Chemical Substances § 721.7286 Amines, N-tallowalkyltripropylenetetra-, citrates. (a) Chemical substances and significant new uses subject to reporting. (1) The chemical substance identified as amines, N...

40 CFR 721.7285 - Amines, N-cocoalkyltrimethylenedi-, citrates.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Amines, N-cocoalkyltrimethylenedi... Specific Chemical Substances § 721.7285 Amines, N-cocoalkyltrimethylenedi-, citrates. (a) Chemical substances and significant new uses subject to reporting. (1) The chemical substance identified as amines, N...

40 CFR 721.7286 - Amines, N-tallowalkyltripropylenetetra-, citrates.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Amines, N-tallowalkyltripropylenetetra... Specific Chemical Substances § 721.7286 Amines, N-tallowalkyltripropylenetetra-, citrates. (a) Chemical substances and significant new uses subject to reporting. (1) The chemical substance identified as amines, N...

Carbon Dioxide-Mediated C(sp3)-H Arylation of Amine Substrates.

PubMed

Kapoor, Mohit; Liu, Daniel; Young, Michael C

2018-05-25

Elaborating amines via C-H functionalization has been an important area of research over the past decade but has generally relied on an added directing group or sterically hindered amine approach. Since free-amine-directed C(sp 3 )-H activation is still primarily limited to cyclization reactions and to improve the sustainability and reaction scope of amine-based C-H activation, we present a strategy using CO 2 in the form of dry ice that facilitates intermolecular C-H arylation. This methodology has been used to enable an operationally simple procedure whereby 1° and 2° aliphatic amines can be arylated selectively at their γ-C-H positions. In addition to potentially serving as a directing group, CO 2 has also been demonstrated to curtail the oxidation of sensitive amine substrates.

A general catalytic β-C-H carbonylation of aliphatic amines to β-lactams.

PubMed

Willcox, Darren; Chappell, Ben G N; Hogg, Kirsten F; Calleja, Jonas; Smalley, Adam P; Gaunt, Matthew J

2016-11-18

Methods for the synthesis and functionalization of amines are intrinsically important to a variety of chemical applications. We present a general carbon-hydrogen bond activation process that combines readily available aliphatic amines and the feedstock gas carbon monoxide to form synthetically versatile value-added amide products. The operationally straightforward palladium-catalyzed process exploits a distinct reaction pathway, wherein a sterically hindered carboxylate ligand orchestrates an amine attack on a palladium anhydride to transform aliphatic amines into β-lactams. The reaction is successful with a wide range of secondary amines and can be used as a late-stage functionalization tactic to deliver advanced, highly functionalized amine products of utility for pharmaceutical research and other areas. Copyright © 2016, American Association for the Advancement of Science.

Conversion of alcohols to enantiopure amines through dual enzyme hydrogen-borrowing cascades

PubMed Central

Mutti, Francesco G.; Knaus, Tanja; Scrutton, Nigel S.; Breuer, Michael; Turner, Nicholas J.

2016-01-01

α-Chiral amines are key intermediates for the synthesis of a plethora of chemical compounds on industrial scale. Here we present a biocatalytic hydrogen-borrowing amination of primary and secondary alcohols that allows for the efficient and environmentally benign production of enantiopure amines. The method relies on the combination of an alcohol dehydrogenase (ADHs from Aromatoleum sp., Lactobacillus sp. and Bacillus sp.) enzyme operating in tandem with an amine dehydrogenase (AmDHs engineered from Bacillus sp.) to aminate a structurally diverse range of aromatic and aliphatic alcohols (up to 96% conversion and 99% enantiomeric excess). Furthermore, primary alcohols are aminated with high conversion (up to 99%). This redox self-sufficient network possesses high atom efficiency, sourcing nitrogen from ammonium and generating water as the sole by-product. PMID:26404833

A novel assay to measure tertiary and quaternary amines in wastewater: An indicator for NDMA wastewater precursors.

PubMed

Woods-Chabane, Gwen C; Glover, Caitlin M; Marti, Erica J; Dickenson, Eric R V

2017-07-01

This study examined the potential of using a novel bulk amine assay as an approximation for the tertiary and quaternary amine load in wastewaters and surface water samples, and this approximation was compared to N-nitrosodimethylamine (NDMA) formation potential using chloramines. An existing colorimetric method was examined and optimized for the detection of amines in environmental water samples. The method consists of liquid-liquid extraction followed by a catalyzed reaction to form a yet-undefined product that is known to be both a strong chromophore and fluorophore. Previous work verified that this reaction was effectively catalyzed by a number of compounds containing tertiary and quaternary amine moieties. Many tertiary and quaternary compounds are also efficient producers of NDMA under chloramination conditions, and a linear correlation was consequently derived from the bulk amine signals vs. NDMA formation potential in various wastewater samples (R 2  = 0.74; n = 24; p-value < 0.05). The results provide evidence that approximately 2% of the tertiary and quaternary amines measured can form NDMA and an estimated 0.01-1.3% of nitrogen in dissolved organic nitrogen originates from these bulk amines. The normalization of NDMA concentration by the amine measurement revealed that ozone effectively destroyed those tertiary and quaternary amine structures more likely to form NDMA in treated wastewater samples. This bulk amine assay illustrates that proxy measurements of tertiary and quaternary amines can be linked to the NDMA formation potential of a given sample, and this approach may prove useful as a characterizing tool for NDMA precursors in wastewater. Copyright © 2017 Elsevier Ltd. All rights reserved.

Soda-amine pulping : reaction of amines with free phenolic [beta]-[omicron]-4 ethers

Treesearch

John R. Obst

1981-01-01

The quinone methide from guaiacylglycol-ß-guaiacyl ether underwent nucleophilic addition to the a-carbon with primary and secondary amines at 40°C. At pulping temperature, 170°C, only the primary amine adduct was detected. The quinone methide from guaiacylglycerol-ß-guaiacyl ether gave analogous adducts at 40°C, but no quinone methide-amine adducts were detected at 170...

Aminative umpolung of aldehydes to α-amino anion equivalents for Pd-catalyzed allylation: an efficient synthesis of homoallylic amines.

PubMed

Ding, Lei; Chen, Jing; Hu, Yifan; Xu, Juan; Gong, Xing; Xu, Dongfang; Zhao, Baoguo; Li, Hexing

2014-02-07

An attractive strategy for generation of α-amino anions from aldehydes with applications in synthesis of homoallylic amines is described. Aromatic aldehydes can be converted to α-amino anion equivalents via amination with 2,2-diphenylglycine and subsequent decarboxylation. The in situ generated α-imino anions are highly reactive for Pd-catalyzed allylation, forming the corresponding homoallylic amines in high yields with excellent regioselectivity.

Monocyte and macrophage-targeted NADPH oxidase mediates antifungal host defense and regulation of acute inflammation in mice

PubMed Central

Grimm, Melissa J.; Vethanayagam, R. Robert; Almyroudis, Nikolaos G.; Dennis, Carly G.; Khan, A. Nazmul H.; D’Auria, Anthony; Singel, Kelly L.; Davidson, Bruce A.; Knight, Paul R.; Blackwell, Timothy S.; Hohl, Tobias M.; Mansour, Michael K.; Vyas, Jatin M.; Röhm, Marc; Urban, Constantin F.; Kelkka, Tiina; Holmdahl, Rikard; Segal, Brahm H.

2013-01-01

Chronic granulomatous disease, an inherited disorder of the NADPH oxidase in which phagocytes are defective in the generation of superoxide anion and downstream reactive oxidant species, is characterized by severe bacterial and fungal infections and excessive inflammation. Although NADPH oxidase isoforms exist in several lineages, reactive oxidant generation is greatest in neutrophils, where NADPH oxidase has been deemed vital for pathogen killing. In contrast, the function and importance of NADPH oxidase in macrophages are less clear. Therefore, we evaluated susceptibility to pulmonary aspergillosis in globally NADPH oxidase-deficient mice versus transgenic mice with monocyte/macrophage-targeted NADPH oxidase activity. We found that the lethal inoculum was more than 100-fold greater in transgenic versus globally NADPH oxidase-deficient mice. Consistent with these in vivo results, NADPH oxidase in mouse alveolar macrophages limited germination of phagocytosed Aspergillus fumigatus spores. Finally, globally NADPH oxidase-deficient mice developed exuberant neutrophilic lung inflammation and pro-inflammatory cytokine responses to zymosan, a fungal cell wall-derived product composed principally of particulate beta-glucans, whereas inflammation in transgenic and wildtype mice was mild and transient. Together, our studies identify a central role for monocyte/macrophage NADPH oxidase in controlling fungal infection and in limiting acute lung inflammation. PMID:23509361

[Respiratory oxidases: the enzymes which use most of the oxygen which living things breathe].

PubMed

Toledo-Cuevas, E M

1997-01-01

The respiratory oxidases are the last enzymes of the aerobic respiratory chain. They catalize the reduction of molecular oxygen to water, with generation of an electrochemical gradient useful for the energy demanding cellular processes. Most of the oxidases belong to the heme-copper superfamily. They possess a heme-copper center, constituted of a high spin heme and a CuB center, where the reduction of oxygen takes place and probably where the link to proton pumping is located. The superfamily is divided in two classes: the quinol- and the cytochrome c-oxidases. The latter are divided in the aa3 and the cbb3-type cytochrome c oxidases. The main difference between quinol- and the aa3-type cytochrome c-oxidases is the CuA center, which is absent in the quinol oxidases. The cbb3-type cytochrome oxidases have the binuclear center, but lack the CuA center. They also does not have the classical subunits II and III. These differences seem not to affect the oxygen reduction or the proton pumping. Probably the oxidases have evolved from some denitrification enzymes and prior the photosynthetic process. Also is possible that the cbb3-type cytochrome oxidases or others very similar have been the first oxidases to appear.

40 CFR 721.1625 - Alkylbenzene sulfonate, amine salt.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Alkylbenzene sulfonate, amine salt... Substances § 721.1625 Alkylbenzene sulfonate, amine salt. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified generically as alkylbenzene sulfonate, amine...

40 CFR 721.1625 - Alkylbenzene sulfonate, amine salt.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Alkylbenzene sulfonate, amine salt... Substances § 721.1625 Alkylbenzene sulfonate, amine salt. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified generically as alkylbenzene sulfonate, amine...