scholarly journals Jacobsen Catalyst as a Cytochrome P450 Biomimetic Model for the Metabolism of Monensin A

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Bruno Alves Rocha ◽  
Anderson Rodrigo Moraes de Oliveira ◽  
Murilo Pazin ◽  
Daniel Junqueira Dorta ◽  
Andresa Piacezzi Nascimento Rodrigues ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Liver Mitochondria ◽  
Biological Properties ◽  
Model Systems ◽  
Mass Spectrometry Analysis ◽  
Rat Liver Mitochondria ◽  
Model Assessment ◽  
Spectrometry Analysis ◽  
Monensin A

Monensin A is a commercially important natural product isolated fromStreptomyces cinnamonensinsthat is primarily employed to treat coccidiosis. Monensin A selectively complexes and transports sodium cations across lipid membranes and displays a variety of biological properties. In this study, we evaluated the Jacobsen catalyst as a cytochrome P450 biomimetic model to investigate the oxidation of monensin A. Mass spectrometry analysis of the products from these model systems revealed the formation of two products: 3-O-demethyl monensin A and 12-hydroxy monensin A, which are the same ones found inin vivomodels. Monensin A and products obtained in biomimetic model were tested in a mitochondrial toxicity model assessment and an antimicrobial bioassay againstStaphylococcus aureus, S. aureusmethicillin-resistant,Staphylococcus epidermidis, Pseudomonas aeruginosa,andEscherichia coli.Our results demonstrated the toxicological effects of monensin A in isolated rat liver mitochondria but not its products, showing that the metabolism of monensin A is a detoxification metabolism. In addition, the antimicrobial bioassay showed that monensin A and its products possessed activity against Gram-positive microorganisms but not for Gram-negative microorganisms. The results revealed the potential of application of this biomimetic chemical model in the synthesis of drug metabolites, providing metabolites for biological tests and other purposes.

scholarly journals N-Terminomics Strategies for Protease Substrates Profiling

Molecules ◽  
2021 ◽  
Vol 26 (15) ◽  
pp. 4699
Author(s):  
Mubashir Mintoo ◽  
Amritangshu Chakravarty ◽  
Ronak Tilvawala
Keyword(s):  
Mass Spectrometry ◽  
Protease Activity ◽  
Viral Infections ◽  
Mass Spectrometry Analysis ◽  
Post Translational Modification ◽  
Spectrometry Analysis ◽  
Physiological Conditions ◽  
Inflammatory Conditions ◽  
Biological Mixtures

Proteases play a central role in various biochemical pathways catalyzing and regulating key biological events. Proteases catalyze an irreversible post-translational modification called proteolysis by hydrolyzing peptide bonds in proteins. Given the destructive potential of proteolysis, protease activity is tightly regulated. Dysregulation of protease activity has been reported in numerous disease conditions, including cancers, neurodegenerative diseases, inflammatory conditions, cardiovascular diseases, and viral infections. The proteolytic profile of a cell, tissue, or organ is governed by protease activation, activity, and substrate specificity. Thus, identifying protease substrates and proteolytic events under physiological conditions can provide crucial information about how the change in protease regulation can alter the cellular proteolytic landscape. In recent years, mass spectrometry-based techniques called N-terminomics have become instrumental in identifying protease substrates from complex biological mixtures. N-terminomics employs the labeling and enrichment of native and neo-N-termini peptides, generated upon proteolysis followed by mass spectrometry analysis allowing protease substrate profiling directly from biological samples. In this review, we provide a brief overview of N-terminomics techniques, focusing on their strengths, weaknesses, limitations, and providing specific examples where they were successfully employed to identify protease substrates in vivo and under physiological conditions. In addition, we explore the current trends in the protease field and the potential for future developments.

scholarly journals Phytotoxicity of Essential Oils on Selected Weeds: Potential Hazard on Food Crops

Plants ◽  
2018 ◽  
Vol 7 (4) ◽  
pp. 79 ◽  
Author(s):  
María Ibáñez ◽  
María Blázquez
Keyword(s):  
Seed Germination ◽  
Essential Oil ◽  
Essential Oils ◽  
Mass Spectrometry Analysis ◽  
Gas Chromatography Mass Spectrometry ◽  
Food Crops ◽  
Potential Hazard ◽  
Spectrometry Analysis

The chemical composition of winter savory, peppermint, and anise essential oils, and in vitro and in vivo phytotoxic activity against weeds (Portulaca oleracea, Lolium multiflorum, and Echinochloa crus-galli) and food crops (maize, rice, and tomato), have been studied. Sixty-four compounds accounting for between 97.67–99.66% of the total essential oils were identified by Gas Chromatography-Mass Spectrometry analysis. Winter savory with carvacrol (43.34%) and thymol (23.20%) as the main compounds produced a total inhibitory effect against the seed germination of tested weed. Menthol (48.23%), menthone (23.33%), and iso-menthone (16.33%) from peppermint only showed total seed germination inhibition on L. multiflorum, whereas no significant effects were observed with trans-anethole (99.46%) from anise at all concentrations (0.125–1 µL/mL). Low doses of peppermint essential oil could be used as a sustainable alternative to synthetic agrochemicals to control L. multiflorum. The results corroborate that in vivo assays with a commercial emulsifiable concentrate need higher doses of the essential oils to reproduce previous in vitro trials. The higher in vivo phytotoxicity of winter savory essential oil constitutes an eco-friendly and less pernicious alternative to weed control. It is possible to achieve a greater in vivo phytotoxicity if less active essential oil like peppermint is included with other active excipients.

scholarly journals The Endopolygalacturonase 1 from Botrytis cinerea Activates Grapevine Defense Reactions Unrelated to Its Enzymatic Activity

2003 ◽  
Vol 16 (6) ◽  
pp. 553-564 ◽  
Author(s):  
Benoît Poinssot ◽  
Elodie Vandelle ◽  
Marc Bentéjac ◽  
Marielle Adrian ◽  
Caroline Levis ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Calcium Influx ◽  
Active Oxygen Species ◽  
Degree Of Polymerization ◽  
Mass Spectrometry Analysis ◽  
Gene Transcript ◽  
Transcript Accumulation ◽  
Spectrometry Analysis ◽  
Defense Reactions

A purified glycoprotein from Botrytis cinerea(strain T4), identified as endopolygalacturonase 1 (T4BcPG1) by mass spectrometry analysis, has been shown to activate defense reactions in grapevine (Vitis vinifera cv. Gamay). These reactions include calcium influx, production of active oxygen species, activation of two mitogen-activated protein kinases, defense gene transcript accumulation, and phytoalexin production. Most of these defense reactions were also activated in grapevine in response to purified oligogalacturonides (OGA) with a degree of polymerization of 9 to 20. In vivo, these active OGA might be a part of the released products resulting from endopolygalacturonase activity on plant cell walls. Nevertheless, the intensity and kinetics of events triggered by OGA were very different when compared with T4BcPG1 effects. Moreover, chemical treatments of T4BcPG1 and desensitization assays have allowed us to discriminate enzymatic and elicitor activities, indicating that elicitor activity was not due to released oligogalacturonides. Thus, BcPG1 should be considered as both an avirulence and a virulence factor. The role of the secreted BcPG1 in the pathogenicity of Botrytis cinerea is discussed.

Corona protein impacts on alternating current biosusceptometry signal and circulation times of differently coated MnFe2O4 nanoparticles

Nanomedicine ◽  
2021 ◽  
Author(s):  
Andre Gonçalves Prospero ◽  
Lais Pereira Buranello ◽  
Carlos AH Fernandes ◽  
Lucilene Delazari dos Santos ◽  
Guilherme Soares ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum ◽  
Signal Intensity ◽  
Alternating Current ◽  
Circulation Time ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis

Background: We evaluated the impacts of corona protein (CP) formation on the alternating current biosusceptometry (ACB) signal intensity and in vivo circulation times of three differently coated magnetic nanoparticles (MNP): bare, citrate-coated and bovine serum albumin-coated MNPs. Methods: We employed the ACB system, gel electrophoresis and mass spectrometry analysis. Results: Higher CP formation led to a greater reduction in the in vitro ACB signal intensity and circulation time. We found fewer proteins forming the CP for the bovine serum albumin-coated MNPs, which presented the highest circulation time in vivo among the MNPs studied. Conclusion: These data showed better biocompatibility, stability and magnetic signal uniformity in biological media for bovine serum albumin-coated MNPs than for citrate-coated MNPs and bare MNPs.

Abstract 54: ML355 in Prevention of Thrombosis in vivo with Minimal Effects on Hemostasis

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Reheman Adili ◽  
Katherine Mast ◽  
Michael Holinstat
Keyword(s):  
Ex Vivo ◽  
Platelet Reactivity ◽  
Thrombus Formation ◽  
Flow Chamber ◽  
Human Platelets ◽  
Mass Spectrometry Analysis ◽  
Vessel Occlusion ◽  
Spectrometry Analysis ◽  
Thrombosis Model

12-lipoxygenase (12-LOX) has been demonstrated to regulate platelet function, hemostasis, and thrombosis ex vivo , supporting a key role for 12-LOX in regulation of in vivo thrombosis. While pharmacologically targeting 12-LOX in vivo has been a challenge to date, the recent development of the 12-LOX selective inhibitor, ML355, as an effective antiplatelet therapeutic in vivo was assessed. ML355 potently inhibited thrombin and other agonist-induced platelet aggregation ex vivo in washed human platelets and inhibited downstream oxylipin production of platelet 12-LOX as confirmed by Mass spectrometry analysis. Ex vivo flow chamber assays confirmed that human platelet adhesion and thrombus formation at arterial shear over collagen was attenuated in human whole blood treated with ML355 to a greater extent compared to aspirin. In vivo , PK assessment of ML355 showed reasonable 12-LOX plasma levels 12 hours following administration of ML355. FeCl 3 -induced injury of the mesenteric arterioles resulted in less stable thrombi in 12-LOX -/- mice and ML355-treated WT mice resulting in impairment of vessel occlusion. Additionally, ML355 dose-dependently inhibited laser-induced thrombus formation in the cremaster arteriole thrombosis model in WT, but not in 12-LOX -/- mice. Importantly, hemostatic plug formation and bleeding following treatment with ML355 were not affected in response to laser ablation on the saphenous vein or in a cremaster microvasculature laser-induced rupture model. Our data strongly supports 12-LOX as a key determinant of platelet reactivity in vivo and inhibition of platelet 12-LOX with ML355 may represent a new class of antiplatelet therapeutics.

scholarly journals Thiol Peroxidase Is an Important Component of Streptococcus pneumoniae in Oxygenated Environments

2012 ◽  
Vol 80 (12) ◽  
pp. 4333-4343 ◽  
Author(s):  
Barak Hajaj ◽  
Hasan Yesilkaya ◽  
Rachel Benisty ◽  
Maayan David ◽  
Peter W. Andrew ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Mass Spectrometry Analysis ◽  
Activity Levels ◽  
Content Type ◽  
Spectrometry Analysis ◽  
Cellular Processes ◽  
Thiol Peroxidase ◽  
Fine Tune

ABSTRACTStreptococcus pneumoniaeis an aerotolerant Gram-positive bacterium that causes an array of diseases, including pneumonia, otitis media, and meningitis. During aerobic growth,S. pneumoniaeproduces high levels of H2O2. SinceS. pneumoniaelacks catalase, the question of how it controls H2O2levels is of critical importance. Thepsalocus encodes an ABC Mn2+-permease complex (psaBCA) and a putative thiol peroxidase,tpxD. This study shows thattpxDencodes a functional thiol peroxidase involved in the adjustment of H2O2homeostasis in the cell. Kinetic experiments showed that recombinant TpxD removed H2O2efficiently. However,in vivoexperiments revealed that TpxD detoxifies only a fraction of the H2O2generated by the pneumococcus. Mass spectrometry analysis demonstrated that TpxD Cys58undergoes selective oxidationin vivo, under conditions where H2O2is formed, confirming the thiol peroxidase activity. Levels of TpxD expression and synthesisin vitrowere significantly increased in cells grown under aerobic versus anaerobic conditions. The challenge with D39 and TIGR4 with H2O2resulted intpxDupregulation, whilepsaBCAexpression was oppositely affected. However, the challenge of ΔtpxDmutants with H2O2did not affectpsaBCA, implying that TpxD is involved in the regulation of thepsaoperon, in addition to its scavenging activity. Virulence studies demonstrated a notable difference in the survival time of mice infected intranasally with D39 compared to that of mice infected intranasally with D39ΔtpxD. However, when bacteria were administered directly into the blood, this difference disappeared. The findings of this study suggest that TpxD constitutes a component of the organism's fundamental strategy to fine-tune cellular processes in response to H2O2.

scholarly journals Arachidonic Acid Inhibits Epithelial Na Channel Via Cytochrome P450 (CYP) Epoxygenase-dependent Metabolic Pathways

2004 ◽  
Vol 124 (6) ◽  
pp. 719-727 ◽  
Author(s):  
Yuan Wei ◽  
Dao-Hong Lin ◽  
Rowena Kemp ◽  
Ganesh S.S. Yaddanapudi ◽  
Alberto Nasjletti ◽  
...  
Keyword(s):  
Arachidonic Acid ◽  
Cytochrome P450 ◽  
Collecting Duct ◽  
Inhibitory Effect ◽  
Mass Spectrometry Analysis ◽  
Gas Chromatography Mass Spectrometry ◽  
Na Channel ◽  
Cortical Collecting Duct ◽  
Patch Clamp Technique ◽  
Spectrometry Analysis

We used the patch-clamp technique to study the effect of arachidonic acid (AA) on epithelial Na channels (ENaC) in the rat cortical collecting duct (CCD). Application of 10 μM AA decreased the ENaC activity defined by NPo from 1.0 to 0.1. The dose–response curve of the AA effect on ENaC shows that 2 μM AA inhibited the ENaC activity by 50%. The effect of AA on ENaC is specific because neither 5,8,11,14-eicosatetraynoic acid (ETYA), a nonmetabolized analogue of AA, nor 11,14,17-eicosatrienoic acid mimicked the inhibitory effect of AA on ENaC. Moreover, inhibition of either cyclooxygenase (COX) with indomethacin or cytochrome P450 (CYP) ω-hydroxylation with N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS) failed to abolish the effect of AA on ENaC. In contrast, the inhibitory effect of AA on ENaC was absent in the presence of N-methylsulfonyl-6-(propargyloxyphenyl)hexanamide (MS-PPOH), an agent that inhibits CYP-epoxygenase activity. The notion that the inhibitory effect of AA is mediated by CYP-epoxygenase–dependent metabolites is also supported by the observation that application of 200 nM 11,12-epoxyeicosatrienoic acid (EET) inhibited ENaC in the CCD. In contrast, addition of 5,6-, 8,9-, or 14,15-EET failed to decrease ENaC activity. Also, application of 11,12-EET can still reduce ENaC activity in the presence of MS-PPOH, suggesting that 11,12-EET is a mediator for the AA-induced inhibition of ENaC. Furthermore, gas chromatography mass spectrometry analysis detected the presence of 11,12-EET in the CCD and CYP2C23 is expressed in the principal cells of the CCD. We conclude that AA inhibits ENaC activity in the CCD and that the effect of AA is mediated by a CYP-epoxygenase–dependent metabolite, 11,12-EET.

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