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Journal ArticleDOI

Measuring reactive oxygen and nitrogen species with fluorescent probes: challenges and limitations.

TL;DR: A critical analysis of the challenges and limitations of the most widely used fluorescent probes for detecting and measuring reactive oxygen and nitrogen species and proposed guidelines that will help present and future researchers with regard to the optimal use of selected fluorescent probes and interpretation of results are presented.
About: This article is published in Free Radical Biology and Medicine.The article was published on 2012-01-01 and is currently open access. It has received 1423 citations till now.
Citations
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Journal ArticleDOI
TL;DR: This review covered ZnO-NPs antibacterial activity including testing methods, impact of UV illumination,ZnO particle properties (size, concentration, morphology, and defects), particle surface modification, and minimum inhibitory concentration.
Abstract: Antibacterial activity of zinc oxide nanoparticles (ZnO-NPs) has received significant interest worldwide particularly by the implementation of nanotechnology to synthesize particles in the nanometer region. Many microorganisms exist in the range from hundreds of nanometers to tens of micrometers. ZnO-NPs exhibit attractive antibacterial properties due to increased specific surface area as the reduced particle size leading to enhanced particle surface reactivity. ZnO is a bio-safe material that possesses photo-oxidizing and photocatalysis impacts on chemical and biological species. This review covered ZnO-NPs antibacterial activity including testing methods, impact of UV illumination, ZnO particle properties (size, concentration, morphology, and defects), particle surface modification, and minimum inhibitory concentration. Particular emphasize was given to bactericidal and bacteriostatic mechanisms with focus on generation of reactive oxygen species (ROS) including hydrogen peroxide (H2O2), OH− (hydroxyl radicals), and O2 −2 (peroxide). ROS has been a major factor for several mechanisms including cell wall damage due to ZnO-localized interaction, enhanced membrane permeability, internalization of NPs due to loss of proton motive force and uptake of toxic dissolved zinc ions. These have led to mitochondria weakness, intracellular outflow, and release in gene expression of oxidative stress which caused eventual cell growth inhibition and cell death. In some cases, enhanced antibacterial activity can be attributed to surface defects on ZnO abrasive surface texture. One functional application of the ZnO antibacterial bioactivity was discussed in food packaging industry where ZnO-NPs are used as an antibacterial agent toward foodborne diseases. Proper incorporation of ZnO-NPs into packaging materials can cause interaction with foodborne pathogens, thereby releasing NPs onto food surface where they come in contact with bad bacteria and cause the bacterial death and/or inhibition.

2,627 citations

Journal ArticleDOI
TL;DR: A review of the state-of-the-art of this multidisciplinary area and identifying the key research challenges is provided in this paper, where the developments in diagnostics, modeling and further extensions of cross section and reaction rate databases are discussed.
Abstract: Plasma–liquid interactions represent a growing interdisciplinary area of research involving plasma science, fluid dynamics, heat and mass transfer, photolysis, multiphase chemistry and aerosol science. This review provides an assessment of the state-of-the-art of this multidisciplinary area and identifies the key research challenges. The developments in diagnostics, modeling and further extensions of cross section and reaction rate databases that are necessary to address these challenges are discussed. The review focusses on non-equilibrium plasmas.

1,078 citations

Journal ArticleDOI
TL;DR: The physicochemical basis for mitochondrial accumulation of lipophilic cations, synthetic chemistry strategies to target compounds to mitochondria, mitochondrial probes, and sensors, and examples of mitochondrial targeting of bioactive compounds are described.
Abstract: Mitochondria are recognized as one of the most important targets for new drug design in cancer, cardiovascular, and neurological diseases. Currently, the most effective way to deliver drugs specifically to mitochondria is by covalent linking a lipophilic cation such as an alkyltriphenylphosphonium moiety to a pharmacophore of interest. Other delocalized lipophilic cations, such as rhodamine, natural and synthetic mitochondria-targeting peptides, and nanoparticle vehicles, have also been used for mitochondrial delivery of small molecules. Depending on the approach used, and the cell and mitochondrial membrane potentials, more than 1000-fold higher mitochondrial concentration can be achieved. Mitochondrial targeting has been developed to study mitochondrial physiology and dysfunction and the interaction between mitochondria and other subcellular organelles and for treatment of a variety of diseases such as neurodegeneration and cancer. In this Review, we discuss efforts to target small-molecule compounds to mitochondria for probing mitochondria function, as diagnostic tools and potential therapeutics. We describe the physicochemical basis for mitochondrial accumulation of lipophilic cations, synthetic chemistry strategies to target compounds to mitochondria, mitochondrial probes, and sensors, and examples of mitochondrial targeting of bioactive compounds. Finally, we review published attempts to apply mitochondria-targeted agents for the treatment of cancer and neurodegenerative diseases.

892 citations

Journal ArticleDOI
22 Oct 2015-Cell
TL;DR: This work reviews mitochondrial ROS-mediated signaling pathways with an emphasis on how they are involved in various basal and adaptive physiological responses that control organismal homeostasis.

826 citations


Cites background from "Measuring reactive oxygen and nitro..."

  • ...The use of commercially fluorescent ROS probes is widespread but oftenwithout the knowledge that these do not always readout specific ROS species faithfully and are prone to other confounding artifacts (Kalyanaraman et al., 2012)....

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Journal ArticleDOI
TL;DR: This article looks back to the antioxidant/free radical field in 1994 and discusses how it has progressed in the past 18 years and suggests that increasing endogenous antioxidant levels (e.g., by supplying "pro-oxidants") may be a better approach to therapeutics and disease prevention than consuming large doses of "dietary antioxidants."
Abstract: This article looks back to the antioxidant/free radical field in 1994 and discusses how it has progressed in the past 18 years. In some areas, there has been little change: the role of oxygen radicals and other reactive oxygen species (ROS) in the origin or progression of most human diseases remains uncertain, with cancer and neurodegenerative disease being likely exceptions. Even in diseases in which ROS are involved there has been little progress in developing effective antioxidant treatments. Mega-doses of dietary antioxidants have also generally failed to prevent human disease, in part because they do not decrease oxidative damage in vivo (as revealed by robust biomarkers). However, some strategies that are known to delay disease onset may act, at least in part, by decreasing oxidative damage levels. Nevertheless, far more is known today about endogenous antioxidant defenses and how they are regulated, which has led to a deeper understanding of how some ROS can act as signaling molecules. Increasing endogenous antioxidant levels (e.g., by supplying “pro-oxidants”) may be a better approach to therapeutics and disease prevention than consuming large doses of “dietary antioxidants.”

721 citations

References
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Book
13 Jun 1985
TL;DR: 1. Oxygen is a toxic gas - an introduction to oxygen toxicity and reactive species, and the chemistry of free radicals and related 'reactive species'
Abstract: 1. Oxygen is a toxic gas - an introductionto oxygen toxicity and reactive species 2. The chemistry of free radicals and related 'reactive species' 3. Antioxidant defences Endogenous and Diet Derived 4. Cellular responses to oxidative stress: adaptation, damage, repair, senescence and death 5. Measurement of reactive species 6. Reactive species can pose special problems needing special solutions. Some examples. 7. Reactive species can be useful some more examples 8. Reactive species can be poisonous: their role in toxicology 9. Reactive species and disease: fact, fiction or filibuster? 10. Ageing, nutrition, disease, and therapy: A role for antioxidants?

21,528 citations


Additional excerpts

  • ..., atherosclerosis, cancer, diabetes, neurodegeneration) [1]....

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Journal ArticleDOI
TL;DR: The pathways that regulate ROS homeostasis are crucial for mitigating the toxicity of ROS and provide strong evidence about specificity in ROS signalling.
Abstract: Reactive oxygen species (ROS) have been shown to be toxic but also function as signalling molecules. This biological paradox underlies mechanisms that are important for the integrity and fitness of living organisms and their ageing. The pathways that regulate ROS homeostasis are crucial for mitigating the toxicity of ROS and provide strong evidence about specificity in ROS signalling. By taking advantage of the chemistry of ROS, highly specific mechanisms have evolved that form the basis of oxidant scavenging and ROS signalling systems.

2,941 citations


"Measuring reactive oxygen and nitro..." refers background in this paper

  • ...At a molecular level, reactive oxygen and nitrogen species exhibit signaling and cellfunction-modifying roles [2]....

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Journal ArticleDOI
TL;DR: Data suggest that H2O2-Fe(2+)-derived oxidant is mainly responsible for the nonenzymatic oxidation of DCFH, which remains an attractive probe as an overall index of oxidative stress in toxicological phenomena.
Abstract: The use of dichlorofluorescin (DCFH) as a measure of reactive oxygen species was studied in aqueous media. Hydrogen peroxide oxidized DCFH to fluorescent dichlorofluorescein (DCF), and the oxidation was amplified by the addition of ferrous iron. Hydrogen peroxide-induced DCF formation in the presence of ferrous iron was completely inhibited by deferoxamine and partially inhibited by ethylenediaminetetraacetic acid, but was augmented by diethylenetriaminepentaacetic acid. Iron-peroxide-induced oxidation of DCFH was partially inhibited by catalase but not by horseradish peroxidase. Nonchelated iron-peroxide oxidation of DCFH was partially inhibited by several hydroxyl radical scavengers, but was independent of the scavenger concentration, and this suggests that free hydroxyl radical is not involved in the oxidation of DCFH in this system. Superoxide anion did not directly oxidize DCFH. Data suggest that H2O2-Fe(2+)-derived oxidant is mainly responsible for the nonenzymatic oxidation of DCFH. In addition, peroxidase alone and oxidants formed during the reduction of H2O2 by peroxidase oxidize DCFH. Since DCFH oxidation may be derived from several reactive intermediates, interpretation of specific reactive oxygen species involved in biological systems should be approached with caution. However, DCFH remains an attractive probe as an overall index of oxidative stress in toxicological phenomena.

2,465 citations


"Measuring reactive oxygen and nitro..." refers background in this paper

  • ...Previous studies implicated a role for redox-active iron in intracellular oxidation of DCFH to DCF [16], although the origin of the cellular iron was not known....

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Journal ArticleDOI
TL;DR: This review examines how target selectivity and antioxidant effectiveness vary for different oxidants and highlights areas where greater understanding is required on the fate of oxidants generated by cellular NADPH oxidases and on the identification of oxidant sensors in cell signaling.
Abstract: There is a vast literature on the generation and effects of reactive oxygen species in biological systems, both in relation to damage they cause and their involvement in cell regulatory and signaling pathways. The biological chemistry of different oxidants is becoming well understood, but it is often unclear how this translates into cellular mechanisms where redox changes have been demonstrated. This review addresses this gap. It examines how target selectivity and antioxidant effectiveness vary for different oxidants. Kinetic considerations of reactivity are used to assess likely targets in cells and how reactions might be influenced by restricted diffusion and compartmentalization. It also highlights areas where greater understanding is required on the fate of oxidants generated by cellular NADPH oxidases and on the identification of oxidant sensors in cell signaling.

2,011 citations


"Measuring reactive oxygen and nitro..." refers background in this paper

  • ...As pointed out in a recent review [3], reactive oxygen and nitrogen species are not single entities but represent a broad range of chemically distinct reactive species with diverse biological reactivities [3,4]....

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  • ...It is unlikely to compete with H2O2-detoxifying enzymes such as the peroxiredoxins, which react 7 orders of magnitude faster with H2O2[3,4]....

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Journal Article
TL;DR: The data indicate that the DCFH oxidation assay is quantitatively related to the oxidative metabolic burst of PMNL, and they strongly suggest that the reaction is mediated by H2O2 generated by the PMNL.
Abstract: We have developed a quantitative assay to monitor the oxidative burst (H2O2 production) of polymorphonuclear leukocytes (PMNL) using single cell analysis by flow cytometry, and have examined whether PMNL respond to membrane stimulation with an all-or-none oxidative burst. During incubation with normal neutrophils, dichlorofluorescin diacetate diffused into the cells, was hydrolyzed to 2',7'-dichlorofluorescin (DCFH) and was thereby trapped within the cells. The intracellular DCFH, a nonfluorescent fluorescein analogue, was oxidized to highly fluorescent 2',7'-dichlorofluorescein (DCF) by PMNL stimulated by phorbol myristate acetate (PMA). That the oxidative product was DCF was shown by excitation/emission spectra and by mass spectrometry of the product from PMA-stimulated PMNL. Normal resting and PMA-stimulated PMNL oxidized 6.9 +/- 0.7 and 160 +/- 13 attomoles DCF per cell, respectively, in 15 min. Absence of calcium and magnesium ions and/or addition of 2 mM EDTA did not inhibit DCF formation by PMNL stimulated by 100 ng/ml PMA. Since EDTA prevented aggregation of PMNL (even when stimulated by 100 ng/ml PMA), which would prevent accurate flow cytometric analysis, further experiments were performed with EDTA in the medium. A close correlation between average DCFH oxidation and hexose monophosphate shunt stimulation was demonstrated using cells from patients whose PMNL had oxidative metabolic defects of varying severity. Intracellular DCFH was also oxidized by reagent H2O2 or oxygen derivatives generated by glucose oxidase + glucose or by xanthine oxidase + acetaldehyde; DCFH oxidation by these systems was inhibited by catalase but unchanged by superoxide dismutase. The data indicate that the DCFH oxidation assay is quantitatively related to the oxidative metabolic burst of PMNL, and they strongly suggest that the reaction is mediated by H2O2 generated by the PMNL. Incubation of PMNL with varying concentrations of PMA caused graded responses by all PMNL present; i.e., 1 ng/ml PMA caused a mean response of 34% maximal with a single population of responding PMNL (rather than 66% resting and 34% fully stimulated as predicted by the all-or-none hypothesis). Thus, with these assay conditions, oxidative product formation by PMNL occurs as a graded response to membrane stimulation by PMA.

1,790 citations