Oxidation Of Ascorbate Research Articles

Novel application of cyclo(-Phe-Pro) in mitigating aluminum toxicity through oxidative stress alleviation in wheat roots

Microbial secondary metabolites are crucial in plant-microorganism interactions, regulating plant growth and stress responses. In this study, we found that cyclo(-Phe-Pro), a proline-based cyclic dipeptide secreted by many microorganisms, alleviated aluminum toxicity in wheat roots by increasing root growth, decreasing callose deposition, and decreasing Al accumulation. Cyclo(-Phe-Pro) also significantly reduced Al-induced reactive oxygen species (ROS) with H2O2, O2•-, and •OH levels decreasing by 19.1%, 42.8%, and 17.9% in root tips, thus protecting the plasma membrane from oxidative damage. Although Al stress increased the activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and ascorbate peroxidase (APX) in wheat roots, cyclo(-Phe-Pro) application reduced these enzyme activities. However, compared to the Al treatment, cyclo(-Phe-Pro) application increased DPPH and FRAP activities by 16.8% and 14.9%, indicating increased non-enzymatic antioxidant capacity in wheat roots. We observed that Al caused the oxidation of ascorbate (AsA) and glutathione (GSH) to dehydroascorbate (DHA) and glutathione disulfide (GSSG), respectively. Under Al stress, cyclo(-Phe-Pro) treatment maintained reduced AsA and GSH levels, as well as high AsA/DHA and GSH/GSSG redox pair ratios in wheat roots. High AsA/DHA and GSH/GSSG ratios can reduce Al toxicity by neutralizing free radicals and restoring redox homeostasis via antioxidant properties. These results suggest that cyclo(-Phe-Pro) maintains ASA- and GSH-dependent redox homeostasis to alleviate oxidative and Al stress in wheat roots. Findings of this study establishes a theoretical foundation for using microbial metabolites to mitigate Al toxicity in acidic soils, highlighting their potential in sustainable agriculture.

Potency of Red Guava Fruitghurt as an Antioxidant

Antioxidants are compounds needed by the body to protect body cells from damage caused by free radicals. Free radicals can damage macromolecules such as cell membrane lipids, DNA, and proteins, and cause cell oxidative stress. Red Guava Fruitghurt is a probiotic with the basic ingredients of fruit. The purpose of this study is to determine the effect of fruitghurt on the MDA levels in the blood of mice induced by CCl4. This was an experimental study conducted using mice (Mus musculus) which were divided into three treatment groups – the negative control group (that were not induced and were not given fruitghurt), the positive control group (that were induced with CCl4 but not given fruitghurt), and the treatment group (that were induced with CCl4 and given fruitghurt). The data obtained in the study were analyzed using the one-way ANOVA test, and the results showed significant differences between the treatment groups. Giving fruitghurt to the P1 group had a significant effect on reducing malondialdehyde levels compared to the positive control group, with a P-value of 0.03. Lactobacillus acidophilus bacteria found in this fruitghurt can reduce oxidative stress by inhibiting ascorbic acid oxidation, reducing activity and capturing radial superoxide anions, hydrogen peroxide, and free radicals, and inhibiting lipid oxidation. So this fruitghurt has antioxidant properties. This is also strengthened by the content of red guava which is rich in phenolics, flavonoids, and vitamin C that have good free radical scavenging activity, so guava fruit is a natural source of antioxidants. Keywords: antioxidants, fruitghurt, malon dialdehyde, probiotics, red guava

Fabrication of non-enzymatic and highly sensitive electrochemical ascorbic acid sensor based on GO/Ag/PMMA nanocomposites

In the present study, Graphene Oxide–Silver–Poly(methyl methacrylate) (GO/Ag/PMMA) nanocomposites were fabricated and investigated for Ascorbic Acid sensing without using any enzyme. First, the graphene oxide was prepared by improved Hummers method. The tri-phase nanocomposites were also prepared by sonication and solution casting method with the ratio of Graphene Oxide (1, 2, 3, 4, and 5 %). The composites were characterized by FTIR, X-ray Diffraction (XRD), TGA, and scanning electron microscopy (SEM) analysis. The electrochemical studies/characterizations of ascorbic acid oxidation were carried out in the acidic medium and the new sensor was found to be a better electrochemical ascorbic acid sensor. The fabricated sensor is highly sensitive to ascorbic acid and shows its sensitivity (778.532 µA mM−1 cm−2), it has a broad linear range (from 0.1 to 6 mM), it is selective, and it shows tolerance towards endogenous species such as glucose, uric acid, hydrogen peroxide, and dopamine.

A novel approach to study ascorbic acid oxidation using hexavalent chromium species—theoretical and practical perspectives of the use of potentiometric titration technique

AbstractThis paper highlights the advantages of using the potentiometric titration technique as a valuable tool for studying the oxidation reaction of ascorbic acid by hexavalent chromium species in aqueous solutions. Particular attention was paid to the method of determining diagnostic points allowing the quantification of ascorbic acid in the sample under study. Additionally, the influence of experimental conditions, specifically the pH of the system and the concentration of the reactants, on the type of hexavalent chromium species involved in the interaction with ascorbic acid was analyzed. It has been shown that the method presented provides a simple, cost-effective, and rapid tool that can be widely used for the direct determination of ascorbic acid.

Versatile Bovine Serum Albumin as Ingenious Electron Operator-Enhanced Photoelectrochemical Biosensing for Ultrasensitive Detection of miRNA.

Bovine serum albumin (BSA) has been widely used in biosensors as a blocking agent. Herein, conformist BSA was first exploited as an ingenious operator to enhance the photocurrent response of (2Z,2'Z)-2,2'-(1,4-phenylene)bis(3-(4-(bis(4-methoxyphenyl)amino)phenyl)acrylonitrile) (TPDCN)-based photoelectrochemical (PEC) platform via manipulating the electron transfer process of the detection system. Concretely, the presence of target molecules triggered catalytic hairpin assembly reaction and subsequently powered terminal deoxynucleotidyl transferase-mediated signal amplification to produce the AgNP@BSA-DNA dendrimer nanostructure. After being treated with HNO3, a large amount of BSA could be released from the dendrimer nanostructure. When they were transferred to the TPDCN-based PEC platform, the photocurrent response of the biosensor was largely enhanced because BSA can manipulate the electrons of TPDCN via a well-matched energy level to form a new electron transfer track. Meanwhile, tryptophan (Trp) in BSA could be oxidized to quinone Trp-O under photoirradiation, which can facilitate the oxidation of ascorbate and generate more H+ to promote the migration of photogenerated electrons. As a result, the proposed PEC biosensor exhibits excellent analytical performance for detection of miRNA-21 (as a model target) over a wide linear range of 0.01 to 10,000 pM with detection limit as low as 4.7 fM. Overall, this strategy provides a new perspective on constructing efficient PEC biosensors, which expands the potential applications in bioanalysis and clinical diagnosis.

Abstract Tu123: Redox Biology of the Mitochondrial Protein mitoNEET and Ascorbate

Background: It has been proposed that mitoNEET is an Fe-S transfer/repair enzyme, playing a role under conditions of oxidative stress. mitoNEET protects mitochondria against oxidative stress in cardiomyocytes. Loss of mitoNEET is linked with age-related heart failure. Oxidized mitoNEET, [2Fe-2S] 2+ , exports Fe-S cluster from mitochondria to cytosol. Redox control of mitochondrion is essential for the proper functioning of the organelle/mitochondria. However, the molecular mechanisms involved in the redox biology of mitoNEET is not fully studied. Hypothesis: Ascorbate regulates redox reactions and Fe-S cluster transport function of mitoNEET in the mitochondria. Aims: The objective of this work is to evaluate the redox reactions between mitoNEET and ascorbate. Methods: Room temperature and low temperature (77 K) electron paramagnetic resonance (EPR) spectroscopy techniques were used to measure the oxidation of ascorbate and reduced mitoNEET, respectively. Results: EPR spectroscopic study shows that mitoNEET oxidizes ascorbate to ascorbate radical. The formation of ascorbate radical increases with increasing concentration of ascorbate. The formation of ascorbate radical is decreased in the presence of menadione. Low temperature EPR study shows that one electron reduction of mitoNEET by ascorbate. Conclusion: This study suggest that ascorbate is a substrate/cofactor for mitoNEET. Ascorbate regulates the Fe-S cluster transport function of mitoNEET. Ascorbate plays an important role in the redox biology of mitoNEET under physiological and pathophysiological conditions.

Dielectric study and electrochemical detection of ascorbic acid using modified electrode treated with europium-doped tricalcium aluminate nanoparticles

The current study involved the dielectric, and sensor properties of trivalent europium (Eu3+) doped tricalcium aluminate (Ca3Al2O6) Nanoparticles. These samples were synthesized through the solution combustion method with urea (NH2CO·NH2) as the fuel. Structural analysis confirms that the samples exhibited a sustainable cubic phase with the Pm-3m space group. The irregular spherical morphology and agglomeration of synthesized particles were observed using a scanning electron microscope. The energy bandgap values were calculated using the Kubelka and Munk relation, which ranged from 5.14 to 5.27 eV. To examine the dielectric behavior at different frequencies, electrical impedance spectroscopy was performed which provided insights into the changes in electrical permittivity and revealed the contributions of both grains and grain boundaries in the samples. The synthesized material exhibited higher values of dielectric constant and loss at lower frequencies, which decreased significantly as the frequency increased. This suggests that the samples are suitable for applications in microwave devices. Also, the electrochemical studies demonstrate the sensor's rapid amperometric response to ascorbic acid oxidation. The current constructed sensor has a sensitivity of 0.0065 A for ascorbic acid and the electrode treated with Ca3-xAl2O6:xEu3+ (x = 0.05) is better able to detect the electrochemical behavior of substances like ascorbic acid.

The Angiotensin Metabolite His-Leu Is a Strong Copper Chelator Forming Highly Redox Active Species.

His-Leu is a hydrolytic byproduct of angiotensin metabolism, whose concentration in the bloodstream could be at least micromolar. This encouraged us to investigate its Cu(II) binding properties and the concomitant redox reactivity. The Cu(II) binding constants were derived from isothermal titration calorimetry and potentiometry, while identities and structures of complexes were obtained from ultraviolet-visible, circular dichroism, and room-temperature electronic paramagnetic resonance spectroscopies. Four types of Cu(II)/His-Leu complexes were detected. The histamine-like complexes prevail at low pH. At neutral and mildly alkaline pH and low Cu(II):His-Leu ratios, they are superseded by diglycine-like complexes involving the deprotonated peptide nitrogen. At His-Leu:Cu(II) ratios of ≥2, bis-complexes are formed instead. Above pH 10.5, a diglycine-like complex containing the equatorially coordinated hydroxyl group predominates at all ratios tested. Cu(II)/His-Leu complexes are also strongly redox active, as demonstrated by voltammetric studies and the ascorbate oxidation assay. Finally, numeric competition simulations with human serum albumin, glycyl-histydyl-lysine, and histidine revealed that His-Leu might be a part of the low-molecular weight Cu(II) pool in blood if its abundance is >10 μM. These results yield further questions, such as the biological relevance of ternary complexes containing His-Leu.

Kinetics of ascorbate and dithiothreitol oxidation by soluble copper, iron, and manganese, and 1,4-naphthoquinone: Influence of the species concentration and the type of fluid

An alternative metric to account for particulate matter (PM) composition-based toxicity is the ability of PM-species to generate reactive oxygen species (ROS) and deplete antioxidants, the so-called oxidative potential (OP). Acellular OP assays are the most used worldwide, mainly those based on ascorbic acid (AA) and dithiothreitol (DTT) depletion; OP values are calculated from AA/DTT concentration over time kinetic curves. Since a great variability in OP-DTT and OP-AA values can be found in the literature, the understanding of those factors affecting the kinetic rate of AA and DTT oxidation in the presence of PM-bound species will improve the interpretation of OP values. In this work, a kinetic study of the oxidation rate of AA and DTT driven by species usually found in PM (transition metals and naphthoquinone (NQ)) was carried out. In particular, the influence of the concentration of Cu(II), Fe(II), Fe(III), Mn(II), Mn(III), and 1,4-NQ, and the type of fluid used in the assay (phosphate buffer (PB), phosphate buffer saline (PBS) and artificial lysosomal fluid (ALF)) is analysed and discussed. The reaction orders with respect to the AA/DTT and the active compound, and the kinetic rate constants were also determined. The results show great variability in OP values among the studied species depending on the fluid used; the OP values were mostly higher in PB0.05 M, followed by PBS1x and ALF. Moreover, different species concentration-responses for OP-DTT/OP-AA were obtained. These differences were explained by the different reaction orders and kinetic rate constants obtained for each active compound in each fluid.

Time-Separated Pulse Release-Activation of an Enzyme from Alginate-Polyethylenimine Hydrogels Using Electrochemically Generated Local pH Changes.

β-Glucosidase (EC 3.2.1.21) from sweet almond was encapsulated into pH-responsive alginate-polyethylenimine (alginate-PEI) hydrogel. Then, electrochemically controlled cyclic local pH changes resulting from ascorbate oxidation (acidification) and oxygen reduction (basification) were used for the pulsatile release of the enzyme from the composite hydrogel. Activation of the enzyme was controlled by the very same pH changes used for β-glucosidase release, separating these two processes in time. Importantly, the activity of the enzyme, which had not been released yet, was inhibited due to the buffering effect of PEI present in the gel. Thus, only a portion of the released enzyme was activated. Both enzymatic activity and release were monitored by confocal fluorescence microscopy and regular fluorescent spectroscopy. Namely, commercially available very little or nonfluorescent substrate 4-methylumbelliferyl-β-d-glucopyranoside was hydrolyzed by β-glucosidase to produce a highly fluorescent product 4-methylumbelliferone during the activation phase. At the same time, labeling of the enzyme with rhodamine B isothiocyanate was used for release observation. The proposed work represents an interesting smart release-activation system with potential applications in biomedical field.

Quest for a stable Cu-ligand complex with a high catalytic activity to produce reactive oxygen species.

Metal ion-catalyzed overproduction of reactive oxygen species (ROS) is believed to contribute significantly to oxidative stress and be involved in several biological processes, from immune defense to development of diseases. Among the essential metal ions, copper is one of the most efficient catalysts in ROS production in the presence of O2 and a physiological reducing agent such as ascorbate. To control this chemistry, Cu ions are tightly coordinated to biomolecules. Free or loosely bound Cu ions are generally avoided to prevent their toxicity. In the present report, we aim to find stable Cu-ligand complexes (Cu-L) that can efficiently catalyze the production of ROS in the presence of ascorbate under aerobic conditions. Thermodynamic stability would be needed to avoid dissociation in the biological environment, and high ROS catalysis is of interest for applications as antimicrobial or anticancer agents. A series of Cu complexes with the well-known tripodal and tetradentate ligands containing a central amine linked to three pyridyl-alkyl arms of different lengths were investigated. Two of them with mixed arm length showed a higher catalytic activity in the oxidation of ascorbate and subsequent ROS production than Cu salts in buffer, which is an unprecedented result. Despite these high catalytic activities, no increased antimicrobial activity toward Escherichia coli or cytotoxicity against eukaryotic AGS cells in culture related to Cu-L-based ROS production could be observed. The potential reasons for discrepancy between in vitro and in cell data are discussed.

Synthesis, characterization and anticancer screening of novel phenylbenzylidene thiosemicarbazone derivatives

Ten derivatives of phenylbenzylidene thiosemicarbazones (1–10) were synthesized by multicomponent reaction between hydrazine hydrate, isothiocyanate and substituted aldehydes in the presence of the phase transfer reagent Triton-B and subsequently heating the mixture. The synthesized compounds 1, 2, 5, 8 demonstrated to act as highly potent anticancer agents. They showed anticancer activity by mobilizing cellular 59Fe, inhibiting cellular uptake of 59Fe from 59Fe2-Tf, and mediating the ascorbate oxidation. Further, the structure activity relationship revealed that the donor atom and its stabilization resulted in better anticancer activity. The softness of the chelating donor atoms N and S yielded redox active iron complexes, which easily mediated ascorbate oxidation. This study identified the selective and potential chelators, which may act as anticancer agents and require further in vivo screening in future.

Respiratory tract lining fluid copper content contributes to pulmonary oxidative stress in patients with systemic sclerosis

Background Systemic sclerosis (SSc) is an autoimmune disease characterized by fibrosis of the skin and internal organs, mostly affecting young and middle-aged women. Significant questions remain as to its pathogenesis, especially the triggers for the associated interstitial lung disease (SSc-ILD). We examined the extent to which SSc and SSc-ILD were related to oxidative stress and altered metal homeostasis at the air-lung interface. Methods In this case-control study, we recruited 20 SSc patients, of which 11 had SSc-ILD. Eighteen healthy individuals were recruited as age-matched healthy controls, for a total of 38 study participants. Low molecular weight antioxidants (ascorbate, urate and glutathione), metal transport and chelation proteins (transferrin and ferritin) and metals (Fe and Cu) concentrations, including a measure of the catalytically active metal pool, were determined in respiratory tract lining fluid (RTLF) collected by bronchoalveolar lavage from the SSc group and compared with healthy controls. Results In the SSc group, 14 individuals were of female sex (70%) and the median age was 57 years (range 35-75). We observed evidence of oxidative stress in the RTLFs of SSc patients, characterised by increased concentrations of glutathione disulphide (GSSG, P<0.01), dehydroascorbate (DHA, P<0.05) and urate (P<0.01). This was associated with elevated RTLF Fe (P=0.07) and Cu (P<0.001), and evidence of a catalytic metal pool, demonstrated by an enhanced rate of ascorbate oxidation in the recovered lavage fluid (p<0.01). Cu concentrations were significantly associated with the ascorbate depletion rate (r=0.76, P<0.001), and GSSG (r=0.38, P<0.05) and protein carbonyl (r=0.44, P<0.01) concentrations. Whilst these markers were all increased in SSc patients, we found no evidence for an association with SSc-ILD. Conclusions These data confirm the presence of oxidative stress in the airways of SSc patients and, for the first time, suggest that an underlying defect in metal homeostasis at the air-lung interface may play a role in disease progression.

A Facile Synthesis of Bimetallic Copper-Silver Nanocomposite and Their Application in Ascorbic Acid Detection

Background:: An important antioxidant, ascorbic acid, must be detected in several industrial samples collected from food, pharmaceuticals, and water treatment plants. Herein, we reported a method to produce a bimetallic copper-silver (Cu-Ag) nanocomposite and used it in the development of very sensitive and selective electrochemical sensor for the detection of ascorbic acid. Methods:: A simple chemistry concept was used during the synthesis process to reduce the cost while minimizing the use of dangerous chemicals and minimizing the environmental impact. The Strobilanthes kunthiana leaves extract effectively reduced the copper and silver ions, resulting in the creation of an extremely stable and evenly distributed Cu-Ag nanocomposite. Results:: As-prepared bimetallic Cu-Ag nanocomposite exhibited outstanding electrochemical activity against ascorbic acid oxidation. The nanocomposite was examined using field emission scanning electron microscopy (FE-SEM), energy dispersive spectroscopy (EDS), elemental mapping (EMap) and X-ray diffraction analysis (XRD) to ascertain its composition, structure, and stability. Using cyclic voltammetry (CV), the electrochemical performance of the nanocomposite and also the detection of ascorbic acid were carried out. The bimetallic Cu-Ag nanocomposite also exhibited better long-term stability and fouling resistance, making it appropriate for use in real-world applications and complex sample matrices. Conclusion:: The bimetallic Cu-Ag nanocomposite coated electrode was used to detect the concentration of ascorbic acid by amperometry. As a result, this study offered a simple chemical method for creating a bimetallic copper-silver nanocomposite with superior electrochemical qualities for the accurate detection of ascorbic acid. conclusion: Its potential use as an electrochemical sensor for the detection of ascorbic acid opens doors for a variety of industries, including biological diagnostics, judging the quality of food, and environmental monitoring. As a result, this study offers a green method for creating a bimetallic copper-silver nanocomposite with superior electrochemical qualities for the accurate detection of ascorbic acid. The created nanocomposite has a lot of potential for improving ascorbic acid detection methods while upholding sustainable material synthesis. other: NA

Электрохимические сенсоры на основе сеток из однослойных углеродных нанотрубок для вольтамперометрического определения аскорбиновой кислоты

Modern highly sensitive and selective sensors are able to determine biologically active substances, which makes this direction one of the most popular areas of analytical chemistry. The study featured the electrochemical properties of new fiber materials based on single-wall carbon nanotubes with prospects of using them in the voltammetry of ascorbic acid. 
 The authors developed a new technology to synthesize films from disordered single-wall carbon nanotubes by chemical vapor deposition. Fibers were produced from a solvent by wet-pulling of single-wall carbon nanotubes networks. Thin films of randomly oriented single-wall carbon nanotube bundles were deposited downstream of a floating aerosol CVD reactor, which included a high temperature furnace with a quartz tube. The synthesis of the single-wall carbon nanotube samples was performed at 825°C. Ethanol served as carbon source while ferrocene was used as catalyst precursor. The single-wall carbon nanotubes were collected on a nitrocellulose filter in the form of films with transmittances of 10% in the middle of the visible wavelength (550 nm). The method was optimized to involve air annealing at 300–320°C and a treatment with strong inorganic acids, i.e., HCl, HNO3 + H2SO4. The voltammetric curves recording included background electrolyte, scan rate, and preconditioning. These parameters were selected experimentally to obtain the maximal sensor response to ascorbic acid content. The anodic peak of ascorbic acid in the phosphate buffer electrolyte (pH 6.86) was observed at a potential of +0.2 V. The current and peak area of ascorbic acid oxidation depended neither on the time nor on the conditioning potential of the sensor. The linear dependences of these parameters on the concentration of ascorbic acid stayed within 50–500 μmol/L (8.8–90 mg/L) at a scan rate of 0.1 mV/s. The single-wall carbon nanotube microsensor had a length of 0.5 cm and an average width of 400 μm. Its sensitivity was two times as high as that of a disk glassy carbon electrode with a diameter of 5 mm. 
 The experimental sensors proved effective in determining ascorbic acid in food products, pharmaceuticals, and biological fluids.

A Simple Flow Injection System for Amperometric Detection of Ascorbic Acid Using Carbon Paste/Copper Schiff Base Composite Electrode

AbstractIn this paper, a simple, sensitive, and precise flow injection system was optimized and applied to ascorbic acid determination with a carbon paste electrode modified with the Schiff base copper complex. The modified sensor showed increased sensitivity toward ascorbic acid oxidation with a decrease of oxidation potential by 200 mV. Under optimized flow parameters, working potential, gasket thickness, sample and reaction coil volumes, and flow rate, the developed flow system gave a linear response from 5 to 75 μmol/L of ascorbic acid. The calculated limit of detection was 4.5 μmol/L, calculated as 3σ/s which corresponds to the absolute value of 40 ng for the 50 μL sample coil. The precision for six consecutive injections of 5 and 75 μmol/L ascorbic acid solutions was determined as a relative standard deviation of 5.8 % and 1.5 %, respectively. The flow injection system enabled up to 120 analyses per hour. An optimized flow system was applied to the determination of ascorbic acid in dietary supplements, after simple preparation.

Ascorbate oxidation stimulates rice root growth via effects on auxin and abscisic acid levels

Ascorbate oxidation stimulates rice root growth via effects on auxin and abscisic acid levels

Electrochemical and Biocatalytic Signal-Controlled Payload Release from a Metal-Organic Framework.

A metal-organic framework (MOF), ZIF-8, which is stable at neutral and slightly basic pH values in aqueous solutions and destabilized/dissolved under acidic conditions, is loaded with a pH-insensitive fluorescent dye, rhodamine-B isothiocyanate, as a model payload species. Then, the MOF species are immobilized at an electrode surface. The local (interfacial) pH value is rapidly decreased by means of an electrochemically stimulated ascorbate oxidation at +0.4V (Ag/AgCl/KCl). Oxygen reduction upon switching the applied potential to -0.8V allows to return the local pH to the neutral/basic pH, then stopping rapidly the release process. The developed method allows electrochemical control over stimulated or inhibited payload release processes from the MOF. The pH variation proceeds in a thin film of the solution near the electrode surface. The switchable release process is realized in a buffer solution and undiluted human serum. As the second option, the pH decrease stimulating the release process is achieved upon an enzymatic reaction using esterase and ester substrate. This approach potentially allows the release activation controlled by numerous enzymes assembled in complex biocatalytic cascades. It is expected that related electrochemical or biocatalytic systems can represent novel signal-responding materials with switchable features for delivering (bio)molecules within biomedical applications.

Thylakoid Rhodanese-like Protein-Ferredoxin:NADP+ Oxidoreductase Interaction Is Integrated into Plant Redox Homeostasis System.

In vascular plants, the final photosynthetic electron transfer from ferredoxin (Fd) to NADP+ is catalyzed by the flavoenzyme ferredoxin:NADP+ oxidoreductase (FNR). FNR is recruited to thylakoid membranes via an integral membrane protein TROL (thylakoid rhodanese-like protein) and the membrane associated protein Tic62. We have previously demonstrated that the absence of TROL triggers a very efficient superoxide (O2•-) removal mechanism. The dynamic TROL-FNR interaction has been shown to be an apparently overlooked mechanism that maintains linear electron flow before alternative pathway(s) is(are) activated. In this work, we aimed to further test our hypothesis that the FNR-TROL pair could be the source element that triggers various downstream networks of chloroplast ROS scavenging. Tandem affinity purification followed by the MS analysis confirmed the TROL-FNR interaction and revealed possible interaction of TROL with the thylakoid form of the enzyme ascorbate peroxidase (tAPX), which catalyzes the H2O2-dependent oxidation of ascorbate and is, therefore, the crucial component of the redox homeostasis system in plants. Further, EPR analyses using superoxide spin trap DMPO showed that, in comparison with the wild type, plants overexpressing TROL (TROL OX) propagate more O2•- when exposed to high light stress. This indicates an increased sensitivity to oxidative stress in conditions when there is an excess of membrane-bound FNR and less free FNR is found in the stroma. Finally, immunohistochemical analyses of glutathione in different Arabidopsis leaf cell compartments showed highly elevated glutathione levels in TROL OX, indicating an increased demand for this ROS scavenger in these plants, likely needed to prevent the damage of important cellular components caused by reactive oxygen species.

Ascorbate oxidation driven by PM2.5-bound metal(loid)s extracted in an acidic simulated lung fluid in relation to their bioaccessibility

The oxidative potential (OP) is defined as the ability of inhaled PM components to catalytically/non-catalytically generate reactive oxygen species (ROS) and deplete lung antioxidants. Although several studies have measured the OP of particulate matter (PM OP) soluble components using different antioxidants under neutral pH conditions, few studies have measured PM OP with acidic lung fluids. This study provides new insights into the use of acidic rather than neutral fluids in OP assays. Thus, the first aim of this study was to clarify the effect of using an acidic lung fluid on ascorbic acid (AA) depletion. This was achieved by measuring the oxidative potential (OP-AA) of individual compounds known to catalyze the AA oxidation (CuSO4, CuCl2, and 1,4-NQ) in artificial lysosomal fluid (ALF, pH 4.5), a commonly used acidic simulated lung fluid, and in a neutral fluid (phosphate-buffered saline (PBS1x), pH 7.4). Our results from these individual compounds showed a significant decrease of OP-AA in the acidic fluid (ALF) with respect to the neutral fluid (PBS). Then, the second aim of this work was to investigate whether the OP-AA assay could be applied to PM2.5 samples extracted in acidic conditions. For this purpose, OP-AA and bioaccessible concentrations of metal(loid)s (V, Mn, Fe, Ni, Cu, Zn, As, Mo, Cd, Sb, and Pb) of PM2.5 samples collected in an urban-industrial area that were extracted in ALF were analyzed. The mean volume-normalized OP (OP-AAv) value was 0.10 ± 0.07 nmol min−1 m−3, clearly lower than the values found in the literature at neutral pH. OP-AAv values were highly correlated with the ALF-bioaccessible concentration of most of the studied metal(loid)s, mainly with Cu and Fe.