Glutathionylation of the Active Site Cysteines of Peroxiredoxin 2 and Recycling by Glutaredoxin

Peroxiredoxin 2 (Prx2) is a thiol protein that functions as an antioxidant, regulator of cellular peroxide concentrations, and sensor of redox signals. Its redox cycle is widely accepted to involve oxidation by a peroxide and reduction by thioredoxin/thioredoxin reductase. Interactions of Prx2 with other thiols are not well characterized. Here we show that the active site Cys residues of Prx2 form stable mixed disulfides with glutathione (GSH). Glutathionylation was reversed by glutaredoxin 1 (Grx1), and GSH plus Grx1 was able to support the peroxidase activity of Prx2. Prx2 became glutathionylated when its disulfide was incubated with GSH and when the reduced protein was treated with H₂O₂ and GSH. The latter reaction occurred via the sulfenic acid, which reacted sufficiently rapidly (k = 500 m⁻¹ s⁻¹) for physiological concentrations of GSH to inhibit Prx disulfide formation and protect against hyperoxidation to the sulfinic acid. Glutathionylated Prx2 was detected in erythrocytes from Grx1 knock-out mice after peroxide challenge. We conclude that Prx2 glutathionylation is a favorable reaction that can occur in cells under oxidative stress and may have a role in redox signaling. GSH/Grx1 provide an alternative mechanism to thioredoxin and thioredoxin reductase for Prx2 recycling.     

Secretory expression and characterization of a novel peroxiredoxin for zearalenone detoxification in Saccharomyces cerevisiae

Zearalenone (ZEN) is a Fusarium mycotoxin, which is considered to be an oestrogenic endocrine disruptor found to cause severe morphological and functional disorders of reproductive organs in livestock. Increasing attention has been paid to the development of an effective strategy for ZEN decontamination. ZEN is oxidized into smaller estrogenic metabolites by a novel peroxiredoxin (Prx) isolated from Acinetobacter sp. SM04. The Prx coding gene was cloned in a secretory vector pYES2-alpha (pYα) with an alpha (α) signal peptide gene inserted into the multiple cloning site of pYES2. The recombinant Prx was secreted from Saccharomyces cerevisiae INVSc1 after inducing with 2% (w/v) galactose for 72 h, and was found to be nearly 20 kDa through 12% SDS-PAGE. The expressed amount of recombinant Prx was 0.24 mg/mL in the extracellular supernatant. Recombinant Prx showed a gradient increase at the beginning of ZEN degradation. The final ZEN degradation amount was 0.43 μg by one unit recombinant Prx after 12 h. Furthermore, the temperature, H₂O₂ concentration, and pH for highest peroxidase activity of recombinant Prx were 80 °C, 20 mM and 9.0, respectively. When compared with other peroxidases, the thermal stability and alkali resistance of recombinant Prx were much better. The results suggest that recombinant Prx is successfully expressed in S. cerevisiae.     

Effects of intraspecific competition on growth and photosynthesis of Atriplex prostrata

We investigated the effect of intraspecific competition on growth parameters and photosynthesis of the salt marsh species Atriplex prostrata Boucher in order to distinguish the effects of density-dependent growth inhibition from salt stress. High plant density caused a reduction of 30% in height, 82% in stem dry mass, 80% in leaf dry mass, and 95% in root dry mass. High density also induced a pronounced 72% reduction in leaf area, 29% decrease in length of mature internodes and 50% decline in net photosynthetic rate. The alteration of net photosynthesis paralleled growth inhibition, decreasing from 7.6 ± 0.9 μmol CO₂ m⁻² s⁻¹ at low density to 3.5 ± 0.4 μmol CO₂ m⁻² s⁻¹ at high density, indicating growth inhibition caused by intraspecific competition is mainly due to a decline in net photosynthesis rate. Plants grown at high density also exhibited a reduction in stomatal conductance from 0.7 ± 0.1 mol H₂O m⁻² s⁻¹ at low density to 0.3 ± 0.1 mol H₂O m⁻² s⁻¹ at high density and a reduction in transpiration rate from 6.0 ± 0.3 mmol H₂O m⁻² s⁻¹ at low density to 4.3 ± 0.3 mmol H₂O m⁻² s⁻¹ at high density. Biomass production was inhibited by an increase in plant density, which reduced the rate of photosynthesis, stomatal conductance and leaf area of plants.     

Interaction of adenanthin with glutathione and thiol enzymes: Selectivity for thioredoxin reductase and inhibition of peroxiredoxin recycling

The diterpenoid, adenanthin, represses tumor growth and prolongs survival in mouse promyelocytic leukemia models (Liu et al., Nat. Chem. Biol. 8, 486, 2012). It was proposed that this was done by inactivating peroxiredoxins (Prxs) 1 and 2 through the formation of an adduct specifically on the resolving Cys residue. We confirmed that adenanthin underwent Michael addition to isolated Prx2, thereby inhibiting oxidation to a disulfide-linked dimer. However, contrary to the original report, both the peroxidatic and the resolving Cys residues could be derivatized. Glutathione also formed an adenanthin adduct, reacting with a second-order rate constant of 25±5 M^–1 s^–1. With 50 µM adenanthin, the peroxidatic and resolving Cys of Prx2 reacted with half-times of 7 and 40 min, respectively, compared with 10 min for GSH. When erythrocytes or Jurkat T cells were treated with adenanthin, we saw no evidence for a reaction with Prxs 1 or 2. Instead, adenanthin caused time- and concentration-dependent loss of GSH followed by dimerization of the Prxs. Prxs undergo continuous oxidation in cells and are normally recycled by thioredoxin reductase and thioredoxin. Our results indicate that Prx reduction was inhibited. We observed rapid inhibition of purified thioredoxin reductase (half-time 5 min with 2 µM adenanthin) and in cells, thioredoxin reductase was much more sensitive than GSH and loss of both preceded accumulation of oxidized Prxs. Thus, adenanthin is not a specific Prx inhibitor, and its reported antitumor and anti-inflammatory effects are more likely to involve more general inhibition of thioredoxin and/or glutathione redox pathways.     

Kinetic analysis for suicide-substrate inactivation of microperoxidase-11: A modified model for bisubstrate enzymes in the presence of reversible inhibitors

Kinetics of microperoxidase-11 (MP-11) as a heme–peptide enzyme model in oxidation reaction of guaiacol (AH) by hydrogen peroxide was studied in the presence of amino acids, taking into account the inactivation of MP-11 during reaction by its suicide substrate, H₂O₂. Reliability of the kinetic equation was evaluated by non-linear mathematical fitting. Fitting of experimental data into a new integrated kinetic relation showed a close match between the kinetic model and the experimental data. Indeed, it was found that the mechanism of suicide-peroxide inactivation of MP-11 in the presence of amino acids is different from MP-11 and/or horseradish peroxidase. In this mechanism, amino acids compete with hydrogen peroxide for the sixth co-ordination position of iron atom in the heme group through a competitive inhibition mechanism.The proposed model can successfully determine the kinetic parameters including inactivation by hydrogen peroxide as well as the inhibitory rate constants by the amino acid inhibitor.Kinetic parameters of inactivation including the initial activity of MP-11, α₀, the apparent inactivation rate constant, k_i and the apparent inhibition rate constant for cysteine, k_I were obtained 0.282 ± 0.006 min^?1, 0.497 ± 0.013 min^?1 and 1.374 ± 0.007 min^?1 at [H₂O₂] = 1.0 mM, 27 °C, phosphate buffer 5.0 mM, pH 7.0. Results showed that inactivation and inhibition of microperoxidase as a peroxidase model enzyme occurred simultaneously even at low concentrations of hydrogen peroxide (0.4 mM). This kinetic analysis based on the suicide-substrate inactivation of microperoxidase-11, provides a tool and model for studying peroxidase models in the presence of reversible inhibitors. The introduced inhibition procedure can be used in designing activity tunable and specific protected enzyme models in the hidden and reversibly inhibited forms, which do not undergo inactivation.     

Amperometric hydrogen peroxide biosensor based on the immobilization of HRP on nano-Au/Thi/poly (p-aminobenzene sulfonic acid)-modified glassy carbon electrode

A novel hydrogen peroxide biosensor was fabricated for the determination of H₂O₂. The precursor film was first electropolymerized on the glassy carbon electrode with p-aminobenzene sulfonic acid (p-ABSA) by cyclic voltammetry (CV). Then thionine (Thi) was adsorbed to the film to form a composite membrane, which yielded an interface containing amine groups to assemble gold nanoparticles (nano-Au) layer for immobilization of horseradish peroxidase (HRP). The electrochemical characteristics of the biosensor were studied by CV and chronoamperometry. The factors influencing the performance of the resulting biosensor were studied in detail. The biosensor responded to H₂O₂ in the linear range from 2.6 × 10^− 6 mol/L to 8.8 × 10^− 3 mol/L with a detection limit of 6.4 × 10^− 7 mol/L. Moreover, the studied biosensor exhibited good accuracy and high sensitivity. The proposed method was economical and efficient, making it potentially attractive for the application to real sample analysis.     

Uprising the antioxidant power of Argania spinosa L. callus through abiotic elicitation

This study was carried out in order to investigate the ability of tissues of Argania spinosa (L.) to undergo unlimited cell divisions by triggering their proliferative potential via callogenesis. Axenic cultures were efficiently established using axillary buds cultured on half-strength Murashige and Skoog (MS) medium after 20 min of surface sterilization with sodium hypochlorite 6% (v/v). The highest callus rate was achieved with 1.0 mg L⁻¹ of naphthaleneacetic acid (NAA) and 1.0 mg L⁻¹ of 2,4-dichlorophenoxyacetic acid (2,4D) or similarly with 0.01 mg L⁻¹ of 6-benzylaminopurine (BAP) and 1.0 mg L⁻¹ of 2,4D at pH of 5.8, under dark conditions. The results of this study show also a significant increase in the callus's antioxidant power under abiotic pressure induced by NaCl. Catalase (CAT), peroxidase (PO), and superoxide dismutase (SOD) activities were significantly triggered, which protected the cells from the stimulated oxidative stress, under hydrogen peroxide (H₂O₂) significant release. This reaction favors subsequently the tissue recover process linked to the low abundance of polyphenol oxidase (PPO) activity and malondialdehyde (MDA) content. This work proves the efficiency of salt stress in boosting the argan cell's antioxidant status, which could be commercially applied in the field of cells regenerative therapy.     

Isoorientin-6″-O-glucoside,a water-soluble antioxidant isolated from Gentiana arisanensis

The antioxidant activities of isoorientin-6″-O-glucoside were studied using various models. Isoorientin-6″-O-glucoside was more potent than Trolox, probucol and butylated hydroxytoluene (BHT) in reducing the stable free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH). It also scavenged superoxide anion, peroxyl and hydroxyl radicals that were generated by xanthine/xanthine oxidase, 2,2′-azobis(2-amidinopropane) dihydrochloride (AAPH) and Fe³⁺–ascorbate–EDTA–H₂O₂ system, respectively. The IC₅₀ value, stoichiometry factor and second-order rate constant were 9.0 ± 0.8 μM, 1.8 ± 0.1 and 2.6 × 10¹⁰ M⁻¹ s⁻¹ for superoxide generation, peroxyl and hydroxyl radicals. However, isoorientin-6″-O-glucoside did not inhibit xanthine oxidase activity or scavenge hydrogen peroxide (H₂O₂), carbon radical or 2,2′-azobis(2,4-dimethylvaleronitrile) (AMVN)-derived peroxyl radical in hexane. Isoorientin-6″-O-glucoside inhibited Cu²⁺-induced oxidation of human low-density lipoprotein (LDL) as measured by fluorescence intensity, thiobarbituric acid-reactive substance formation and electrophoretic mobility. Since isoorientin-6″-O-glucoside did not possess pro-oxidant activity, it may be an effective water-soluble antioxidant that can prevent LDL against oxidation.     

Oxidative stress caused by pyocyanin impairs CFTR Cl− transport in human bronchial epithelial cells

Pyocyanin (N-methyl-1-hydroxyphenazine), a redox-active virulence factor produced by the human pathogen Pseudomonas aeruginosa, is known to compromise mucociliary clearance. Exposure of human bronchial epithelial cells to pyocyanin increased the rate of cellular release of H₂O₂ threefold above the endogenous H₂O₂ production. Real-time measurements of the redox potential of the cytosolic compartment using the redox sensor roGFP1 showed that pyocyanin (100 μM) oxidized the cytosol from a resting value of − 318 ± 5 mV by 48.0 ± 4.6 mV within 2 h; a comparable oxidation was induced by 100 μM H₂O₂. Whereas resting Cl⁻ secretion was slightly activated by pyocyanin (to 10% of maximal currents), forskolin-stimulated Cl⁻ secretion was inhibited by 86%. The decline was linearly related to the cytosolic redox potential (1.8% inhibition/mV oxidation). Cystic fibrosis bronchial epithelial cells homozygous for ΔF508 CFTR failed to secrete Cl⁻ in response to pyocyanin or H₂O₂, indicating that these oxidants specifically target the CFTR and not other Cl⁻ conductances. Treatment with pyocyanin also decreased total cellular glutathione levels to 62% and cellular ATP levels to 46% after 24 h. We conclude that pyocyanin is a key factor that redox cycles in the cytosol, generates H₂O₂, depletes glutathione and ATP, and impairs CFTR function in Pseudomonas-infected lungs.     

Termination of a toxic Alexandrium bloom with hydrogen peroxide

The dinoflagellate Alexandrium ostenfeldii is a well-known harmful algal species that can potentially cause paralytic shellfish poisoning (PSP). Usually A. ostenfeldii occurs in low background concentrations only, but in August of 2012 an exceptionally dense bloom of more than 1 million cells L⁻¹ occurred in the brackish Ouwerkerkse Kreek in The Netherlands. The A. ostenfeldii bloom produced both saxitoxins and spirolides, and is held responsible for the death of a dog with a high saxitoxin stomach content. The Ouwerkerkse Kreek routinely discharges its water into the adjacent Oosterschelde estuary, and an immediate reduction of the bloom was required to avoid contamination of extensive shellfish grounds. Previously, treatment of infected waters with hydrogen peroxide (H₂O₂) successfully suppressed cyanobacterial blooms in lakes. Therefore, we adapted this treatment to eradicate the Alexandrium bloom using a three-step approach. First, we investigated the required H₂O₂ dosage in laboratory experiments with A. ostenfeldii. Second, we tested the method in a small, isolated canal adjacent to the Ouwerkerkse Kreek. Finally, we brought 50 mg L⁻¹ of H₂O₂ into the entire creek system with a special device, called a water harrow, for optimal dispersal of the added H₂O₂. Concentrations of both vegetative cells and pellicle cysts declined by 99.8% within 48 h, and PSP toxin concentrations in the water were reduced below local regulatory levels of 15 μg L⁻¹. Zooplankton were strongly affected by the H₂O₂ treatment, but impacts on macroinvertebrates and fish were minimal. A key advantage of this method is that the added H₂O₂ decays to water and oxygen within a few days, which enables rapid recovery of the system after the treatment. This is the first successful field application of H₂O₂ to suppress a marine harmful algal bloom, although Alexandrium spp. reoccurred at lower concentrations in the following year. The results show that H₂O₂ treatment provides an effective emergency management option to mitigate toxic Alexandrium blooms, especially when immediate action is required.     

Peroxynitrite and hydrogen peroxide elicit similar cellular stress responses mediated by the Ccp1 sensor protein

Peroxynitrite [ONOO(H)] is an oxidant associated with deleterious effects in cells. Because it is an inorganic peroxide that reacts rapidly with peroxidases, we speculated that cells may respond to ONOO(H) and H₂O₂ challenge in a similar manner. We exposed yeast cells to SIN-1, a well-characterized ONOO(H) generator, and observed stimulation of catalase and peroxiredoxin (Prx) activities. Previously, we reported that H₂O₂ challenge increases these activities in wild-type cells and in cells producing the hyperactive mutant H₂O₂ sensor Ccp1^W191F but not in Ccp1-knockout cells (ccp1Δ). We find here that the response of ccp1Δ and ccp1^W191F cells to SIN-1 mirrors that to H₂O₂, identifying Ccp1 as a sensor of both peroxides. SIN-1 simultaneously releases ^•NO and O₂^•−, which react to form ONOO(H), but exposure of the three strains separately to an ^•NO donor (spermine-NONOate) or an O₂^•− generator (paraquat) mainly depresses catalase or Prx activity, whereas co-challenge with the NONOate and paraquat stimulates these activities. Because Ccp1 appears to sense ONOO(H) in cells, we examined its reaction with ONOO(H) in vitro and found that peroxynitrous acid (ONOOH) rapidly (k₂>10⁶ M⁻¹ s⁻¹) oxidizes purified Ccp1 to an intermediate with spectral and ferrocytochrome-oxidizing properties indistinguishable from those of its well-characterized compound I formed with H₂O₂. Importantly, the nitrite released from ONOOH is not oxidized to ^•NO₂ by Ccp1^׳s compound I, unlike peroxidases involved in immune defense. Overall, our results reveal that yeast cells mount a common antioxidant response to ONOO(H) and H₂O₂, with Ccp1 playing a pivotal role as an inorganic peroxide sensor.     

Batch and continuous fermentative production of hydrogen with anaerobic sludge entrapped in a composite polymeric matrix

Cell immobilization techniques were adopted to biohydrogen production using immobilized anaerobic sludge as the seed culture. Sucrose-based synthetic wastewater was converted to H₂ using batch and continuous cultures. A novel composite polymeric material comprising polymethyl methacrylate (PMMA), collagen, and activated carbon was used to entrap biomass for H₂ production. Using the PMMA immobilized cells, the favorable conditions for batch H₂ fermentation were 35 °C, pH 6.0, and an 20 g COD l⁻¹ of sucrose, giving a H₂ production rate of 238 ml h⁻¹ l⁻¹ and a H₂ yield of 2.25 mol H₂ mol sucrose⁻¹. Under these optimal conditions, continuous H₂ fermentation was conducted at a hydraulic retention time (HRT) of 4–8 h, giving the best H₂-producing rate of 1.8 l h⁻¹ l⁻¹ (over seven-fold of the best batch result) at a HRT of 6 h and a H₂ yield of 2.0 mol H₂ mol sucrose⁻¹. The sucrose conversion was essentially over 90% in all runs. The biogas consisted of only H₂ and CO₂. The major soluble metabolites were butyric acid, acetic acid, and 2,3-butandiol, while a small amount of ethanol also detected. The PMMA-immobilized-cell system developed in this work seems to be a promising H₂-producing process due to the high stability in continuous operations and the capability of achieving a competitively high H₂ production rate under a relatively low organic loading rate.     

An amperometric biosensor based on horseradish peroxidase immobilized onto maize tassel-multi-walled carbon nanotubes modified glassy carbon electrode for determination of heavy metal ions in aqueous solution

A biosensor for trace metal ions based on horseradish peroxidase (HRP) immobilized on maize tassel-multiwalled carbon nanotube (MT-MWCNT) through electrostatic interactions is described herein. The biosensor was characterized using Fourier transform infrared (FTIR), UV–vis spectrometry, voltammetric and amperometric methods. The FTIR and UV–vis results inferred that HRP was not denatured during its immobilization on MT-MWCNT composite. The biosensing principle was based on the determination of the cathodic responses of the immobilized HRP to H₂O₂, before and after incubation in trace metal standard solutions. Under optimum conditions, the inhibition rates of trace metals were proportional to their concentrations in the range of 0.092–0.55 mg L⁻¹, 0.068–2 mg L⁻¹ for Pb²⁺ and Cu²⁺ respectively. The limits of detection were 2.5 μg L⁻¹ for Pb²⁺ and 4.2 μg L⁻¹ for Cu²⁺. Representative Dixon and Cornish-Bowden plots were used to deduce the mode of inhibition induced by the trace metal ions. The inhibition was reversible and mixed for both metal ions. Furthermore, the biosensor showed good stability, selectivity, repeatability and reproducibility.     

Effects of flooding on photosynthesis and root respiration in saltcedar (Tamarix ramosissima), an invasive riparian shrub

The introduced shrub Tamarix ramosissima invades riparian zones, but loses competitiveness under flooding. Metabolic effects of flooding could be important for T. ramosissima, but have not been previously investigated. Photosynthesis rates, stomatal conductance, internal (intercellular) CO₂, transpiration, and root alcohol dehydrogenase (ADH) activity were compared in T. ramosissima across soil types and under drained and flooded conditions in a greenhouse. Photosynthesis at 1500 μmol quanta m⁻² s⁻¹ (A₁₅₀₀) in flooded plants ranged from 2.3 to 6.2 μmol CO₂ m⁻² s⁻¹ during the first week, but A₁₅₀₀ increased to 6.4–12.7 μmol CO₂ m⁻² s⁻¹ by the third week of flooding. Stomatal conductance (g_s) at 1500 μmol quanta m⁻² s⁻¹ also decreased initially during flooding, where g_s was 0.018 to 0.099 mol H₂O m⁻² s⁻¹ during the first week, but g_s increased to 0.113–0.248 mol H₂O m⁻² s⁻¹ by the third week of flooding. However, photosynthesis in flooded plants was reduced by non-stomatal limitations, and subsequent increases indicate metabolic acclimation to flooding. Root ADH activities were higher in flooded plants compared to drained plants, indicating oxygen stress. Lower photosynthesis and greater oxygen stress could account for the susceptibility of T. ramosissima at the onset of flooding. Soil type had no effect on photosynthesis or on root ADH activity. In the field, stomatal conductance, leaf water potential, transpiration, and leaf δ¹³C were compared between T. ramosissima and other flooded species. T. ramosissima had lower stomatal conductance and water potential compared to Populus deltoides and Phragmites australis. Differences in physiological responses for T. ramosissima could become important for ecological concerns.     

Adsorption characteristics of sulfur powder by bamboo charcoal to restrain sulfur allergies

Exposures to particulate matter with a diameter of 2.5 μm or less (PM2.5) may influence the risk of birth defects and make you allergic, which causes serious harm to human health. Bamboo charcoal can adsorb harmful substances,that was of benefitto people’s health. In order to figure out the optimal adsorbtion condition and the intrinsic change of bamboo charcoal, five chemicals were adsorbed by bamboo charcoal and were analyzed by FT-IR. The optimal blast time was 80 min of Na₂SO₃, 100 min of Na₂S₂O₈, 20 min of Na₂SO₄, 120 min of Fe₂(SO₄)₃ and 60 min or 100 min of S. FT-IR spectra showed that bamboo charcoal had five characteristic peaks of SS stretch, H₂O stretch, OH stretch, CO stretch or CC stretch, and NO₂ stretch at 3850 cm⁻¹, 3740 cm⁻¹, 3430 cm⁻¹, 1630 cm⁻¹ and 1530 cm⁻¹, respectively. For Na₂SO₃, the peaks at 3850 cm⁻¹, 3740 cm⁻¹, 3430 cm⁻¹, 1630 cm⁻¹ and 1530 cm⁻¹ achieved the maximum at 20 min. For Na₂S₂O₈, the peaks at 3850 cm⁻¹, 3740 cm⁻¹, 3430 cm⁻¹ and 1530 cm⁻¹ achieved the maximum at 40 min. For Na₂SO₄, the peaks at 3850 cm⁻¹, 3740 cm⁻¹ and 1530 cm⁻¹ achieved the maximum at 40 min. For Fe₂(SO₄)₃, the peaks at 3850 cm⁻¹, 3740 cm⁻¹, 1630 cm⁻¹ and 1530 cm⁻¹ achieved the maximum at 120 min. For S, the peaks at 3850 cm⁻¹ and 3740 cm⁻¹ achieved the maximum at 40 min, the peaks at 1630 cm⁻¹ and 1530 cm⁻¹ achieved the maximum at 40 min. It proved that bamboo charcoal could remove sulfur powder from air to restrain sulfur allergies.     

Enhanced exopolysaccharide production and biological activity of Lactobacillus rhamnosus ZY with calcium and hydrogen peroxide

Exopolysaccharides (EPS) are important food and drug additives with beneficial antioxidant, anticancer, and immune-related effects on human health. However, the EPS is limited by low yields and the need for complex culture conditions in fermentation. Here, we report that hydrogen peroxide and calcium stimulated probiotic activity and production of crude exopolysaccharide (c-EPS) by Lactobacillus rhamnosus ZY. Accordingly, supplementation with 3 mM H₂O₂ allowed c-EPS biosynthesis to reach 567 mg/L after 24 h. Addition of both CaCl₂ and H₂O₂ resulted in a c-EPS yield of 2498 mg/L after 12 h, over 9-fold higher than that of an anaerobic culture. We observed that exposure to calcium and hydrogen peroxide made the cells more hydrophobic and led to the over-expression of GroEL, NADH peroxidase, and glyceraldehyde 3-phosphate dehydrogenase, thus increasing energy storage and EPS production. Chromatographic analysis revealed c-EPS was composed mainly of mannose (5.1%), galactose (15.3%), glucose (20–30%), and rhamnose (50–60%). Preliminary in vitro tests revealed that H₂O₂ and CaCl₂ enhanced the 2,2-diphenyl-1-picrylhydrazyl and hydroxyl radical scavenging capacities, resulting in a notable protective effect against oxidative damage in NIH/3T3 cells. Our study provides a simple and cost-effective approach for achieving high yields of good quality EPS using Lactobacillus rhamnosus.     

Examination of the superoxide/hydrogen peroxide forming and quenching potential of mouse liver mitochondria

Pyruvate dehydrogenase (PDHC) and α-ketoglutarate dehydrogenase complex (KGDHC) are important sources of reactive oxygen species (ROS). In addition, it has been found that mitochondria can also serve as sinks for cellular hydrogen peroxide (H₂O₂). However, the ROS forming and quenching capacity of liver mitochondria has never been thoroughly examined. Here, we show that mouse liver mitochondria use catalase, glutathione (GSH), and peroxiredoxin (PRX) systems to quench ROS. Incubation of mitochondria with catalase inhibitor 3-amino-1,2,4-triazole (triazole) induced a significant increase in pyruvate or α-ketoglutarate driven O₂⁻/H₂O₂ formation. 1-Choro-2,4-dinitrobenzene (CDNB), which depletes glutathione (GSH), elicited a similar effect. Auranofin (AF), a thioredoxin reductase-2 (TR2) inhibitor which disables the PRX system, did not significantly change O₂⁻/H₂O₂ formation. By contrast catalase, GSH, and PRX were all required to scavenging extramitochondrial H₂O₂. In this study, the ROS forming potential of PDHC, KGDHC, Complex I, and Complex III was also profiled. Titration of mitochondria with 3-methyl-2-oxovaleric acid (KMV), a specific inhibitor for O₂⁻/H₂O₂ production by KGDHC, induced a ~ 86% and ~ 84% decrease in ROS production during α-ketoglutarate and pyruvate oxidation. Titration of myxothiazol, a Complex III inhibitor, decreased O₂⁻/H₂O₂ formation by ~ 45%. Rotenone also lowered ROS production in mitochondria metabolizing pyruvate or α-ketoglutarate indicating that Complex I does not contribute to ROS production during forward electron transfer from NADH. Taken together, our results indicate that KGDHC and Complex III are high capacity sites for O₂⁻/H₂O₂ production in mouse liver mitochondria. We also confirm that catalase plays a role in quenching either exogenous or intramitochondrial H₂O₂.     

Response surface analysis for enzymatic decolorization of Congo red by manganese peroxidase

The enzymatic decolorization process of manganese peroxidase (MnP) is a complex system, which is greatly affected by the concentrations of H₂O₂, Mn²⁺, dye and enzyme. This work aimed to study these factors and investigate the combined interactions between them by applying response surface methodology (RSM) for decolorization of Congo red with MnP from Schizophyllum sp. F17, meanwhile conventional one-factor-at-a-time analysis was carried out. Through the one-factor-at-a-time analysis the optimized H₂O₂, Mn²⁺, Congo red and MnP extract was 0.2 mM, 0.5 mM, 50 mg/l and 0.8 ml, respectively, and the maximum decolorization attained under such conditions was 24.2%. Response surface analysis was conducted through Box–Behnken design and a second-order polynomial model (R² = 0.8565) was generated to describe the combined effect and the interactions quantificationally. ANOVA analysis indicated that the interactions between H₂O₂ and MnP, between dye and MnP were significant; the optimum condition through RSM was found to be 0.35 mM H₂O₂, 0.5 mM Mn²⁺, 75 mg/l Congo red and 1.4 ml MnP extract, for maximum decolorization of 30.8%.     

Desulphurization characteristics of bamboo charcoal from sulfur solution

Sulfur powder and sulfur dioxide (SO₂) often floated in air, produced acid rain and algal blooms, and could cause diseases. Bamboo charcoal could have adsorption and filtration properties. In order to figure out the optimal adsorption condition and the intrinsic change of the bamboo charcoal, five chemicals were adsorbed by bamboo charcoal and were analyzed by FT-IR. Fe₂(SO₄)₃’s, Na₂SO₄’s, Na₂S₂O₈’s, S’s, and Na₂SO₃’s optimal adsorption condition was the concentration of 19 g/1000 g and stir time of 20 min, 21 g/1000 g and stir time of 60 min, 7 g/1000 g and stir time of 120 min, 11 g/1000 g and stir time of 120 min, 21 g/1000 g and stir time of 60 min, respectively. FT-IR spectra showed that for FT-IR spectra of Fe₂(SO₄)₃, the transmissivity of the peaks at 3435 cm⁻¹ and 2925 cm⁻¹ achieved the maximum for 60 min and the concentration was 19 g/1000 g, the transmissivity of the peaks at 1630 cm⁻¹, 1060 cm⁻¹ and 660 cm⁻¹ achieved the maximum for 60 min and the concentration was 7 g/1000 g. For FT-IR spectra of Na₂SO₄, the transmissivity of the peaks at 1630 cm⁻¹, 1060 cm⁻¹ and 660 cm⁻¹ achieved the maximum for 20 min and the concentration was 13 g/1000 g. For FT-IR spectra of Na₂S₂O₈, the transmissivity of the peaks at 3435 cm⁻¹, 2925 cm⁻¹, 1630 cm⁻¹ and 1060 cm⁻¹ achieved the maximum for 120 min and the concentration was 19 g/1000 g. For FT-IR spectra of S, the transmissivity of the peaks at 3435 cm⁻¹, 2925 cm⁻¹, 1630 cm⁻¹ and 1060 cm⁻¹ achieved the maximum for 20 min and the concentration was 11 g/1000 g, 17 g/1000 g and 21 g/1000 g. For FT-IR spectra of Na₂SO₃, the transmissivity of the peaks at 3435 cm⁻¹ achieved the maximum for 120 min and the concentration was 5 g/1000 g, the transmissivity of the peaks at 2925 cm⁻¹, 1630 cm⁻¹ and 1060 cm⁻¹ achieved the maximum for 120 min and the concentration was 11 g/1000 g. In these states, the number of the transmissivity of the maximum peaks is the largest.