The aim of this paper was to assess the total antioxidant capacity of some commercial fruit juices (namely citrus), spectrophotometrically and by the biamperometric method, using the redox couple DPPH· (2,2-diphenyl-1-picrylhydrazyl)/DPPH (2,2-diphenyl-1-picrylhydrazine). Trolox® was chosen as a standard antioxidant. In the case of the spectrophometric method, the absorbance decrease of the DPPH· solution was followed. For the biamperometric method, the influence of some parameters like the potential diference, ΔE, DPPH· concentration, and Trolox® concentration was investigated. The calibration graph obtained for Trolox® presents linearity between 5 and 30 µM, (y = 0.059 x + 0.0564, where y represents the value of current intensity, expressed as μA and x the value of Trolox® concentration, expressed as μM; r2 = 0.9944). The R.S.D. value for the biamperometric method was 1.29% (n = 10, c = 15 μM Trolox®). In the case of the spectrophotometric method, the calibration graph obtained for Trolox® presents linearity between 0.01 and 0.125 mM (y = -9.5789 x+1.4533, where y represents the value of absorbance and x, the value of Trolox® concentration, expressed as mM; r2 = 0.9963). The R.S.D. value for the spectrophotometric method was 2.05%. Both methods were applied to total antioxidant activity determination in real samples (natural juices and soft drinks) and the results were in good agreement.
α-Hexachlorocyclohexane (α-HCH) is the only chiral isomer of the eight 1,2,3,4,5,6-HCHs and we have developed an enantiomer-specific stable carbon isotope analysis (ESIA) method for the evaluation of its fate in the environment. The carbon isotope ratios of the α-HCH enantiomers were determined for a commercially available α-HCH sample using a gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) system equipped with a chiral column. The GC-C-IRMS measurements revealed δ-values of -32.5 ± 0.8‰ and -32.3 ± 0.5‰ for (-) α-HCH and (+) α-HCH, respectively. The isotope ratio of bulk α-HCH was estimated to be -32.4 ± 0.6‰ which was in accordance with the δ-values obtained by GC-C-IRMS (-32.7 ± 0.2‰) and elemental analyzer-isotope ratio mass spectrometry (EA-IRMS) of the bulk α-HCH (-32.1 ± 0.1‰). The similarity of the isotope ratio measurements of bulk α-HCH by EA-IRMS and GC-C-IRMS indicates the accuracy of the chiral GC-C-IRMS method. The linearity of the α-HCH ESIA method shows that carbon isotope ratios can be obtained for a signal size above 100 mV. The ESIA measurements exhibited standard deviations (2σ) that were mostly < ± 0.5‰. In order to test the chiral GC-C-IRMS method, the isotope compositions of individual enantiomers in biodegradation experiments of α-HCH with Clostridium pasteurianum and samples from a contaminated field site were determined. The isotopic compositions of the α-HCH enantiomers show a range of enantiomeric and isotope patterns, suggesting that enantiomeric and isotope fractionation can serve as an indicator for biodegradation and source characterization of α-HCH in the environment.
Carbon isotope fractionation factors were determined with the dichloro elimination of gamma-hexachlorocyclohexane (gamma-HCH) by the sulfate-reducing bacteria Desulfococcus multivorans DSM 2059 and Desulfovibrio gigas DSM 1382. Both strains are known for cometabolic HCH dechlorination. Degradation experiments with gamma-HCH in concentrations of 22-25 gammaM were carried out using benzoate (for D. multivorans) and lactate (for D. gigas) as electron donors, respectively. Gamma-HCH was dechlorinated by both bacterial strains within four weeks, and the metabolites gamma-3,4,5,6-tetrachlorocyclohexene (gamma-TCCH), chlorobenzene (CB), and benzene were formed. The carbon isotope fractionation of gamma-HCH dechlorination was quantified by the Rayleigh model, using a bulk enrichment factor (epsilon C) of -3.9 +/- 0.6 for D. gigas and -3.4 +/- 0.5 for D. multivorans, which correspond to apparent kinetic isotope effect (AKIEc) values of 1.023 +/- 0.004 or 1.02 +/- 0.003 for stepwise Cl-C bond cleavage. The extent and range of isotope fractionation suggest that gamma-HCH dechlorination can be monitored in anoxic environments by compound-specific isotope analysis (CSIA).
The Ca2+‐dependent response to oxidative stress caused by H2O2 or tert‐butylhydroperoxide (tBOOH) was investigated in Saccharomyces cerevisiae cells expressing transgenic cytosolic aequorin, a Ca2+‐dependent photoprotein. Both H2O2 and tBOOH induced an immediate and short‐duration cytosolic Ca2+ increase that depended on the concentration of the stressors. Sublethal doses of H2O2 induced Ca2+ entry into the cytosol from both extracellular and vacuolar sources, whereas lethal H2O2 shock mobilized predominantly the vacuolar Ca2+. Sublethal and lethal tBOOH shocks induced mainly the influx of external Ca2+, accompanied by a more modest vacuolar contribution. Ca2+ transport across the plasma membrane did not necessarily involve the activity of the Cch1p/Mid1p channel, whereas the release of vacuolar Ca2+ into the cytosol required the vacuolar channel Yvc1p. In mutants lacking the Ca2+ transporters, H2O2 or tBOOH sensitivity correlated with cytosolic Ca2+ overload. Thus, it appears that under H2O2‐induced or tBOOH‐induced oxidative stress, Ca2+ mediates the cytotoxic effect of the stressors and not the adaptation process.
A method was developed for assessing ascorbic acid concentration in commercial fruit juice by cyclic voltammetry. The anodic oxidation peak for ascorbic acid occurs at about 490 mV on a Pt disc working electrode (versus SCE). The influence of the potential sweep speed on the peak height was studied. The obtained calibration graph shows a linear dependence between peak height and ascorbic acid concentration in the domain (0.1–10 mmol·L−1). The equation of the calibration graph was y = 6.391x + 0.1903 (where y represents the value of intensity measured for the anodic peak height, expressed as μA and x the analyte concentration, as mmol·L−1, r2 = 0.9995, r.s.d. = 1.14%, n = 10, Cascorbic acid = 2 mmol·L−1). The developed method was applied to ascorbic acid assessment in fruit juice. The ascorbic acid content determined ranged from 0.83 to 1.67 mmol·L−1 for orange juice, from 0.58 to 1.93 mmol·L−1 for lemon juice, and from 0.46 to 1.84 mmol·L−1 for grapefruit juice. Different ascorbic acid concentrations (from standard solutions) were added to the analysed samples, the degree of recovery being comprised between 94.35% and 104%. Ascorbic acid determination results obtained by cyclic voltammetry were compared with those obtained by the volumetric method with dichlorophenol indophenol. The results obtained by the two methods were in good agreement.
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