Effect of Zinc nanoparticles on oxidative stress-related genes and antioxidant enzymes activity in the brain of Oreochromis niloticus and Tilapia zillii

This study was carried out to determine the median lethal concentrations (LC₅₀) of Zinc nanoparticles (ZnNPs) on Oreochromis niloticus and Tilapia zillii. The biochemical and molecular potential effects of ZnNPs (500 and 2000 μg L⁻¹) on the antioxidant system in the brain tissue of O. niloticus and T. zillii were investigated. Four hundred fish were used for acute and sub-acute studies. ZnNP LC₅₀ concentrations were investigated in O. niloticus and T. zillii. The effect of 500 and 2000 μg L⁻¹ ZnNPs on brain antioxidants of O. niloticus and T. zillii was investigated. The result indicated that 69 h LC₅₀ was 5.5 ± 0.6 and 5.6 ± 0.4 for O. nilotica and T. zillii, respectively. Fish exposed to 500 μg L⁻¹ ZnNPs showed a significant increase in reduced glutathione (GSH), total glutathione (tGSH) levels, superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx) and glutathione-S-transferase (GST) activity and gene expression. On the contrary, malondialdehyde (MDA) levels significantly decreased. Meanwhile, fish exposed to 2000 μg L⁻¹ ZnNPs showed a significant decrease of GSH, tGSH levels, SOD, CAT, GR, GPx and GST activity and gene expression. On the contrary, MDA levels significantly increased. It was concluded that, the 96 h LC₅₀ of ZnNPs was 5.5 ± 0.6 and 5.6 ± 0.4 for O. nilotica and T. zillii, respectively. ZnNPs in exposure concentrations of 2000 μg/L induced a deleterious effect on the brain antioxidant system of O. nilotica and T. zillii. In contrast, ZnNPs in exposure concentrations of 500 μg L⁻¹ produced an inductive effect on the brain antioxidant system of O. nilotica and T. zillii.     

MTs in Palaemonetes argentinus as potential biomarkers of zinc contamination in freshwaters

Aquatic invertebrates take up and accumulate essential and non-essential trace metals even when both are likely to be poisonous. In order to study the potential of the metallothioneins (MTs) as biomarkers of metal contamination in native shrimp Palaemonetes argentinus, organisms have been exposed at 0, 5, 50 and 500 μg L⁻¹ of zinc for 96 h. Moreover, accumulation and subcellular distribution of this essential metal were evaluated. A significant Zn accumulation was observed in different body sections. Higher Zn levels occurred in cephalothorax compared to abdomen, especially at the highest exposure concentration (500 μg Zn L⁻¹). A clear differential subcellular metal distribution between cephalothorax and abdomen was also observed. In cephalothorax Zn was similarly distributed between the soluble and insoluble fractions; while in abdomen, when total Zn increased, insoluble metal augmented more markedly than the soluble one. Cytosolic Zn levels increased more in cephalothorax than in abdomen of shrimps exposed to 500 μg Zn L⁻¹ when compared to control. Finally, a significant induction of MTs was observed in cephalothorax at 500 μg Zn L⁻¹. A potential role for MTs as biomarkers in P. argentinus should be further studied to enhance the sensitivity of the response, although it is likely that MTs play a key role in metal detoxification since the increase of these proteins is linked to metal challenge.     

Insecticidal properties of Pimpinella anisum essential oils against the Culex quinquefasciatus and the non-target organism Daphnia magna

The efficacy of an essential oil obtained from Pimpinella anisum fruits and its major compound, trans-Anethole, was tested on the eggs, larvae and adults of Culex quinquefasciatus. While causing no significant mortality on eggs, other tested stages were very sensitive to the essential oil and trans-Anethole. LC₅₀ for the 2nd to 4th instar larvae was estimated as 26–27 μL·L^− 1 and 15–19 μL·L^− 1 for the essential oil and trans-Anethole, respectively. As for the essential oil applied on adults, LC(LD)₅₀ was estimated as 9.3 μL mL^− 1 (spray test), 1.9 μL L^− 1 (fumigation test) and 0.6 μg cm^− 2 (tarsal test), and for trans-Anethole as 8.1 μL mL^− 1 (spray test), 2.1 μL L^− 1 (fumigation test) and 0.4 μg cm^− 2 (tarsal test). The time needed to achieve 50% mortality after application of LC(LD)₉₉ of the essential oil was significantly different; for example, in larvicidal assays it ranged from 15 to 235 min depending on the larval instar, and from 9 to 180 min when applied to adults, depending on the mode of application. It was also found that temperature had an important effect on the larvicidal efficacy of the essential oil, and oviposition deterrent activity was studied.The essential oil and trans-Anethole were toxic for Daphnia magna (62–92% mortality) and significantly reduced its fertility at high concentrations (35–50 μL mL^− 1) and long exposure (48 h). However, no negative effect on Daphnia mortality or fertility was found at shorter exposure times (6 h) and/or lower concentrations (20 μL mL^− 1).Based on the results of this study, we can recommend the essential oil from P. anisum as a suitable active substance for potential botanical insecticides.     

Accumulation and detoxification dynamic of cyanotoxins in the freshwater shrimp Palaemonetes argentinus

The uptake and accumulation of microcystin-LR (MC-LR) in the shrimp Palaemonetes argentinus was investigated using both laboratory and field assays. Shrimps were exposed in aquarium during 1, 2, 3 and 7 days to 1, 10 and 50 μg L⁻¹ MCLR. Accumulation (0.7 ± 0.2 μg MC-LR g⁻¹) was observed after three days exposures to 50 μg L⁻¹ toxin. Then, shrimps were relocated in fresh water (free of MCLR) to verify the detoxification dynamic, showing a drop to 0.18 ± 0.01 μg MCLR g⁻¹ after three days. The activity of glutathione-S-transferase, measured in the microsomal fraction (mGST), was significantly increased during the exposure period, with further increment during the detoxification period. Furthermore, cytosolic GST (sGST) and glutathione reductase (GR) increased their activities during detoxification, while inhibition was observed for catalase (CAT) with no significant changes for glutathione peroxidase (GPx). Current results suggest that GSH conjugation could be an important MC detoxification mechanism in P. argentinus and that MCLR induce oxidative stress in this shrimp.Field exposures were carried out in San Roque Reservoir (Córdoba, Argentina) after a cyanobacteria bloom. Nodularin (Nod) presence was measured for the first time in this waterbody (0.24 ± 0.04 μg L⁻¹), being the first report of Nod in South America freshwaters. Nod was also detected in Palaemonetes argentinus (0.16 ± 0.03 μg g⁻¹) after three weeks of exposure in this reservoir, with the concomitant activation of mGST, sGST and CAT.Although internal doses of Nod were low throughout the exposure, they were enough to cause biochemical disturbances in Palaemonetes argentinus.Further laboratory studies on Nod accumulation and antioxidant responses in Palaemonetes argentinus are necessary to fully understand these field results. P. argentinus should be considered a potential vector for transferring cyanotoxins to higher trophic levels in aquatic environments.     

Synergistic effect of Se-methylselenocysteine and vitamin E in ameliorating the acute ethanol-induced oxidative damage in rat

The present study was conduced to investigate the synergistic effects of combined treatments with Se-methylselenocysteine (SeMSC) and vitamin E (Vit E) in reversing oxidative stress induced by ethanol in serum and different tissues of rats. Sixty female rats were randomly divided into six groups for 30 days’ consecutive pretreatments as followed: control (I), physiological saline (II), 2.8 μg kg⁻¹ Se as SeMSC (III), 2.8 μg kg⁻¹ Se as sodium selenite (Na₂SeO₃, IV), 5 mg kg⁻¹ α-tocopherol as α-tocopherol acetate (Vit E, V), 5 mg kg⁻¹ α-tocopherol as α-tocopherol acetate and 2.8 μg kg⁻¹ Se as SeMSC (VI). All animals in groups II–VI were treated by ethanol treatment to cause oxidative stress. After 6 h of ethanol treatment, the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), the contents of total antioxidant capacity (T-AOC), malondialdehyde (MDA), glutathione (GSH) and carbonyl protein (CP) in the serum, liver, heart and kidney were measured. The result showed that the individual SeSMC, Na₂SeO₃ and vitamin E could effectively increase the SOD, T-AOC, GSH-Px and GSH contents as well as significantly decrease the MDA and CP concentrations in the tissues of ethanol-induced rats. At the same dose on different forms of Se, SeMSC showed greater antioxidant activity than Na₂SeO₃. Moreover, group VI (SeMSC and α-tocopherol acetate) showed much better antioxidant activity than individual group III (SeMSC) and V (α-tocopherol acetate) due to the synergistic effect.     

A green and efficient procedure for the preconcentration and determination of cadmium,nickel and zinc from freshwater,hemodialysis solutions and tuna fish samples by cloud point extraction and flame atomic absorption spectrometry

Cloud point extraction (CPE) was used to simultaneously preconcentrate trace-level cadmium, nickel and zinc for determination by flame atomic absorption spectrometry (FAAS). 1-(2-Pyridilazo)-2-naphthol (PAN) was used as a complexing agent, and the metal complexes were extracted from the aqueous phase by the surfactant Triton X-114 ((1,1,3,3-tetramethylbutyl)phenyl-polyethylene glycol). Under optimized complexation and extraction conditions, the limits of detection were 0.37 μg L⁻¹ (Cd), 2.6 μg L⁻¹ (Ni) and 2.3 μg L⁻¹ (Zn). This extraction was quantitative with a preconcentration factor of 30 and enrichment factor estimated to be 42, 40 and 43, respectively. The method was applied to different complex samples, and the accuracy was evaluated by analyzing a water standard reference material (NIST SRM 1643e), yielding results in agreement with the certified values.     

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.     

Effects of exposure to sublethal propiconazole on intestine-related biochemical responses in rainbow trout,Oncorhynchus mykiss

The effect of long-term (30 days) exposure to PCZ (0.2, 50, and 500 μg l^?1) on intestine-related biochemical markers in rainbow trout was investigated. Multiple biomarkers were measured, including digestive enzymes (proteolytic enzymes and amylase), antioxidant responses (TBARS, CP, SOD, CAT, GR and GPx) and energy metabolic parameters (RNA/DNA ratio, Na⁺-K⁺-ATPase). Exposure to 500 μg l^?1 PCZ led to significantly inhibited (p < 0.01) proteolytic enzyme and amylase activity. Activities of the antioxidant enzymes SOD, CAT, and GPx gradually increased at lower PCZ concentrations (0.2 and 50 μg l^?1). At the highest concentration (500 μg l^?1), oxidative stress was apparent as significant higher (p < 0.05) lipid peroxidation and protein carbonyls, associated with an inhibition of antioxidant enzymes activity. Moreover, energy metabolic parameters (RNA/DNA ratio, Na⁺-K⁺-ATPase) were significantly inhibited (p < 0.01) in the intestines of fish exposed to 500 μg l^?1 PCZ, compared with controls. We suggest that long-term exposure to PCZ could result in several responses in intestine-related biochemical markers, which potentially could be used as indicators for monitoring residual PCZ present in the aquatic environment.     

Isolation and characterization of Pseudomonas stutzeri QZ1 from an anoxic sulfide-oxidizing bioreactor

Bacterial strain QZ1 was isolated from sludge of anoxic sulfide-oxidizing (ASO) reactor. Based on 16S rDNA sequence analysis and morphological characteristics, the isolate was identified as Pseudomonas stutzeri. The isolate was found to be a facultative chemolithotroph, using sulfide as electron donor and nitrite as electron acceptor. The strain QZ1 produced sulfate as the major product of sulfide oxidation, depending on the initial sulfide and nitrite concentrations. The isolate was capable of growth under strictly autotrophic conditions. The growth and substrate removal of Pseudomonas stutzeri QZ1 were optimal at an initial pH of 7.5–8.0 at 30 °C. The specific growth rate (μ) was found as 0.035 h⁻¹ with a doubling time of 21.5 h. For isolate QZ1, the EC₅₀ values both for sulfide and nitrite were found to be 335.95 mg S L⁻¹ and 512.38 mg N L⁻¹, respectively, showing that the sulfide oxidation into sulfate by Pseudomonas stutzeri QZ1 was badly affected beyond these substrate concentrations.     

Bench-scale fermentation of Trichoderma viride on wastewater sludge: Rheology,lytic enzymes and biocontrol activity

Conidiation and lytic enzyme production by Trichoderma viride at different solids concentration of pre-treated municipal wastewater sludge was examined in a 15-L fermenter. The maximum conidia concentration (5.94 × 10⁷ CFU mL⁻¹ at 96 h) was obtained at 30 g L⁻¹ suspended solids. The maximum lytic enzyme activities were achieved around 12–30 h of fermentation. Bioassay against a fungal phytopathogen, Fusarium sp. showed maximum activity in the sample drawn around 96 h of fermentation at 30 g L⁻¹ suspended solids concentration. Entomotoxicity against spruce budworm larvae showed maximum value ≈17290 SBU μL⁻¹ at 30 g L⁻¹ suspended solids concentration at the end of fermentation (96 h). Plant bioassay showed dual action of T. viride, i.e., disease prevention and growth promotion. The rheological analyses of fermentation sludges showed the pseudoplastic behaviour. In order to maintain required dissolved oxygen concentration ≥30%, the agitation and aeration requirements significantly increased at 35 g L⁻¹ compared to 30 and 25 g L⁻¹. The oxygen uptake rate and volumetric oxygen mass transfer coefficient, k_La at 35 g L⁻¹ did not increase in comparison to 30 g L⁻¹ due to rheological complexity of the broth during fermentation. Thus, the successful fermentation operation of the biocontrol fungus T. viride is a rational indication of its potential for mass-scale production for agriculture and forest sector as a biocontrol agent.     

A new fungal peroxidase with alkaline-tolerant,chloride-enhancing activity and dye decolorization capacity

A new fungal peroxidase (Pspd) from Perenniporia subacida was purified by ammonium sulfate precipitation, DEAE-cellulose DE52 anionic exchange and Sepharose GL-6B chromatography, resulting in a high specific activity of 9.138 U mg⁻¹, 3.622-fold higher than that of crude enzyme at the same level. Polyacrylamide gel electrophoresis and UV–vis adsorption spectrum analysis showed that the purified enzyme is a heme-containing monomer with a molecular mass of 43.0 kDa. Optimal peroxidase activity was obtained at pH 5.5 and 30 °C when using 100.0 mM n-propanol as substrate, and under these conditions, the catalytic efficiency (k_cat/K_m) is 1.57 s⁻¹ μM⁻¹. Pspd was inhibited by l-cysteine, dithiothreitol, EDTA and sodium azide, but stimulated by Mn²⁺, Na⁺, Mg²⁺ and K⁺. The enzyme is stable over a broad pH range of 7.0–8.5 after incubation for 72 h, which indicated that the enzyme is lasting alkaline-tolerant. It was worth noting that the chloride at relatively low concentrations can enhance the peroxidase activity, with concomitant increase in substrate affinity. Additionally, Pspd performed high decolorization capability toward structurally various dyes and the capability was independent of the oxidizing mediators, with 75.31% of Neutral Red (50.0 mg L⁻¹) being decolorized by 1.5 U mL⁻¹ pure enzyme after incubation for 72 h. These properties demonstrated that Pspd has potentials for textile dyes decolorization applications.     

The effects of light intensity on the growth of Japanese Gambierdiscus spp. (Dinophyceae)

Marine toxic dinoflagellates of the genus Gambierdiscus are the causative agents of ciguatera fish poisoning (CFP), a form of seafood poisoning that is widespread in tropical, subtropical and temperate regions worldwide. The distributions of Gambierdiscus australes, Gambierdiscus scabrosus and two phylotypes of Gambierdiscus spp. type 2 and type 3 have been reported for the waters surrounding the main island of Japan. To explore the bloom dynamics and the vertical distribution of these Japanese species and phylotypes of Gambierdiscus, the effects of light intensity on their growth were tested, using a photoirradiation-culture system. The relationship between the observed growth rates and light intensity conditions for the four species/phylotypes were formulated at R > 0.92 (p < 0.01) using regression analysis and photosynthesis-light intensity (P-L) model. Based on this equation, the optimum light intensity (L_max) and the semi-optimum light intensity range (L_s-opt) that resulted in the maximum growth rate (μ_max) and ≥80% μ _max values of the four species/phylotypes, respectively, were as follows: (1) the L_max and L_s-opt of G. australes were 208 μmol photons m⁻² s⁻¹ and 91–422 μmol photons m⁻² s⁻¹, respectively; (2) those of G. scabrosus were 252 and 120–421 μmol photons m⁻² s⁻¹, respectively; (3) those of Gambierdiscus sp. type 2 were 192 and 75–430 μmol photons m⁻² s⁻¹, respectively; and (4) those of Gambierdiscus sp. type 3 were ≥427 and 73–427 μmol photons m⁻² s⁻¹, respectively. All four Gambierdiscus species/phylotypes required approximately 10 μmol photons m⁻² s⁻¹ to maintain growth. The light intensities in coastal waters at a site in Tosa Bay were measured vertically at 1 m intervals once per season. The relationships between the observed light intensity and depth were formulated using Beer’s Law. Based on these equations, the range of the attenuation coefficients at Tosa Bay site was determined to be 0.058–0.119 m⁻¹. The values 1700 μmol photons m⁻² s⁻¹, 500 μmol photons m⁻² s⁻¹, and 200 μmol photons m⁻² s⁻¹ were substituted into the equations to estimate the vertical profiles of light intensity at sunny midday, cloudy midday and rainy midday, respectively. Based on the regression equations coupled with the empirically determined attenuation coefficients for each of the four seasons, the ranges of the projected depths of L_max and L_s-opt for the four Gambierdiscus species/phylotypes under sunny midday conditions, cloudy midday conditions, and rainy midday conditions were 12–38 m and 12–54 m, 1–16 m and 1–33 m, and 0 m and 0–16 m, respectively. These results suggest that light intensity plays an important role in the bloom dynamics and vertical distribution of Gambierdiscus species/phylotypes in Japanese coastal waters.     

Myriophyllum aquaticum as a biomonitor of water heavy metal input related to agricultural activities in the Xanaes River (Córdoba,Argentina)

The aim of the present study was to assess the temporal variation of the heavy metal content (Co, Cu, Fe, Mn, Ni, Pb, and Zn) in surface water and sediments in relation to agricultural practices in the Xanaes River (Córdoba, Argentina). A second objective was to analyze possible relationships between the input of heavy metals on surface water and sediment, heavy metal accumulation and physiological changes in the aquatic plant Myriophyllum aquaticum. Samples were taken from the river at two contrasting sites (between April 2010 and August 2010): (1) a pristine area (mountain site), and (2) an area with intensive agricultural activity located at 60 km down river (agricultural site). The total concentration of heavy metals in surface water was higher in samples collected at the agricultural site but in sediments only the Mn concentration was higher than at the mountain site. The Fe and Mn concentrations in surface water at the agricultural site exceeded the recommended values for Argentinean Legislation of 300 μg L⁻¹ for Fe and 100 μg L⁻¹ for Mn. The accumulations of Zn and Mn in M. aquaticum were higher at the agricultural site and more elevated than the Zn and Mn concentrations in sediments at the same sites and sampling times. At the agricultural site, temporal variations of Cu, Fe and Zn were relatively similar for plants and water column, but the levels of the metals in plants were displaced over time. These results suggest that the levels of pollutants in the river came in pulses from the riverbank. These results show the potential use of M. aquaticum as a suitable accumulation biomonitor at the early stages of heavy metal pollution in rivers.     

Copper,zinc superoxide dismutase of Curcuma aromatica is a kinetically stable protein

This study details on cloning and characterization of Cu,Zn superoxide dismutase (Ca–Cu,Zn SOD) from a medicinally important plant species Curcuma aromatica. Ca–Cu,Zn SOD was 692 bp with an open reading frame of 459 bp. Expression of the gene in Escherichia coli cells followed by purification yielded the enzyme with K_m of 0.047 ± 0.008 μM and V_max of 1250 ± 24 units/mg of protein. The enzyme functioned (i) across a temperature range of −10 to +80 °C with temperature optima at 20 °C; and (ii) at pH range of 6–9 with optimum activity at pH 7.8. Ca–Cu,Zn SOD retained 50% of the maximum activity after autoclaving, and was stable at a wide storage pH ranging from 3 to 10. The enzyme tolerated varying concentrations of denaturating agent, reductants, inhibitors, trypsin, was fairly resistant to inactivation at 80 °C for 180 min (k_d, 6.54 ± 0.17 × 10⁻³ min⁻¹; t_1/2, 106.07 ± 2.68 min), and had midpoint of thermal transition (T_m) of 70.45 °C. The results suggested Ca–Cu,Zn SOD to be a kinetically stable protein that could be used for various industrial applications.     

Nitrite inhibition and intermediates effects on Anammox bacteria: A batch-scale experimental study

A batch test procedure, based on manometric measurements, was used to study the Anammox process, in particular the inhibition due to nitrite and the effects of hydroxylamine and hydrazine, indicated as possible intermediates of the process. The maximum nitrite removal rate (MNRR) was measured. The method showed good reliability with a standard error of 4.5 ± 3.3% (n: 41). All the tests were carried out on samples taken from a pilot plant with Anammox suspended biomass. The tests were used also to monitor the reactor activity. By testing different spiked additions of nitrite (10–75 mg NO₂⁻-N L⁻¹), a short-term inhibition, with more than 25% MNRR decrease, was found at concentrations higher than 60 mg NO₂⁻-N L⁻¹. Repeated additions of nitrite higher than 30 mg NO₂⁻-N L⁻¹ caused losses of activity. After a complete loss of activity, spiked additions of hydroxylamine (30 mg N L⁻¹ in total) determined a 20% permanent recovery. Low amounts of the intermediates (1–3 mg N L⁻¹) applied on partially inhibited samples and uninhibited samples produced temporary increases in activity up to 50% and 30%, respectively.     

Diel activities of antioxidant enzymes,photosynthetic pigments and malondialdehyde content in stationary-phase cells of Tetraselmis gracilis (Prasinophyceae)

The activities of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX), as well as photosynthetic pigment contents and free malondialdehyde (MDA), were determined in senescent batch cultures of Tetraselmis gracilis (Kylin) Butcher, under a cyclic light regime. A 2.6-fold increase in SOD activity (from 53 to 137 U mg⁻¹ protein) was observed in the light phase, contrasting with a 9-fold increase in CAT (from 1 to 9 μmol H₂O₂ min⁻¹ mg⁻¹ protein) and a 1.7-fold increase in APX (from 3 to 5 μmol ascorbate min⁻¹ mg⁻¹ protein) activities, both enzymes peaking in the dark phase. The β-carotene and lycopene content did not vary significantly with the light–dark cycle. The Chl a, Chl b, lutein, zeaxanthin, violaxanthin and neoxanthin pigments exhibited the highest values in the first half (3–6 h) of the light phase, followed by a declining trend and a plateau or a slight increase 3 h from the beginning of the dark phase onwards. The highest values for prasinoxanthin were observed in the second half of the dark phase and the first half of the light phase. None of the pigments displayed any discernible cyclic trend. The possibility of the xanthophyll cycle occurring during senescence is discussed in light of the high value (∼0.9) obtained for the zeaxanthin/(zeaxanthin + violaxanthin) ratio. The free MDA content was enhanced during the experimental period, which may be an indicator of oxidative stress in aging cell cultures. Our results indicated the occurrence of an imbalance between the production of reactive oxygen species and the antioxidant defense in stationary T. gracilis cells.     

Treatment of acidic wastewater arising from the refining of vegetable oil by crossflow microfiltration at very low transmembrane pressure

The refining process of vegetable oils generates acidic wastewater with the following characteristics: pH (1–1.5), COD (10–30 g O₂ L⁻¹), suspended solids (7–12 g L⁻¹) and fats (2–4 g L⁻¹). In order to reduce the effluent load and recover a fraction of the fats without using any additives, a microfiltration (0.2–1.4 μm) process involving ceramic membranes at very low transmembrane pressure values (0.1–1 bar) was assessed. Four batches of acidic wastewater from different manufacturing runs were tested. Trials with a constant volumetric reduction ratio of 30 were carried out for periods of more than 5 h. With a 0.5 μm membrane it was possible to maintain a permeate flux of 100 L h⁻¹ m⁻² for 24 h and achieve a 91% reduction in SS, a 96% reduction in fat and a COD reduction of more than 60%. In addition, the retentate thus extracted separated spontaneously into two phases, both of which could be exploited: the upper phase mainly consisting of fats as a by-product and the lower clarified phase which could be mixed into the permeate.     

Urea loading from a spring storm—Knysna estuary,South Africa

Urea concentrations were tracked in the Knysna estuary, South Africa, following a strong initial storm of the austral summer rainy season in 2000. These post-storm urea concentrations are compared against a 1-year survey of these concentrations and demonstrate the initial and longer-term impacts of urea loading on a near-pristine estuary. Fifteen stations along the main channel of the estuary were sampled four times each, 2–4 weeks before the storm, providing and average urea concentration of 1.4 μmol N L⁻¹. When these same stations were sampled 12 h after the storm the average urea concentration had increased to 9.4 μmol N L⁻¹. The average urea concentration at these 15 stations remained high, averaging 10.2 μmol N L⁻¹ and 10.0 μmol N L⁻¹ 36 and 84 h post-storm, respectively. The urea concentrations in the rivers supplying fresh water to the Knysna estuary, one draining pasturelands and the other draining informal housing settlements that relied on pit toilets and open sewers are also compared before and after this storm. These data suggest that spring storms significantly influence nutrient availability, which in turn, may be related to the large dinoflagellates blooms witnessed in the Knysna estuary during the later portion of the wet summer season.     

Continuous succinic acid production by Actinobacillus succinogenes in a biofilm reactor: Steady-state metabolic flux variation

Continuous anaerobic fermentations were performed in a novel external-recycle, biofilm reactor using d-glucose and CO₂ as carbon substrates. Succinic acid (SA) yields were found to be an increasing function of glucose consumption with the succinic acid to acetic acid ratio increasing from 2.4 g g⁻¹ at a glucose consumption of 10 g L⁻¹, to 5.7 g g⁻¹ at a glucose consumption of 50 g L⁻¹. The formic acid to acetic acid ratio decreased from an equimolar value (0.77 g g⁻¹) at a glucose consumption of 10 g L⁻¹ to a value close to zero at 50 g L⁻¹. The highest SA yield on glucose and highest SA titre obtained were 0.91 g g⁻¹ and 48.5 g L⁻¹ respectively. Metabolic flux analysis based on the established C₃ and C₄ metabolic pathways of Actinobacillus succinogenes revealed that the increase in the succinate to acetate ratio could not be attributed to the decrease in formic acid and that an additional source of NADH was present. The fraction of unaccounted NADH increased with glucose consumption, suggesting that additional reducing power is present in the medium or is provided by the activation of an alternative metabolic pathway.     

Purification and characterization of yellow laccase from Trametes hirsuta MTCC-1171 and its application in synthesis of aromatic aldehydes

A yellow laccase from the culture filtrate of Trametes hirsuta MTCC-1171 has been purified. The purification methods involved concentration of the culture filtrate by ammonium sulphate precipitation and an anion exchange chromatography on diethylaminoethyl cellulose. The sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and native polyacrylamide gel electrophoresis gave single protein band indicating that the enzyme preparation was pure. The molecular mass of the enzyme determined from SDS-PAGE analysis was 55.0 kDa. Using 2,6-dimethoxyphenol, 2,2′[azino-bis-(3-ethylbonzthiazoline-6-sulphonic acid) diammonium salt] and 3,5-dimethoxy-4-hydroxybenzaldehyde azine as the substrates, the K_m, k_cat and k_cat/K_m values of the laccase were found to be 420 μM, 13.04 s⁻¹, 3.11 × 10⁴ M⁻¹ s⁻¹, 225 μM, 13.03 s⁻¹, 1.3 × 10⁵ M⁻¹ s⁻¹ and 100 μM, 13.04 s⁻¹, 5.8 × 10⁴ M⁻¹ s⁻¹, respectively. The pH and temperature optima were 4.5 and 60 °C, respectively while pH and temperature stabilities were pH 4.5 and 50 °C. The activation energy for thermal denaturation of the enzyme was 18.6 kJ/mol/K. The purified laccase has yellow colour and does not show absorption band around 610 nm like blue laccases. The purified laccase transforms toluene, 3-nitrotoluene, 4-nitrotoluene, 3-chlorotoluene, 4-chlorotoluene and 3,4-dimethoxytoluene to benzaldehyde, 3-nitrobenzaldehyde, 4-nitrobenzaldehyde, 3-chlorobenzaldehyde, 4-chlorobenzaldehyde and 3,4-dimethoxybenzaldehyde in the absence of mediator molecules in high yields.