Fluorometric fiber optic drop sensor for atmospheric hydrogen sulfide

A fluorometric technique based on a liquid drop excited from its interior by an optical fiber is described for the measurement of low concentrations of atmospheric hydrogen sulfide (H2S). A drop of alkaline fluorescein mercuric acetate (FMA) solution is suspended in a flowing air sample stream and serves as a renewable sensor. An optical fiber contained within the conduit that forms the drop, brings in the excitation beam; the fluorescence emission is measured by an inexpensive photodiode positioned close to the drop. As H2S in the sample is collected by the alkaline drop, it reacts rapidly with FMA resulting in a significant decrease in fluorescence intensity, proportional to the concentration of H2S sampled. The chemistry of this uniquely selective reaction has been well established for many years, the present technique permits a simple fast inexpensive near real-time measurement with very little reagent consumption. Even without prolonged sampling/preconcentration steps, limits of detection (LODs) in the double digit ppbv range is readily attainable. (C) 1997 Elsevier B.V. B.V.

Electrochemical behavior of the N-nosyl-protected amino acids in N,N-dimethylformamide

The electrochemical reduction of serine, glycine, and leucine protected by the 4-nitrobenzenesulfonyl, group in N,N-dimethylformamide at mercury cathode occurs at two steps. The first one at -0.8 V vs. SCE, after a one-electron transfer, leads the anion radical formation that dimerizes and adsorbs at electrode. In the second step at -1.4 V, an instable dianion forms which then cleaves. The mechanism is discussed.

THE CATHODIC CLEAVAGE OF THE NITROBENZENESULFONYL GROUP FROM ALIPHATIC-AMINES IN N,N-DIMETHYLFORMAMIDE

The reduction of benzenesulfonyl derivatives of n-butylamine and N,N-di-n-butylamine with nitro substituents at the 2, 3 and 4 positions of the phenyl ring in N,N-dimethylformamide is reported. The N,N-di-n-butyl-4- and N-n-butyl-2-nitrobenzenesulfonamides are reduced in two cathodic steps. The first one, at about -0.90 V vs. SCE, a reversible one-electron process, gives a stable anion radical. The second reduction step at -1.70 V vs. SCE leads to cleavage of the S-N bond in good yields (> 70%). It is shown that the reduction of the N-n-butyl-3- and N-n-butyl-4-nitrobenzenesulfonamide is different, with three reduction steps. The first reduction step occurs with the formation of an unstable anion radical, which decomposes via N-H bond cleavage. The reduction of this sulfonamide anion occurs at -1.16 V vs. SCE and the third cathodic step arises at -1.70 V vs. SCE when the remaining radical anion is reduced to its dianion. The S-N bond cleavage is rapid but is always a minor process. The mechanisms of the reduction are discussed.

Electroreduction of benznidazole in dimethylsulfoxide

The electrochemical behavior of benznidazole has been investigated in dimethylsulfoxide by cyclic voltammetry and controlled-potential electrolysis. The reduction occurs in two one-electron steps, where the first electron transfer corresponds to the reversible formation of the radical anion followed by a slow chemical reaction. The second electron transfer is attributed to the reduction of the radical anion to a dianion by an electrodic process involving a Very fast cleavage of the dianion with the formation of a lactam derivative as the principal product of reduction in aprotic medium. (C) 2001 the Electrochemical Society. All rights reserved.

The cathodic deprotection of the nitrobenzoyl group from phenyl nitrobenzoates in N,N-dimethylformamide

The reduction of phenyl benzoates with nitro substituents at the 2-,3- and 4-positions of the benzoates in N,N-dimethylformamide is reported. The phenyl 4- and 3-nitrobenzoate are reduced in two cathodic steps. The first one, at about -0.9 V vs. SCE, a reversible one-electron process, gives a rather stable anion radical. The second reduction step at potentials between -1.5 and -2.0 V vs. SCE leads to formation of the dianion, which decomposes giving free phenol in good yields (> 80%). on the other hand, the phenyl 2-nitrobenzoate is reduced in one cathodic step. This step occurs at -0.9 V with formation of an unstable anion radical which decomposes via C-O bond cleavage, giving phenol with a yield of ca. 80%. The mechanisms of the reduction of these compounds are discussed. (C) 1997 Elsevier B.V. S.A.

Role of roscovitine and IBMX on kinetics of nuclear and cytoplasmic maturation of bovine oocytes in vitro

The 3-isobutyl-1-methylxanthine (IBMX) is able to prevent resumption of meiosis by maintaining elevated cyclic AMP (cAMP) concentrations in the oocyte, and roscovitine, a purine known to specifically inhibit MPF kinase activity, maintains bovine oocytes at the germinal vesicle (GV) stage. The present study was conducted to analyze whether cytoplasmic maturation (examined by the pattern of cortical granule (CG) distribution) of bovine oocytes is improved during meiotic arrest with IBMX and roscovitine. Oocytes were matured in vitro in a 10% Knockout(SR) supplemented TCM-199 medium (Control) with either 0.5 mM IBMX or 25 mu M roscovitine (ROSC). Oocytes were stained with fluorescein isothiocyanate conjugated Lens culinaris agglutinin (FITC-LCA) for CG evaluation and with Hoechst 33342 for nuclear stage assessment. At 16 h of culture, the percentage of oocytes remaining in the GV stage was higher (P < 0.05) in the ROSC group (32.41%) compared with the Control and IBMX groups (8.61% and 9.73%, respectively). At 24h of culture, progression of meiosis to M II stage was retarded (P < 0.05) in the ROSC group (24.05%) compared to the Control (60.20%), whereas the IBMX group (33.88%) showed no significant difference to the other two groups. At 16h of maturation, the proportion of oocytes with CG in clusters (immature cytoplasm) was similar between the groups, as was the percentage of peripheral CG (mature) at 24h of maturation. The results of the present study demonstrated that the meiotic inhibitors IBMX and roscovitine delay the progression of nuclear maturation without affecting cytoplasmic maturation, assessed by the analysis of CG repositioning. (c) 2006 Elsevier B.V. All rights reserved.

The electrochemical cleavage of the nitrobenzoyl group from butyl nitrobenzoates in N,N-dimethylformamide

The applicability of the nitrobenzoyl group [NO2C6H4CO-] to protecting the functional hydroxyl group was investigated through study of the electrochemical behaviour of the butyl 4-, 3- and 2-nitrobenzoate compounds. These isomers are reduced in two cathodic steps. The first, at potentials of ca. -0.9 V vs. SCE, is attributed to the formation of rather stable anion radicals, involving one-electron transfer. The second, at potentials of ca. -1.7 V vs. SCE, occurs with a two-electron transfer in an ECE process, in which the dianion produced undergoes scission of the C-O bond giving n-butanoate ions with high yields (similar to 80%)

Influence of membrane-solution interface on the selectivity of SnO2 ultrafiltration membranes

SnO2 supported membranes, presenting 3.0 nm average pore size, have been produced by sol casting on alumina tubular substrate using aqueous colloidal suspensions prepared by sol-gel route. The selectivity and flux throughout SnO2 membrane were analyzed by permeation experiments, using a laboratory tangential filtration pilot equipped with a monotubular membrane. To evaluate the effect of the surface charge at the membrane-solution interface, aqueous salt solutions (NaCl, Na2SO4, CaCl, and CaSO4) of different ionic strength have been filtered and the results correlated with the values of zeta potential measured at several pH. The results show that the retention coefficient is dependent on the electrolyte present in aqueous solution decreasing as: (dication, monoanion) > (monocation, monoanion) approximate to (monocation, dianion) > (dication, dianion). The surface charge and the cation adsorption capacity play a determinant role in these selectivity sequences. (C) 2001 Elsevier B.V. B.V. All rights reserved.

Panretinal photocoagulation versus PRP plus intravitreal bevacizumab for high-risk proliferative diabetic retinopathy (IBeHi study)

Purpose: To evaluate the effects of panretinal photocoagulation (PRP) compared with PRP plus intravitreal bevacizumab on best corrected visual acuity (BCVA) and total area of fluorescein leakage from active new vessels (NVs) in patients with high-risk proliferative diabetic retinopathy (PDR).Methods: We carried out a prospective study of patients with high-risk PDR and no prior laser treatment who were randomly assigned to receive PRP (PRP group) or PRP plus intravitreal injection of 1.5 mg of bevacizumab (PRP-plus group). In all patients, the PRP was administered at two time-points (weeks 1 and 3), with the intravitreal bevacizumab delivered at the end of the second laser episode in the PRP-plus group. Standardized ophthalmic evaluation including Early Treatment Diabetic Retinopathy Study BCVA as well as stereoscopic fundus photography and fluorescein angiography were performed at baseline and at weeks 4, 9 (+/- 1) and 16 (+/- 2). Main outcome measures included changes in BCVA and in total area of fluorescein leakage from active NVs.Results: Twenty-two (n = 30 eyes) consecutive patients completed the 16-week follow-up. There was no significant difference between the PRP and PRP-plus groups with respect to age, gender, type or duration of diabetes, area of fluorescein leakage from active NVs or BCVA. No significant difference in BCVA was observed between the groups throughout the study period. However, the total area of actively leaking NVs was significantly reduced in the PRP-plus group compared with the PRP group at weeks 4, 9 and 16 (p < 0.001). No major adverse events were identified.Conclusions: In the short-term, the adjunctive use of intravitreal bevacizumab with PRP was associated with a greater reduction in the area of active leaking NVs than PRP alone in patients with high-risk PDR.

Sex determination of murine and bovine embryos using cytotoxicity and immunofluorescence assays

Murine and bovine embryos were sexed by using polyclonal and monoclonal anti-H-Y antibodies in cytotoxic and immunofluorescence assays. The specificity of 3 different sources of antibodies are compared. Sexing was accomplished by complement-mediated cytotoxicity or by H-Y antigen detection using a anti-mouse Ig fluorescein conjugate. Cytogenetic ascertainment for correct sex identification yielded a score of about 85% in both species, while the immunofluorescence assay yielded a score of 83%. © 1993.

Effect of α-tocopherol, taurine and selenium on the attenuation of ischemia/reperfusion injury of splanchnic organs

Background: Splanchnic artery occlusion shock is caused by increased capillary permeability and cellular injury precipitated by oxygen derived free radicals following ischemia and reperfusion of splanchnic organs. The purpose of this study was to assess the role of several well-known oxygen- derived free radical scavengers in ameliorating or preventing this syndrome. Study design: Anesthetized rats were subjected to periods of occlusion of the visceral arteries and reperfusion. Tocopherol, taurine, selenium or a 'cocktail' of these three agents was injected subcutaneously for 4 consecutive days prior to operation. Mean arterial blood pressure was measured throughout the experimental period. Fluorometry and technetium-99m pyrophosphate counting of the visceral organs were performed as well as a histologic grading system for intestinal viability. Results: Final mean arterial blood pressure associated with the 'cocktail' and selenium groups was 79.1 ± 27.4 mmHg and 83.6 ± 17.8 mmHg, respectively. These values were significantly higher than the control group, 40.8 ± 11.4 mmHg (P < 0.05). Similar patterns of the benefit of selenium in contrast with the other groups were obtained with fluorescein perfusion, radioisotopic activity and histologic analysis. Conclusion: Pretreatment with selenium of splanchnic ischemia and reperfusion in the rat improves mean arterial blood pressure and microcirculatory visceral perfusion. Further analysis of the precise protective mechanism of selenium for reperfusion injury will enable visceral organs to withstand the consequences of increased capillary leakage and oxidant injury.

Evaluation of homologous, heterologous, and affinity conjugates for the serodiagnosis of Toxoplasma gondii and Neospora caninum in maned wolves (Chrysocyon brachyurus)

Use of serological tests in the diagnosis of infectious diseases in wild animals has several limitations, primarily the difficulty of obtaining species-specific reagents. Wild canids, such as maned wolves (Chrysocyon brachyurus), are highly predisposed to infection by Toxoplasma gondii and, to a lesser extent, to Neospora caninum. The aim of the present study was to evaluate homologous, heterologous, and affinity conjugates in enzyme-linked immunosorbent assays (ELISAs) and indirect fluorescent antibody tests (IFATs) for detecting immunoglobulin (Ig) G antibodies against T. gondii and N. caninum in maned wolves. Serum samples were obtained from 59 captive animals in Brazil and tested by ELISA for T. gondii serology and IFAT for N. caninum serology using 3 different enzymatic and fluorescent conjugates: homologous (guinea pig anti-maned wolf IgG-peroxidase and -fluorescein isothiocyanate [FITC]), heterologous (rabbit anti-dog IgG-peroxidase and -FITC), and affinity (protein A-peroxidase and -FITC). Seropositivity to T. gondii was comparable among the homologous (69.5%), heterologous (74.6%), and affinity (71.2%) enzymatic conjugates. A significant positive correlation was found between the antibody levels determined by the 3 enzymatic conjugates. The highest mean antibody levels (ELISA index = 4.5) were observed with the protein A-peroxidase conjugate. The same seropositivity to N. caninum (8.5%) was found with the homologous and heterologous fluorescent conjugates, but protein A-FITC was not able to detect or confirm any positive samples with homologous or heterologous conjugates. Our results demonstrate that homologous, heterologous, and affinity conjugates might be used in ELISA for serological assays of T. gondii in wild canids, whereas for N. caninum infection, only the homologous or heterologous fluorescent conjugates have been shown to be useful. © American Society of Parasitologists 2005.

In vitro susceptibility of oral Candida albicans strains to different pH levels and calcium hydroxide saturated aqueous solution

Candida albicans is present in the oral cavity and in the whole digestive tract of humans and other animals, being frequently related to endodontic treatment failure. The present study determined the incidence of C. albicans in the oral cavity and the susceptibility of isolates to different pH values and saturated calcium hydroxide aqueous solution at pH 12.5. Sixty-five patients attending the Endodontic Clinic at the Sagrado Coração University participated in the study. The collected samples were cultivated in selective media for C. albicans and the isolates were tested in terms of resistance to both alkaline pH and saturated aqueous solution of calcium hydroxide. In relation to time variables, yeast viability was assessed by the Sabouraud's agar culture and fluorescein diacetate and ethidium bromide fluorescent staining method. Results from the different pHs and experimental times, including those from different techniques measuring fungal viability, were compared using the chi-square and Fisher's exact tests (α=0.05). The yeasts became completely inviable after 48 h of contact with the calcium hydroxide solution. On the other hand, when exposed to the alkaline culture broth, the yeasts were found to be viable at pHs 9.5 and 10.5 for up to 7 days. In conclusion, C. albicans can only be completely inhibited by direct contact with saturated calcium hydroxide aqueous solution after 48 h of exposure.

Bothrops leucurus venom induces nephrotoxicity in the isolated perfused kidney and cultured renal tubular epithelia

Bites from snake (Bothrops genus) cause local tissue damage and systemic complications, which include alterations such as hemostatic system and acute renal failure (ARF). Recent studies suggest that ARF pathogenesis in snakebite envenomation is multifactorial and involves hemodynamic disturbances, immunologic reactions and direct nephrotoxicity. The aim of the work was to investigate the effects of the Bothrops leucurus venom (BlV) in the renal perfusion system and in cultured renal tubular cells of the type MDCK (Madin-Darby Canine kidney). BlV (10 μg/mL) reduced the perfusion pressure at 90 and 120 min. The renal vascular resistance (RVR) decreased at 120 min of perfusion. The effect on urinary flow (UF) and glomerular filtration rate (GFR) started 30 min after BlV infusion, was transient and returned to normal at 120 min of perfusion. It was also observed a decrease on percentual tubular transport of sodium (%TNa+) at 120 min and of chloride (%TCl-) at 60 and 90 min. The treatment with BlV caused decrease in cell viability to the lowest concentration tested with an IC50 of 1.25 μg/mL. Flow cytometry with annexin V and propidium iodide showed that cell death occurred predominantly by necrosis. However, a cell death process may involve apoptosis in lower concentrations. BlV treatment (1.25 μg/mL) led to significant depolarization of the mitochondrial membrane potential and, indeed, we found an increase in the expression of cell death genes in the lower concentrations tested. The venom also evoked an increase in the cytosolic Ca2+ in a concentration dependent manner, indicating that Ca2+ may participate in the venom of B. leucurus effect. The characterization of the effects in the isolated kidney and renal tubular cells gives strong evidences that the acute renal failure induced by this venom is a result of the direct nephrotoxicity which may involve the cell death mechanism. © 2012.

Cryoprotective effect of different glycerol concentrations on domestic cat spermatozoa

Cryopreservation of spermatozoa is a pivotal tool in assisted reproduction, and studies aiming to establish optimal freezing/thawing protocols are essential to enhance sperm survival. The objectives of the present study were to (1) compare the cryoprotective efficiency of three different glycerol concentrations (3%, 5%, and 7%) on the basis of post-thaw sperm quality and (2) investigate whether the incidence of morphologically abnormal sperm in fresh samples is related to cryodamage sensitivity. Semen was collected from six tomcats using an artificial vagina (total 18 ejaculates). Each ejaculate was diluted using Tris-egg yolk-based extender (TEY), evaluated, equally divided into three aliquots, and rediluted using TEY with and without glycerol to achieve final concentrations of 3%, 5%, and 7%. Samples were loaded into 0.25 mL straws, equilibrated for 60 minutes at 5 °C, frozen, and then thawed at 46 °C for 12 seconds. Fresh and frozen-thawed samples were evaluated for sperm motion parameters (computer-assisted sperm analysis), plasma membrane integrity (PMI; propidium iodide and carboxyfluorescein diacetate), and DNA integrity (acridine orange). Plasma and acrosomal membrane integrity were assessed by flow cytometry (propidium iodide and fluorescein isothiocyanate-conjugated pea (Pisum sativum) agglutinin) immediately after thawing. Sperm motion parameters were also evaluated at 30 and 60 minutes of postincubation. For all treatment groups, cryopreservation significantly impaired the PMI and sperm motion parameters, except for straightness and amplitude of lateral head displacement. DNA integrity showed a slight reduction (P < 0.05) when 3% glycerol was used. The percentage of total motility, progressive motility, and rapid spermatozoa were significantly lower immediately after thawing and up to 60 minutes of incubation for the 3% glycerol group when compared with 5% and 7%. No difference (P > 0.05) was found for PMI, acrosome integrity, and DNA integrity among post-thaw groups. However, higher (P < 0.05) incidence of viable cells with reacted acrosome and dead cells with intact acrosome were observed with 7% and 3% glycerol, respectively. Percentage of morphologically abnormal spermatozoa in fresh sample was positively correlated with PMI only in the 3% glycerol group and negatively correlated with sperm motility in the 5% and 7% groups. In conclusion, the final concentration of 5% glycerol offered better cryoprotective effect for ejaculated cat sperm, and the relationship found between prefreezing sperm morphology and post-thaw sperm quality showed to be dependent on final glycerol concentration. © 2013 Elsevier Inc.