A determinação de enxofre em plantas pelo método quelatométrico do BDTA

Neste trabalho são relatados estudos a respeito do método quelatométrico de determinação de sulfato, baseado na titulação do bário residual com solução de EDTA, após a precipitação do citado ânion na forma de sulfato de bário. Foram estudados aspectos relativos ao emprego de diferentes indicadores e passou-se a preconizar o uso de solução de púrpura de ftaleína a 0,2%. São descritos experimentos relativos à determinação de sulfato em soluções puras, observando-se o período de precipitação e a influência de diversos ânions. Finalmente, tendo por base os dados obtidos, é preconizada urna marcha analítica para determinação de enxôfre total e enxôfre-sulfato em material vegetal. Os resultados obtidos quando se empregou a técnica proposta na determinação de sulfato em materiais vegetais, foram comparados com aquêles fornecidos pelo método gravimétrico, baseado na pe-sagem do sulfato de bário.

Determinação do teor total e do teor solúvel, em diversas soluções, do cobre do solo

O presente trabalho tem por objetivo o estudo de métodos e técnicas de determinação do teor total e do teor solúvel, em diversas soluções extratoras, do cobre do solo, Foram utilizados cinco diferentes solos do Estado de São Paulo e as determinações foram feitas pelo método colorimétrico do dietilditiocarbamato de sódio (DDC-Na). Para a determinação do teor total de cobre, os extratos dos solos foram preparados através de ataque de 250 mg de solo com os ácidos HClO4 e HF, 0 teor de cobre total encontrado oscilou de 23 a 126 ppm, para os cinco solos estudados. Quanto ao teor de cobre solúvel, foram empregados como extratores, soluções de HCl 0,05 N e 0,10 N, de EDTA dissódico a 1% e de CH3COOH 0,10 N. As extrações com as soluções de HCl foram conduzidas por agitação de 2,5 e 5,0 g de solo por 50 ml de solução, com a duração de 10, 15 e 30 minutos. Os teores de cobre solubilizado oscilaram de 0,5 a 14,6 ppm, com HCl 0,10 N e de 0,3 a 10,2 ppm, com HCl 0,05 N. Os extratos em soluções de EDTA dissódico e CH3COOH foram obtidos a partir de 5,0 g de solo por 50 ml de solução, agitando também 10, 15 e 30 minutos. Os teores de cobre solubilizado variaram de 0,8 a 15,0 ppm, com solução de EDTA, e de 0,0 a 0,5 ppm, com solução de CH3COOH.

Extração e determinação do teor solúvel de zinco do solo

Foram estudadas diversas soluções extratoras e várias técnicas para extração do zinco solúvel do solo. Foram utilizadas cinco amostras de solos do Estado de São Paulo e as determinações foram feitas empregando-se o método baseado na espectrofotometria de chama de absorção atômica. Para extração do zinco solúvel foram empregadas as seguintes soluções: HCl 0,05 e 0,10N; CH3COOH 0,10N; EDTA dissódico a 1%; MgCl2 0,10 e 0,50N; H2SO4 0,05 e 0,10N. As extrações com as soluções de HCl, CH3COOH, EDTA dissódico e MgCl2 , foram conduzidas por agitação de 2,5 e 5,0 g de solo com 50 ml de solução, durante 10, 15 e 30 minutos. Com as soluções de H2SO4, as extrações foram feitas apenas a partir de 5,0 g de solo para 50 ml de solução, conservando-se as demais condições. Os dados obtidos revelaram que de um modo geral as soluções de EDTA a 1%, de HC1 0,05 e 0,10N, e de H2S0(4) 0,05 e 0,10N, foram mais eficientes na extração do zinco do que as soluções de MgCl2 0,10 e 0,50N e de CH3COOH 0,10N. O tempo de agitação para extração do zinco durante 15 minutos forneceu resultados mais elevados (significativos ao nível de 5% de probabilidade) do que os obtidos com 10 minutos. No entanto a agitação durante 30 minutos apresentou resultados que não diferiram dos conseguidos com 15 minutos. A proporção de 2,5:50 (pêso da terra em gramas para volume em ml de solução extratora) apresentou resultados mais elevados, e significativos ao nível de 5%, do que os obtidos com a proporção de 5:50.

Influência de alguns compostos químicos na retenção do ácido ascórbico em frutas liofilizadas"

No presente trabalho foram estudados os efeitos do EDTA, do metabissulfito de potássio e de metabissulfito + ácido tartárico na retenção do ácido ascórbico durante a liofilização e posterior armazenamento de cereja das Antilhas e de morango. Os resultados e as análises estatísticas mostraram que todos os tratamentos foram significativamente superiores às testemunhas, observando-se também que o tratamento metabissulfito de potássio + ácido tartárico foi ligeiramente melhor que os demais tratamentos, porém não estatisticamente significativo.

O emprego dos testes microbiológicos na extração do zinco em solos de Piracicaba

Foram feitas avaliações do zinco no solo através do teste microbiológico do Aspergillus nigêr (WALLACE, 1961) em amostras de solo colhidas dos horizontes de 8 perfis de solos de 8 séries de solos do município de Piracicaba, Estado de São Paulo, Brasil. De cada sub horizonte foram tomados 0,25 g de solo onde se adicionou 50 ml de solução nutritiva sem zinco deixando-se em incubação em estufa adequada, à temperatura de 28° C, por um período de 7 dias. Passado este período colheu-se o micélio produzido e pesou-se após ser sêco a 70 - 80° C. Estes testes foram comparados com provas em branco e soluções (adições) contendo doses diferentes de zinco. No ensaio microbiológico fez-se, também, em amostras correspondentes ao Ap dos solos, a aplicação de doses crescentes de zinco (0 a 16 microgramas de zinco), nas mesmas condições usadas do teste, para se verificar as reações de cada solo à adição do zinco. Nas mesmas amostras colhidas dos perfis determinou-se em extratos, zinco solúvel, respectivamente de HCl 0,1 N de EDTA-(NH4)2CO3 e de Ditizona-acetato de amônio. Foram estudadas as correlações entre os resultados do Zinco das soluções extratoras e do teste microbiológico. O trabalho permitiu as seguintes conclusões: - O teste microbiológico do Aspergillus niger revelou-se eficiente na avaliação do zinco do solo. - Das 3 solúveis extratoras apenas a ditizona mostrou correlação com o teste microbiológico. - O teste microbiológico permitiu separar os solos em 3 grupos; segundo a sua reação a aplicação de zinco: Bem suprida em Zinco "Luiz de Queiroz"; medianamente supridas: Quebra-Dente, Bairrinho e Lageadinho; mal supridas: Iracema, Monte Olimpo, Guamium e Paredão Vermelho.

Estudos sobre a esporulação de uma amostra de bacillus: IV - Outras evidências sobre a atividade do íon Mn[2+] na esporulação endotrófica

Foram feitas experimentações om o intuito de se buscar mais evidências sobre a participação do íon Mn[2+] no mecanismo esporogenético de uma amostra de Bacillus licheniformis. Quando as formas vegetativas desta bactéria eram depositadas em um meio mineral, carente de fonte de carbono utilizável, em conjunto com um agente seqüestrante de metais como EDTA, a esporulação endotrófica deixava de ocorrer. Entretanto, a esporulação pôde ser protegida quando as células eram previamente saturadas com um excesso de Mn[2+] exógeno. As formas esporuladas obtidas nas condições estudadas mostraram termorresistência a 85ºC durante 20 minutos.

Contribution to the study of immune hemolysis by toad complement

EA (sheep erythrocytes carrying rabbit antibody) are lysed by toad complement under optimal conditions which include a low concentration of cells (1.54 x 10*8/ml), a low temperature of incubation (30°C) and the same amounts of Ca++ and Mg++ as required for the titration of guinea-pig complement. Kinetic studies of the role of cations mentioned above in immune lysis by toad C have disclosed a fundamental difference as compared to guinea-pig C. In a limited complement system, the lysis by amphibian C is completely blocked by EDTA, even when the chelating agent is added as late as 15 minutes after zero-time. Inhibition by EGTA is only partial and the findings suggest that Mg++ is required not only at the beginning, but also at late stages of the lytic process. It has been speculated that the activation of amphibian complement proceeds mainly by the alternative pathway.

High-throughput hydrophilic interaction chromatography coupled to tandem mass spectrometry for the optimized quantification of the anti-Gram-negatives antibiotic colistin A/B and its pro-drug colistimethate.

Colistin is a last resort's antibacterial treatment in critically ill patients with multi-drug resistant Gram-negative infections. As appropriate colistin exposure is the key for maximizing efficacy while minimizing toxicity, individualized dosing optimization guided by therapeutic drug monitoring is a top clinical priority. Objective of the present work was to develop a rapid and robust HPLC-MS/MS assay for quantification of colistin plasma concentrations. This novel methodology validated according to international standards simultaneously quantifies the microbiologically active compounds colistin A and B, plus the pro-drug colistin methanesulfonate (colistimethate, CMS). 96-well micro-Elution SPE on Oasis Hydrophilic-Lipophilic-Balanced (HLB) followed by direct analysis by Hydrophilic Interaction Liquid Chromatography (HILIC) with Ethylene Bridged Hybrid - BEH - Amide phase column coupled to tandem mass spectrometry allows a high-throughput with no significant matrix effect. The technique is highly sensitive (limit of quantification 0.014 and 0.006μg/mL for colistin A and B), precise (intra-/inter-assay CV 0.6-8.4%) and accurate (intra-/inter-assay deviation from nominal concentrations -4.4 to +6.3%) over the clinically relevant analytical range 0.05-20μg/mL. Colistin A and B in plasma and whole blood samples are reliably quantified over 48h at room temperature and at +4°C (<6% deviation from nominal values) and after three freeze-thaw cycles. Colistimethate acidic hydrolysis (1M H2SO4) to colistin A and B in plasma was completed in vitro after 15min of sonication while the pro-drug hydrolyzed spontaneously in plasma ex vivo after 4h at room temperature: this information is of utmost importance for interpretation of analytical results. Quantification is precise and accurate when using serum, citrated or EDTA plasma as biological matrix, while use of heparin plasma is not appropriate. This new analytical technique providing optimized quantification in real-life conditions of the microbiologically active compounds colistin A and B offers a highly efficient tool for routine therapeutic drug monitoring aimed at individualizing drug dosing against life-threatening infections.

Mean platelet volume in the early phase of acute ischemic stroke is not associated with severity or functional outcome.

Background: Previous studies reported an increase of mean platelet volume (MPV) in patients with acute ischemic stroke. However, its correlation with stroke severity has not been investigated. Moreover, studies on the association of MPV with functional outcome yielded inconsistent results. Methods: We included all consecutive ischemic stroke patients admitted to CHUV (Centre Hospitalier Universitaire Vaudois) Neurology Service within 24 h after stroke onset who had MPV measured on admission. The association of MPV with stroke severity (NIHSS score at admission and at 24 h) and outcome (Rankin Scale score at 3 and 12 months) was analyzed in univariate analysis. The chi(2) test was performed to compare the frequency of minor strokes (NIHSS score </=4) and good functional outcome (Rankin Scale score </=2) across MPV quartiles. The ANOVA test was used to compare MPV between stroke subtypes according to the TOAST classification. Student's two-tailed unpaired t test was performed to compare MPV between lacunar and nonlacunar strokes. MPV was generated at admission by the Sysmex XE-2100 automated cell counter (Sysmex Corporation, Kobe, Japan) from EDTA blood samples. Results: There was no significant difference in the frequency of minor strokes (p = 0.46) and good functional outcome (p = 0.06) across MPV quartiles. MPV was not associated with stroke severity or outcome in univariate analysis. There was no significant difference in MPV between stroke subtypes according to the TOAST classification (p = 0.173) or between lacunar and nonlacunar strokes (10.50 +/- 0.91 vs. 10.40 +/- 0.81 fl, p = 0.322). Conclusions: MPV, assessed within 24 h after ischemic stroke onset, is not associated with stroke severity or functional outcome.

Avaluació del filtrat glomerular en candidats a donant renal viu

En una població de pacients candidats a ser donant renal viu, la determinació acurada i precisa de la de la taxa de filtrat glomerular (TFG) és important ja que d’ella en depen que s’accepti o rebutji un canditat a donant renal viu. En aquest estudi es compara la determinació de la TFG calculada segons el mètode isotòpic amb 51Cr-EDTA amb la TFG calculada amb mètodes estimatius basats en substàncies endògenes com la creatinina i la cistatina.

Multiplex real-time PCR for the diagnosis of malaria: correlation with microscopy.

Malaria is generally diagnosed by microscopy and rapid antigen testing. Molecular methods become more widely used. In the present study, the contribution of a quantitative multiplex malaria PCR was investigated. We assessed: (i) the agreement between PCR-based identification and microscopy and (ii) the correlation between the parasite load as determined by quantitative PCR and by microscopy. For 83 patients positive by microscopy for Plasmodium spp., the first EDTA-blood sample was tested by multiplex PCR to confirm smear-based species identification. Parasite load was assessed daily using both microscopy and PCR. Among the 83 patients tested, one was positive by microscopy only and 82 were positive by microscopy and PCR. Agreement between microscopy and PCR for the identification at the species level was 89% (73/82). Six of the nine discordant results corresponded to co-infections by two or three species and were attributed to inaccurate morphological identification of mixed cases. The parasite load generally decreased rapidly after treatment had been started, with similar decay curves being obtained using both microscopy and PCR. Our PCR proved especially useful for identifying mixed infections. The quantification obtained by PCR closely correlated with microscopy-based quantification and could be useful for monitoring treatment efficacy, at least in clinical trials.

Protease activity in Giardia duodenalis trophozoites of axenic strains isolated from symptomatic and asymptomatic patients

We have examined by gelatin-SDS-PAGE the protease activity in cell lysates of Giardia duodenalis trophozoites of two axenic strains isolated in Brazil from a symptomatic patient (BTU-11) and an asymptomatic carrier (BTU-10), and the reference strain Portland 1 (P1). The proteolysis band patterns showed differences among strains isolated from asymptomatic and symptomatic individuals. The lysate of the strain BTU-10, showed only five hydrolysis bands, while a greater number of bands (10-11 bands) was seen in strains BTU-11 and P1. The protease activity in all lysates was inhibited by cysteine (E-64 and iodoacetamide) and serine proteases (TPCK and TLCK) inhibitors, but not by PMSF and EDTA. In general, the results revealed protease activities in G. duodenalis trophozoites of Brazilian axenic strains and the predominance of cysteine proteinases. It should be stressed the inter-strain difference in hydrolysis band patterns observed between strains isolated from symptomatic patients and the strain obtained from an asymptomatic carrier.

Kidney transplantation in patients with Fabry disease

Introduction and Aims: Fabry disease is an X-linked lysosomal storage disorder caused by absence or deficient activity of the lysosomal enzyme alpha-galactosidase A. Renal manifestations occur early in life in a significant proportion of children, in many women and in almost all men with Fabry disease. These manifestations ultimately progress to end-stage renal disease in nearly all males and in some female patients. Data on kidney transplantation in patients with Fabry disease who are receiving enzyme replacement therapy (ERT), however, are scarce. Methods: We examined the clinical characteristics of kidney transplant recipients (KTRs) in the Fabry Outcome Survey (FOS) - a European database of patients with Fabry disease that was established to monitor the safety and outcome of ERT. Results: Of the 752 patients enrolled in FOS up to October 2005, 34 (4.5%) were reported to be KTRs. The mean age of these 32 male and 2 female patients was 45 ± 9 years, the median time since the transplant was 9 years, the median estimated glomerular filtration rate (eGFR) was 46 mL/min/1.73 m2 and the median level of proteinuria was 180 mg/24 hours. ERT was well tolerated, with mild infusion-related reactions reported in only one patient. Amongst these patients, 53% were reported to have hypertension, 71% left ventricular hypertrophy, 27% cardiac valve disease and 27% arrhythmia. A total of 23 (68%) of the patients (1 female, 22 males) were receiving ERT with agalsidase alfa (Replagal; Shire Human Genetic Therapies, UK), with a median duration of treatment of 2.5 years. There were no differences in age or time since transplantation between treated and untreated patients. The median eGFRs were 46 and 49 mL/min/1.73 m2 and the median levels of proteinuria were 200 and 160 mg/24 hours, respectively. Conclusions: KTRs represent a significant minority of individuals enrolled in a large international registry of patients with Fabry disease (FOS). Approximately two-thirds of KTRs with Fabry disease enrolled in FOS receive ERT with agalsidase alfa, which is well tolerated. Comparison of treated and untreated patients has the potential to examine effects of ERT on the progression of renal and cardiovascular disease.

Effect of groundwater composition on arsenic detection by bacterial biosensors.

A luminescent bacterial biosensor was used to quantify bioavailable arsenic in artificial groundwater. Its light production above the background emission was proportional to the arsenite concentration in the toxicologically relevant range of 0 to 0.5 mu M. Effects of the inorganic solutes phosphate, Fe(II) and silicate on the biosensor signal were studied. Phosphate at a concentration of 0.25 g L-1 phosphate slightly stimulated the light emission, but much less than toxicologically relevant concentrations of the much stronger inducer arsenite. No effect of phosphate was oberved in the presence of arsenite. Freshly prepared sodium silicate solution at a concentration of 10 g L-1 Si reduced the arsenite-induced light production by roughly 37%, which can be explained by transient polymerization leading to sequestration of some arsenic. After three days of incubation, silicate did not have this effect anymore, probably because depolymerization occurred. In the presence of 0.4 g L-1 Fe(II), the arsenite-induced light emission was reduced by up to 90%, probably due to iron oxidation followed by arsenite adsorption on the less soluble Fe(III) possibly along with some oxidation to the stronger adsorbing As(V). Addition of 100 mu M EDTA was capable of releasing all arsenic from the precipitate and to transform it into the biologically measurable, dissolved state. The biosensor also proved valuable for monitoring the effectiveness of an arsenic removal procedure based on water filtration through a mixture of sand and iron granules.

Efficacy of benznidazol treatment for asymptomatic chagasic patients from state of Rio Grande do Sul evaluated during a three years follow-up

The efficacy of benznidazol on the treatment of chagasic patients from the state of Rio Grande do Sul was evaluated during a three-year follow-up. A cohort of 80 asymptomatic chronic chagasic patients or blood bank donors (49 male and 31 female) was studied. Their ages varied from 17-42 years, with a mean and a median of 30 and 35 years, respectively. The 80 patients presented positive serology, hemoculture and polymerase chain reaction (PCR). They were treated with 5 mg/Kg benznidazol twice a day for 60 days. Serological, parasitological and PCR methods were used to evaluate response. Serology was performed using commercial ELISA and indirect immunofluorescence (IFI) tests, parasitemia was monitored by hemoculture in LIT medium and PCR with primers S35/S36 was used to amplify a Trypanosoma cruzi 330 bp kDNA repetitive sequence. PCR positivity of 240 seropositive individuals was compared using DNA preparations from whole blood/guanidine EDTA (GE), buffy-coat/GE and frozen buffy-coat. Fifty non-chagasic individuals were used as negative controls. PCR positivity was 86.7% for the frozen buffy-coat, 71.7% for the GE/buffy-coat and 69.2% for the GE/whole blood. The hemocultures became negative just after treatment and remained negative during the three years of follow-up. In the third year after treatment, 9/80 (11.3%) patients presented negative PCR and, from those, four also presented negative serological tests. Furthermore, a reduction in three serological titers was observed in 27/80 (33.8%) of the patients treated. Taken together, the results show that four of the 80 (5.0%) chronic chagasic patients from the state of Rio Grande do Sul were cured after treatment with benznidazol.