3,3',5,5'-Tetramethylbenzidine in hypochlorous acid and taurine chloramine scavenging assays: interference of dimethyl sulfoxide and other vehicles.

This article shows how six vehicles interfere in the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by hypochlorous acid (HOCl) and taurine chloramine in vitro. All the tested vehicles inhibited TMB oxidation by HOCl; dimethyl sulfoxide had a remarkable effect at concentrations as low as 0.00005% (v/v). Cremophor EL and ethanol inhibited TMB oxidation by taurine chloramine at concentrations higher than 0.05 and 25% (v/v), respectively; the other vehicles did not affect this reaction. The results will help to guide the choice of solvent for the TMB oxidation assay performed under viable experimental conditions for evaluation of the HOCl and taurine chloramine scavenging ability of drugs.

Superoxide anion production by neutrophils is associated with prevalent clinical manifestations in systemic lupus erythematosus.

To determine the relation between neutrophil function and the clinical characteristics of systemic lupus erythematosus (SLE), the superoxide anion (O2-) production by neutrophils, mediated by FcgammaR and FcgammaR/CR cooperation, was studied in 64 SLE patients classified according to their prevalent clinical manifestations. Three clinically distinct patterns were designated: (1) manifestations associated with the occurrence of cytotoxic antibodies (SLE-I group); (2) manifestations associated with circulating immune complexes (IC; SLE-II group), and (3) manifestations associated with IC and cytotoxic antibodies (SLE-III group). O2- production was evaluated by a lucigenin-dependent chemiluminescent assay in neutrophils stimulated with IC-IgG opsonized or not with complement. No difference in O2- production was observed when neutrophil responses from healthy controls were compared to the unclassified patients. However, when the SLE patient groups were considered, the following differences were observed: (1) SLE-I neutrophils showed lower O2- production mediated by the IgG receptor (FcgammaR) with the cooperation of complement receptors (FcgammaR/CR) than observed in the SLE-II, SLE-III, and healthy groups; (2) neutrophils from the SLE-II group showed a decreased [Formula: see text] production mediated by FcgammaR/CR compared to the SLE-III group, (3) SLE-III neutrophils produced more O(2)(-) than neutrophils from the SLE-II and control groups, and (4) CR showed inefficiency in mediating the O2- production by neutrophils from the SLE-I group. Comparative experiments on the kinetics of chemiluminescence (CL; Tmax and CLmax) disclosed differences only for the SLE-I group. Taken together, these results suggest that differences in oxidative metabolism of neutrophils mediated by FcgammaR/CR may reflect an acquired characteristic of disease associated with distinct clinical manifestations.

Inhibition of immune complex-mediated neutrophil oxidative metabolism: a pharmacophore model for 3-phenylcoumarin derivatives using GRIND-based 3D-QSAR and 2D-QSAR procedures.

In this study, twenty hydroxylated and acetoxylated 3-phenylcoumarin derivatives were evaluated as inhibitors of immune complex-stimulated neutrophil oxidative metabolism and possible modulators of the inflammatory tissue damage found in type III hypersensitivity reactions. By using lucigenin- and luminol-enhanced chemiluminescence assays (CL-luc and CL-lum, respectively), we found that the 6,7-dihydroxylated and 6,7-diacetoxylated 3-phenylcoumarin derivatives were the most effective inhibitors. Different structural features of the other compounds determined CL-luc and/or CL-lum inhibition. The 2D-QSAR analysis suggested the importance of hydrophobic contributions to explain these effects. In addition, a statistically significant 3D-QSAR model built applying GRIND descriptors allowed us to propose a virtual receptor site considering pharmacophoric regions and mutual distances. Furthermore, the 3-phenylcoumarins studied were not toxic to neutrophils under the assessed conditions.

Neutrophil effector functions triggered by Fc-gamma and/or complement receptors are dependent on B-ring hydroxylation pattern and physicochemical properties of flavonols.

Tissue damage in autoimmune diseases involves excessive production of reactive oxygen species (ROS) triggered by immune complexes (IC) and neutrophil (PMN) interactions via receptors for the Fc portion of IgG (FcgammaR) and complement receptors (CR). Modulation of both the effector potential of these receptors and ROS generation may be relevant to the maintenance of body homeostasis. In the present study, the modulatory effect of four flavonols (myricetin, quercetin, kaempferol, galangin) on rabbit PMN oxidative metabolism, specifically stimulated via FcgammaR, CR or both classes of receptors, was evaluated by luminol- and lucigenin-dependent chemiluminescence assays. Results showed that flavonol inhibitory effect was not dependent on the cell membrane receptor class stimulated but related to the lipophilicity of the compounds (their apparent partition coefficient values were obtained by high-performance liquid chromatography), and was also inversely related to the number of hydroxyl groups in the flavonol B ring and the ROS-scavenger activity (assessed by the luminol--H2O2--horseradish peroxidase reaction). Under the experimental conditions the flavonols tested were not toxic to PMNs (evaluated by lactate dehydrogenase release and trypan blue exclusion) and did not interfere with IC-induced phagocytosis (evaluated by transmission electron microscopy). Our results suggested that inhibition of IC-stimulated PMNs effector functions by the flavonols tested herein was the result of cooperation of different cellular mechanisms.

Antibacterial screening of anthocyanic and proanthocyanic fractions from cranberry juice.

The effect of anthocyanin- and proanthocyanidin-rich fractions isolated from cranberry juice was studied for their antibacterial activity against nine bacterial strains. Activity was assessed by the agar diffusion assay. Staphylococcus aureus ATCC 6538 was the only strain to exhibit some susceptibility to four out of 10 anthocyanin-rich fractions tested. A variable susceptibility of S. aureus, Enterococcus faecalis ATCC 10541, and Micrococcus luteus ATCC 9341 to proanthocyanidin- rich fractions was also observed. Streptococcus mutans strains as well as Escherichia coli ATCC 10538 and Pseudomonas aeruginosa ATCC 27853 were not susceptible to any of the cranberry juice samples or fractions at the tested concentrations. There was no clear correlation between Gram-positive or Gram-negative bacterial susceptibility to cranberry juice. In this work, the role of cranberry juice anthocyanic and proanthocyanic fractions upon bacterial viability is discussed.

A simple method to study the activity of natural compounds on the chemiluminescence of neutrophils upon stimulation by immune complexes.

INTRODUCTION: Neutrophils (PMNs) are the main effector cells involved in the immune response to microorganisms. However, in various noninfectious states, such as autoimmune and immune complex (ICs) diseases, ICs are found to be deposited in various organs, leading to recruitment and activation of PMNs at these sites of deposition. Consequently, reactive oxygen species (ROS) and lysosomal enzymes are extensively released by activated PMNs into the extracellular milieu, leading to host tissue injury. METHODS: In the present study, we discuss some experimental conditions of a luminol-enhanced chemiluminescence (LECL) assay to study the effect of natural compounds on the production of ROS by rabbit PMNs stimulated with precipitated ICs. Moreover, we evaluated the activities of quercetin and 7-allyloxycoumarin on this ROS-producing system and their toxicity to PMNs. RESULTS: Both compounds had concentration-dependent inhibitory effects on LECL. Quercetin at concentration of 5 micromol/l inhibited 94.5+/-1.0% of LECL, whereas 7-allyloxycoumarin at concentration of 200 micromol/l inhibited 53.8+/-2.4% of LECL. Neither compound was toxic to PMNs under the tested conditions. DISCUSSION: The proposed method may be useful for the screening of nontoxic compounds that can modulate ROS production by IC-stimulated PMNs. Special attention should be devoted to natural compounds from higher plants, since their potential as sources of new drugs is still largely unexplored.

Construction and evaluation of an inexpensive device that simulates oral clearance / monstrução e avaliação de um dispositivo de baixo custo simulador do clearace oral

O perfil do clearance de uma determinada substância é muito importante para se estimar a sua disponibilidade oral. Dispositivos ou modelos que simulam o clearance na cavidade oral são importantes para se estudar os efeitos e tempos de retenção de alimentos e fármacos. Este trabalho descreve um eficiente dispositivo, desenvolvido em nosso laboratório com materiais de baixo custo, que pode ser utilizado para o estudo do clearance de substratos cariogênicos, enxagatórios bucais e fármacos de liberação programada, bem como, avaliar a adsorção de fármacos ao esmalte. O dispositivo apresenta três câmaras com volumes mínimo e máximo variáveis, que podem ser eluidas, simultaneamente, em diferentes fluxos. Os volumes de deglutição simulados são ajustáveis e a razão entre os volumes mínimo e máximo pode ser programada. Também são apresentados os resultados de um estudo de avaliação do dispositivo onde determinou-se o clearance de fluoreto proveniente de um enxaguatório bucal, o clearance de uma solução de glicose a 1 por cento e a liberação programada de fluoreto a partir de um adesivo de metacrilato, usando saliva artificial como eluente

In vivo and in vitro studies of a cetylamine fluoride mouthrinse: evaluation of a decive used for in vitro experiments / Estudos in vivo e in vitro de enxaguatório bucal contendo cetilamino fluoreto: validaçäo de dispositivo usado nos experimentos in vitro

Um enxaguatório bucal contendo cetilamino fluoreto (230 ppm de fluoreto) foi preparado para estudos in vitro do clearance de fluoreto e adsorçäo deste íon pelo esmalte dental e/ou hidroxiapatita, utilizando um dispositivo que simula a cavidade oral. Os estudos in vivo do clearance de fluoreto a partir desse enxaguatório bucal foram realizados e comparados com outras fontes de fluoreto. A quantidade de fluoreto absorvida aos blocos de dente ou hidroxiapatita em pó, ambos tratados com o enxaguatório bucal de cetilamino fluoreto, uma ou duas vezes ao dia, foi determinada. Os resultados desses estudos demonstraram ser possível a preparaçäo de enxaguatório bucal contendo cetilamino fluoreto como alternativa para a prevençäo e terapêutica de cáries dentárias

Método para avaliaçäo clínica da capacidade tamponante salivar / A method for the clinical evaluation of salivary buffer capacity

O presente estudo teve por objetivo o desenvolvimento e a padronizaçäo de um método colorimétrico para avaliar a capacidade tamponante salivar média de nossa populaçäo. As amostras de saliva estimulada foram tituladas sem eliminaçäo do CO². Estudos foram realizados para verificar a influência da perda de CO² das amostras durante o período necessário para a realizaçäo das titulaçöes e para a padronizaçäo do tempo de leitura do pH. A partir das titulaçöes realizadas nas amostras de saliva de 206 indivíduos de ambos os sexos na faixa etária de 5 aos 50 anos, na proporçäo de 0,77 : 1 : 0,44, respectivamente para crianças, jovens e adultos, foram feitas as distribuiçöes de freqüências dos casos, a determinaçäo média do pH inicial, do pKa, microequivalentes relativos ao final da titulaçäo, pH final da titulaçäo e uma estimativa da porcentagem da saliva titulada em pH 5...

Enamel fluoride concentrations in unerupted third molars and the influence of fluoridated water on caries scores / Concentração do esmalte fluoretado em terceiros molares não erupcionados e a influência da água fluoretada em escores de cárie

Enamel fluoride concentrations in buccal and lingual surfaces were determined in successive layers obtained by the acid etch method. The 48 unerupted third molar teeth were from subjects who had lived continuosly since birth in two Brazilian towns (Barretos and Ribeirão Prêto) with different fluoride levels in the drinking water but with similar socioeconomic conditions. The DMFT index was determined for children 7-12 years old living in the two towns. Enamel fluoride concentrations were significantly higher in the teeth from the town with higher fluoride levels in drinking water. The children residing in the community with near-optimum fluoride concentration in the water had mean DMFT scores that were less than half those found in the community with a low level of fluoride in the drinking water

Avaliação do teor de fluoreto na água de abastecimento de cidades na região de Ribeirão Preto / Evaluation of community water supply in towns of Ribeirão Preto region

O teor de fluoreto na água de abastecimento de 68 cidades da região de Ribeirão Preto (SP) foi avaliada pelo método do eletrodo seletivo. As amostras foram recolhidas em diferentes pontos das cidades, em função de seu tamanho, com o objetivo de avaliar, naquele instante, a situação de fluoretação da água de abastecimento. Os resultados mostraram que 31 (45,6//) cidades não apresentam água fluoretada, 17 cidades (25,0//) apresentam teor de fluoreto de 0,1 a 0,5 ppm, 13 cidades (19,1//) com 0,5 a 0,7 ppm e 7 cidades (10,3//) com concentração de fluoreto na faixa de 0,7 a 1,0 ppm