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1.
J Appl Toxicol ; 30(2): 172-82, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19839026

RESUMO

Limited experimental models exist to assess drug toxicity in pediatric populations. We recently reported how a multi-age rat model could be used for pre-clinical studies of comparative drug toxicity in pediatric populations. The objective of this study was to expand the utility of this animal model, which previously demonstrated an age-dependent sensitivity to the classic nephrotoxic compound, gentamicin, to another nephrotoxicant, namely cisplatin (Cis). Sprague-Dawley rats (10, 25, 40 and 80 days old) were injected with a single dose of Cis (0, 1, 3 or 6 mg kg(-1) i.p.). Urine samples were collected prior and up to 72 h after treatment in animals that were >or= 25 days old. Several serum, urinary and 'omic' injury biomarkers as well as renal histopathology lesions were evaluated. Statistically significant changes were noted with different injury biomarkers in different age groups. The order of age-related Cis-induced nephrotoxicity was different than our previous study with gentamicin: 80 > 40 > 10 > 25 day-old vs 10 >or= 80 > 40 > 25-day-old rats, respectively. The increased levels of kidney injury molecule-1 (Kim-1: urinary protein/tissue mRNA) provided evidence of early Cis-induced nephrotoxicity in the most sensitive age group (80 days old). Levels of Kim-1 tissue mRNA and urinary protein were significantly correlated to each other and to the severity of renal histopathology lesions. These data indicate that the multi-age rat model can be used to demonstrate different age-related sensitivities to renal injury using mechanistically distinct nephrotoxicants, which is reflected in measurements of a variety of metabolite, gene transcript and protein biomarkers.


Assuntos
Envelhecimento/fisiologia , Cisplatino/toxicidade , Nefropatias/induzido quimicamente , Rim/metabolismo , Fatores Etários , Animais , Biomarcadores/metabolismo , Biomarcadores/urina , Criança , Suscetibilidade a Doenças/metabolismo , Suscetibilidade a Doenças/patologia , Gentamicinas/toxicidade , Humanos , Rim/patologia , Nefropatias/patologia , Nefropatias/urina , Modelos Animais , Pediatria , Ratos , Ratos Sprague-Dawley , Sensibilidade e Especificidade
3.
Rapid Commun Mass Spectrom ; 14(10): 911-7, 2000.
Artigo em Inglês | MEDLINE | ID: mdl-10825256

RESUMO

Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectra of bacterial proteins were obtained from water, lettuce and cloth samples contaminated with Shigella flexneri, Escherichia coli, and Aeromonas hydrophila. Spectra were obtained using proteins directly isolated from water (or water used for rinsing samples) without culturing the bacteria. For S. flexneri and E. coli, two marker ions for specific proteins associated with a virulence-related property (acid resistance) were easily detected. For A. hydrophila, ions from two specifically selected marker proteins, as well as ions from the larger group of proteins isolated from pure cultures, all matched spectra from a contaminated water sample, providing strong evidence that A. hydrophila was the bacterial contaminant. Rinse water from contaminated lettuce and cloth samples showed the same marker ions as the contaminated water samples.


Assuntos
Proteínas de Bactérias/análise , Microbiologia de Alimentos , Gossypium/microbiologia , Lactuca/microbiologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Microbiologia da Água , Aeromonas hydrophila/isolamento & purificação , Biomarcadores/análise , Escherichia coli/isolamento & purificação , Íons , Lasers , Shigella flexneri/isolamento & purificação
4.
Anal Chem ; 71(15): 3226-30, 1999 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-10450164

RESUMO

Characteristic ions in the MALDI TOF mass spectra from bacterial cells have been associated with four known proteins. The proteins, observed both from cells and in filtered cellular suspensions, were isolated by HPLC and identified on the basis of their mass spectra and their partial amino acid sequence, determined using the Edman method (10-15 residues). The acid resistance proteins HdeA and HdeB give rise to ions near m/z 9735 and 9060 in MALDI TOF mass spectra from cells and from extracts of both Escherichia coli 1090 and Shigella flexneri PHS-1059. However, the proteins associated with proteolytic cleavage by the peptidase Lep, rather than the precursor proteins, were observed, both using cells and from cellular extracts. A cold-shock protein, CspA, was associated with the ion near m/z 7643 from Pseudomonas aeruginosa. Similarly, a cold-acclimation protein, CapB, was identified as the source of the ion near m/z 7684 in P. putida. This last protein was homologous with a known CapB from P. fragi. While these experiments involved the detection of known or homologous proteins from typical bacteria, this same approach could also be applied to the detection of unique proteins or biomarker proteins associated with other bacteria of public health significance.


Assuntos
Proteínas de Bactérias/análise , Proteínas de Escherichia coli , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Sequência de Aminoácidos , Proteínas de Bactérias/isolamento & purificação , Proteínas de Transporte/análise , Proteínas de Transporte/isolamento & purificação , Cromatografia Líquida de Alta Pressão , Temperatura Baixa , Escherichia coli/química , Proteínas de Choque Térmico/análise , Proteínas de Choque Térmico/isolamento & purificação , Dados de Sequência Molecular , Pseudomonas aeruginosa/química , Homologia de Sequência de Aminoácidos , Shigella flexneri/química
5.
Chem Biol Interact ; 123(3): 197-217, 1999 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-10654839

RESUMO

The peroxidase from Coprinus cinereus (CPX) catalyzed oxidative oligomerization of 4-chloroaniline (4-CA) forming several products: N-(4-chlorophenyl)-benzoquinone monoamine (dimer D), 4,4'-dichloroazobenzene (dimer E); 2-(4-chloroanilino)-N-(4-chlorophenyl)-benzoquinone (trimer F); 2-amino-5-chlorobenzoquinone-di-4-chloroanil (trimer G); 2-(4-chloroanilino)-5-hydroxybenzoquinone-di-4-chloroanil (tetramer H) and 2-amino-5-(-4-chlroanilino)-benzoquinone-di-4-chloroanil (tetramer 1). In the presence of 4-CA and H2O2, CPX was irreversibly inactivated within 10 min. Inactivation of CPX in the presence of H2O2 was a time-dependent, first-order process when the concentration of 4-CA was varied between 0 and 2.5 mM. The apparent dissociation constant (Ki) for CPX and 4-CA was 0.71 mM. The pseudo-first order rate constant for inactivation (k(inact)), was 1.15 x 10(-2) s(-1). Covalent incorporation of 20 mole 14C-4-CA per mole of inactivated CPX was observed. The partition ratio was about 2200 when either 4-CA or H2O2 was used as the limiting substrate. These results show that 4-CA is a metabolically activated inactivator (i.e. a suicide substrate). Unmodified heme and hydroxymethyl heme were isolated from native, 4-CA-inactivated and H2O2-incubated CPX. Inactivation resulted in significant losses in both heme contents. Analysis of tryptic peptides from 4-CA-inactivated CPX by MALDI-TOF/ MS and UV-VIS spectrophotometry suggested that trimer G and tetramer H were the major 4-CA derivatives that were covalently bound, including to a peptide (MGDAGF-SPDEVVDLLAAHSLASQEGLNSAIFR) containing the heme binding site. These studies show that heme destruction and covalent modification of the polypeptide chain are both important for the inactivation of CPX. These results were compared with similar studies on 4-CA-inactivated horseradish peroxidase (HRP) and bovine lactoperoxidase (LPO) during the oxidation of 4-CA.


Assuntos
Compostos de Anilina/farmacologia , Coprinus/enzimologia , Inibidores Enzimáticos/farmacologia , Peroxidase do Rábano Silvestre/antagonistas & inibidores , Lactoperoxidase/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Bovinos , Cromatografia Líquida de Alta Pressão , Peroxidase do Rábano Silvestre/química , Peroxidase do Rábano Silvestre/metabolismo , Peróxido de Hidrogênio/farmacologia , Cinética , Lactoperoxidase/química , Lactoperoxidase/metabolismo , Dados de Sequência Molecular , Oxirredução , Mapeamento de Peptídeos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
6.
Bioorg Med Chem Lett ; 8(16): 2231-4, 1998 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-9873519

RESUMO

Problems were encountered during attempts to prepare N-terminal cysteine-substituted peptide nucleic acids (PNAs) from commercially available, Fmoc-protected monomers. These problems have been surmounted by the use of an S-t-butylmercapto protecting group on the cysteine moiety. The solid-phase syntheses are carried out via a simplified procedure which should be generally useful for manual PNA synthesis.


Assuntos
Cisteína , Oligodesoxirribonucleotídeos/química , Ácidos Nucleicos Peptídicos/síntese química , Sequência de Bases , Cromatografia Líquida de Alta Pressão , Indicadores e Reagentes , Oligodesoxirribonucleotídeos/síntese química , Ácidos Nucleicos Peptídicos/química , Resinas Vegetais , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
7.
Histochem J ; 29(3): 239-48, 1997 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-9472386

RESUMO

Fucus serratus eggs were examined for evidence of the existence of a lysosome-like body using enzyme histochemical and vital staining techniques. Simultaneous coupling azo-dye techniques for lysosomal acid phosphatase proved inappropriate owing to endogenous phenolic binding artefacts. The large number of alginate polysaccharide and polyphenolic egg vesicles interfered with vital staining techniques for lysosomes. Lysosomal esterase activity was detected in the abundant egg lipid bodies. The role of the egg lipid body as an equivalent lysosome-like body of higher plants, the spherosome, is discussed in relation to egg fertilization and early zygote development.


Assuntos
Lisossomos/ultraestrutura , Alga Marinha/citologia , Zigoto/ultraestrutura , Fosfatase Ácida/metabolismo , Laranja de Acridina , Corantes , Esterases/metabolismo , Corantes Fluorescentes , Histocitoquímica , Lisossomos/enzimologia , Vermelho Neutro , Oxazinas , Polissacarídeos/metabolismo , Alga Marinha/enzimologia , Alga Marinha/ultraestrutura , Zigoto/enzimologia
8.
Rapid Commun Mass Spectrom ; 10(10): 1227-32, 1996.
Artigo em Inglês | MEDLINE | ID: mdl-8759332

RESUMO

Matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) was investigated as a method for the rapid identification of whole bacteria, either by comparison with archived reference spectra or by co-analysis with cultures of known bacteria. Bacteria were sampled from colonies on an agar plate, mixed with the matrix, air-dried, and introduced in batches into the mass spectrometer for analysis. In the first experiment, both bacterial strains that had been previously analyzed to obtain reference spectra and other strains that had not been analyzed were blind-numbered and their spectra were obtained. Those strains that matched reference spectra were found to be correctly identified. A second experiment involved co-analysis of reference strains and bind-numbered strains under identical conditions; species-specific identification was demonstrated by comparison of spectra of the blind-numbered strains with those of the standards. In all of the spectra obtained in these experiments, each bacterial strain showed a few characteristic high-mass ions which are thought to be derived from bacterial proteins. This work represents the first reported instance of successful bacterial chemotaxonomy by MALDI-TOFMS analysis of whole cells. For the strains tested, the method is rapid and simple.


Assuntos
Bactérias/química , Proteínas de Bactérias/análise , Calibragem , Padrões de Referência , Especificidade da Espécie , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
9.
Anal Chem ; 65(9): 1140-6, 1993 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-8503502

RESUMO

Naturally occurring mycotoxins are separated using micellar electrokinetic capillary chromatography. Trends in the retention of these toxins, resulting from changes in mobile-phase composition and pH, are reported and presented as a means of alleviating coelution problems. Two sets of mobile-phase conditions are determined that provide unique separation selectivity. The facile manner by which mobile-phase conditions can be altered, without changes in instrumental configuration, allowed the acquisition of two distinctive, fully resolved chromatograms of 10 mycotoxins in a period of approximately 45 min. By adjusting retention times, using indigenous or added components in mycotoxin samples as normalization standards, it is possible to obtain coefficients of variation in retention time that average less than 1%. The qualitative capabilities of this methodology are evaluated by separating randomly generated mycotoxin-interferent mixtures. In this study, the utilization of normalized retention times applied to separations obtained with two sets of mobile-phase conditions permitted the identification of all the mycotoxins in five unknown samples without any misidentifications.


Assuntos
Cromatografia/métodos , Micotoxinas/análise , Eletroquímica , Reprodutibilidade dos Testes
10.
Talanta ; 39(9): 1139-47, 1992 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18965505

RESUMO

Micellar electrokinetic capillary chromatography (MECC) is applied to the high-speed analysis of aflatoxins. Baseline separation of the four common aflatoxins G(1), G(2), B(1) and B(2), is accomplished in less than 30 sec. Small (25 mum) internal diameter capillaries are found to be critical in maintaining high efficiency under rapid MECC separation conditions. Van Deemter-like plots are generated in order to study the effects of capillary diameter and organic solvent on efficiency under high electric field conditions. Other experimental parameters affecting efficiency are investigated, including buffer concentration, surfactant concentration, and detector time constant. Simple on-column laser-based fluorescence detection, employing helium-cadmium laser radiation at 325 nm for excitation, allows for limits of detection in the range of 0.05-0.9 femtomoles injected for underivatized aflatoxins. Considerations important in the analysis of aflatoxins in real matrices are presented.

14.
Skeletal Radiol ; 5(3): 145-9, 1980.
Artigo em Inglês | MEDLINE | ID: mdl-6938060

RESUMO

Skeletal lesions were radiographically apparent in 6 of 36 (16%) patients with chronic myelogenous leukemia. The spectrum of radiographic changes including diffuse osteoporosis, focal osteolytic and osteoblastic lesions, chloroma, and arthritis, and their clinical behavior is discussed.


Assuntos
Osso e Ossos/diagnóstico por imagem , Leucemia Mieloide/diagnóstico por imagem , Adolescente , Adulto , Artrite/diagnóstico por imagem , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Osteoporose/diagnóstico por imagem , Radiografia
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