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J K Nicholson

Publications and source records attributed to J K Nicholson.

At least 91 records · Page 5Linked to original sources
PubMed2000

Evaluation of TruCount absolute-count tubes for determining CD4 and CD8 cell numbers in human immunodeficiency virus-positive adults. Site Investigators and The NIAID DAIDS New Technologies Evaluation Group.

A single-platform technology that uses an internal bead standard and three-color flow cytometry to determine CD4 and CD8 absolute counts was evaluated for reproducibility and agreement. Values obtained using TruCount absolute-count tubes were compared to those obtained using a two-color predicate methodology. Sixty specimens from human immunodeficiency virus type 1-infected donors were shipped to five laboratories. Each site also analyzed replicates of 14 human immunodeficiency virus type 1-infected local specimens at 6 h and again at 24 h. The interlaboratory variability was significantly less with TruCount (median difference in percent coefficient of variation [%CV] between the two methods was -8% and -3% for CD4 and CD8, respectively) than with the predicate method. Intralaboratory variability was smaller, with a median difference in %CV of -1% for both CD4 and CD8 with 6-h samples and -2% and -3% for CD4 and CD8, respectively, with 24-h samples. Use of TruCount for shipped samples resulted in a median CD4 count change of 7 cells (50th estimated percentile) when all laboratories and CD4 strata were combined. For on-site samples, the median CD4 count change was 10 CD4 cells for 6-h samples and 2 CD4 cells for 24-h samples. Individual site biases occurred in both directions and cancelled each other when the data were combined for all laboratories. Thus, the combined data showed a smaller change in median CD4 count than what may have occurred at an individual site. In summary, the use of TruCount decreased both the inter- and intralaboratory variability in determining absolute CD4 and CD8 counts.

Acquired Immunodeficiency Syndrome↗
PubMed1999

Evaluation of stabilized whole blood control materials for lymphocyte immunophenotyping.

We evaluated stabilized whole blood control materials for their performance as a routine control material for immunophenotyping using flow cytometry. These materials serve as the method control, controlling for both the binding of monoclonal antibodies to the cells and the efficiency of the red-blood-cell lysing reagent. Three products (FluoroTrol, StatusFlow, and CD-Chex): were tested. The controls performed very well in evaluating both CD4 and CD8 percentages and absolute cell counts using flow cytometry. Light scattering patterns and fluorescence intensity of antibody binding were slightly different from those of normal whole blood. These products are not appropriate for quantitative fluorescence standards. Cytometry (Comm. Clin. Cytometry) 38:268-273, 1999. Published 1999 Wiley-Liss, Inc.

Blood Chemical Analysis↗
PubMed1999

Dietary minerals in the gastrointestinal tract: hydroxypolymerisation of aluminium is regulated by luminal mucins.

The regulation of mineral absorption in the gastrointestinal tract is poorly understood. Recent work has identified an intracellular metal-ion transporter but considerable evidence suggests that both soluble and mucosally associated luminal metal-binding ligands regulate initial uptake. Molecules ranging from low molecular weight organic acids to large glycoproteins have been suggested but a definite role for any such species has remained elusive. Here, a series of analytical techniques, allowing for this wide variation in potential binding ligands, was applied to the study of intestinal contents and tissue of rats following different feeding protocols. Aluminium, that has a low endogenous background and maintains a high concentration in the gastrointestinal tract, was investigated as a suitable dietary metal with hydrolytic behaviour similar, for example, to copper, iron and zinc. High resolution nuclear magnetic resonance spectroscopy identified a number of endogenous low molecular weight weak ligands that are secreted into the intestinal lumen. These may slow the rate of hydroxy-polymerisation of hydrolytic metals, allowing their effective donation to less mobile, higher molecular weight binding ligands. Histochemical staining suggested that such species may be soluble mucins as these were consistently associated with luminal aluminium. Significantly, this interaction prevented hydroxy/phosphate precipitation of aluminium, even at supraphysiological levels of the element. This was confirmed with X-ray micro-analysis investigations of ex vivo luminal contents. Nevertheless, from phase distribution experiments, the majority (60-95%) of luminal aluminium was associated with the intestinal solid phase and further histochemistry confirmed this to be gelatinous mucus, chiefly as the mucosally adherent layer. All results suggest a major role for mucus in regulating the gastrointestinal absorption of aluminium. It is proposed that, initially, soluble luminal mucus prevents the hydroxy-precipitation of hydrolytic metals at intestinal pH, allowing their effective donation to the mucus layer. Based on the differing reported metal-mucus interactions, elements that bind well to mucus (Al3+, Fe3+), with kinetically slow rates of ligand exchange (Al3+ < Fe3+) will be less well absorbed than poorly bound elements with kinetically faster rates of ligand exchange (Cu2+, Zn2+ etc.). This mechanism would readily explain many of the reported observations on mineral availability, including the marked variation in absorption of different elements, the differential effects of dietary ligands on mineral uptake and the competition for absorption between different metals.

Aluminum↗
PubMed1999

Characterization of metabolites in intact Streptomyces citricolor culture supernatants using high-resolution nuclear magnetic resonance and directly coupled high-pressure liquid chromatography-nuclear magnetic resonance spectroscopy.

A novel NMR spectroscopic approach to the direct biochemical characterization of bacterial culture broths is presented. A variety of one- and two-dimensional 1H NMR spectroscopic methods were used to characterize low-molecular-weight organic components of broth supernatants from cultures of Streptomyces citricolor. By applying 1H NMR spectroscopy to analyze whole, untreated culture supernatants, it was possible to identify and monitor simultaneously a range of media substrates and excreted metabolites. Identified metabolites include 2-phenylethylamine, trehalose, succinate, acetate, uridine, and aristeromycin, a secondary metabolite with antibiotic properties. Directly coupled HPLC-NMR spectroscopy was also applied to the analysis of broth supernatants for the first time, to aid spectral assignments, especially where signals were extensively overlapped in the 1H NMR spectra of the whole broth mixtures. Two-dimensional NMR methods such as 1H-1H correlation spectroscopy, 1H-13C heteronuclear single quantum correlation, and 1H-13C heteronuclear multiple bond correlation aided the structure elucidation and peak assignments of individual components in the mixtures by providing information on 1H-1H coupling networks and 13C chemical shifts. This work shows that high-resolution NMR spectroscopic methods provide a rapid and efficient means of investigating microbial metabolism directly without invasive or destructive sample pretreatment.

Acetic Acid↗
PubMed1999

High-resolution magic angle spinning 1H NMR spectroscopic studies on intact rat renal cortex and medulla.

High-resolution magic angle spinning 1H NMR (MAS-NMR) spectroscopy was used to investigate the biochemical composition of normal renal cortex and renal papilla samples from rats, and results were compared with those from conventional 1H NMR analysis of protein-free tissue extracts. 1H MAS NMR spectra of samples obtained from inner and outer cortex were found to be broadly similar in terms of metabolite profile, and intra- and inter-animal variability was small. However, the MAS NMR spectra from renal papilla samples were qualitatively and quantitatively different from those obtained from cortex. High levels of free amino acids and several organic acids were detected in the cortex, together with choline, glucose, and trimethylamine-N-oxide. The dominant metabolite resonances observed in papillary tissue were from glycerophosphocholine (GPC), betaine, myo-inositol, and sorbitol. On increasing the magic angle spinning rate from 4,200 to 12,000 Hz, the lipid MAS 1H NMR signal profile remained largely unchanged in papillary tissue, whereas "new" resonances from triglycerides appeared in the spectra of cortical tissue, this effect being reversible on returning the spinning rate to 4,200 Hz. Further investigation into the behavior of the lipid components under different spinning rates suggested that the lipids in the cortex were present in more motionally constrained environments than those in the papilla. 1H MAS NMR spectra of tissues are of value both in interrogation of the biochemical composition of whole tissue, and in obtaining information on the mobility and compartmentalization of certain metabolites.

Animals↗
PubMed1999

Comparison of in vivo 1H MRS of human brain tumours with 1H HR-MAS spectroscopy of intact biopsy samples in vitro.

High resolution magic angle spinning (MAS) 1H nuclear magnetic resonance (NMR) spectroscopy has been employed to study intact human brain tumour tissue and comparison with the corresponding in vivo spectrum has been made. Two dimensional 1H MAS-NMR measurements, including J-resolved and homonuclear shift correlation spectra, were obtained to aid metabolite signal assignment. MAS gave greatly improved line-shape and reduced line-width in comparison to conventional high resolution in vivo 1H MRS of intact tissue, permitting the simultaneous detection of cellular lipids and metabolites. The technique provides the most direct method for comparison of in vivo spectra with high resolution spectra in vitro and hence allows more reliable peak assignment of in vivo 1H MRS spectra.

Astrocytoma↗
PubMed1999

Degradation of 4-fluorobiphenyl in soil investigated by 19F NMR spectroscopy and 14C radiolabelling analysis.

The incubation of the model pollutant [U-14C]'-4-fluorobiphenyl (4FBP) in soil, in the presence and absence of biphenyl (a co-substrate), was carried out in order to study the qualitative disposition and fate of the compound using 14C-HPLC and 19F NMR spectroscopy. Components accounted for using the radiolabel were volatilization, CO2 evolution, organic solvent extractable and bound residue. Quantitative analysis of these data gave a complete mass balance. After sample preparation. 14C-HPLC was used to establish the number of 4FBP related components present in the organic solvent extract. 19F NMR was also used to quantify the organic extracts and to identify the components of the extract. Both approaches showed that the composition of the solvent extractable fractions comprised only parent compound with no metabolites present. As the 14C radiolabel was found to be incorporated into the soil organic matter this indicates that metabolites were being generated, but were highly transitory as incorporation into the SOM was rapid. The inclusion of the co-substrate biphenyl was to increase the overall rate of degradation of 4FBP in soil. The kinetics of disappearance of parent from the soil using the data obtained were investigated from both techniques. This is the first report describing the degradation of a fluorinated biphenyl in soil.

Biodegradation, Environmental↗
PubMed1999

Quantitative structure-toxicity relationships for halogenated substituted-benzenes to Vibrio fischeri, using atom-based semi-empirical molecular-orbital descriptors.

Quantitative structure-toxicity relationships (QSTR's) are derived for an extensive series of halogenated benzenes, anilines, phenols, nitrobenzenes, toluenes and other substituted benzenes against Vibrio fischeri using a wide range of whole molecule and atom-based descriptors derived from semi-empirical molecular-orbital calculations. In terms of direct statistical correlation with toxicity it was found that the molar refractivity was the most important parameter, closely followed by the solvent accessible surface area of the compound. The accuracy of these descriptors in fitting the numerous fluoro- and chloro-mono-aromatic compounds was compared with bromine and iodine analogues, where the 'best' descriptors for the former were found in general to be less accurate for the latter in the case of multi-halogen substitution. The equations obtained were also used to classify the compounds into narcosis-based mechanisms of toxicity and those with respiratory uncoupling potential. A combination of the molar refractivity and the nucleophilic susceptibility of one of the meta ring carbons predicted the toxicity of the halo-benzenes and toluenes, along with anisoles, benzonitriles, nitrobenzenes and most of the anilines. The relevance of these descriptors to developing coherent and more generally applicable models for QSTR's of mono-aromatic compounds to other species in environmental toxicology is discussed.

Benzene Derivatives↗
PubMed1999

Impurity profiling in bulk pharmaceutical batches using 19F NMR spectroscopy and distinction between monomeric and dimeric impurities by NMR-based diffusion measurements.

The impurity profile of production batches of fluorine-containing drugs can be characterised efficiently using 19F NMR spectroscopy. This yields the number and proportions of impurities in the bulk drug to a level of approximately equal 0.1 mole% in a few minutes of NMR experiment time. The approach has been exemplified using a partially purified batch of the steroidal product fluticasone propionate, the impurities in which include a number of dimeric species. Further distinction between the monomer and dimer impurities has been achieved through high resolution chemical shift-resolved NMR measurement of molecular diffusion coefficients on the intact mixture using 19F NMR spectroscopy. The ability of NMR-based diffusion coefficient determination to distinguish between monomeric and dimeric substances was validated using a standard mixture of authentic materials containing both monomers and dimers.

Androstadienes↗
PubMed1999

Directly-coupled HPLC-NMR spectroscopic studies of metabolism and futile deacetylation of phenacetin in the rat.

The metabolism and futile deacetylation of phenacetin has been investigated in the rat via 1H NMR spectroscopic analysis of urine. Animals were dosed with either phenacetin or phenacetin-C2H3 and urine samples were collected for -24-0 (pre-dosing), 0-8. 8-24, and 24-48 h post-dosing. Drug metabolites of the two compounds were concentrated from the urine using solid-phase extraction prior to the use of directly-coupled HPLC-1H NMR spectroscopy for separation and identification. Following dosing of phenacetin, the metabolites identified were paracetamol glucuronide, paracetamol and N-hydroxyparacetamol, whilst paracetamol and N-hydroxyparacetamol sulphate were identified following dosing of phenacetin-C2H3. Quantitatively the percentage futile deacetylation of phenacetin-C2H3 metabolites was found to be 32% in both paracetamol and N-hydroxyparacetamol sulphate. This study further indicated the importance of futile deacetylation in simple analgesics and the value of directly-coupled HPLC-NMR spectroscopy for the study of this process.

Acetaminophen↗
PubMed1999

Studies on the metabolism of 4-fluoroaniline and 4-fluoroacetanilide in rat: formation of 4-acetamidophenol (paracetamol) and its metabolites via defluorination and N-acetylation.

1. The urinary metabolic fate of 4-fluoroaniline (4-FA) and 1-[13C]-4-fluoroacetanilide (4-FAA) has been studied using NMR-based methods after 50 and 100 mg kg(-1) i.p. doses respectively to the male Sprague-Dawley rat. 2. 4-FA was both ortho- and para-hydroxylated. The major metabolite produced by ortho-hydroxylation was 2-amino-5-fluorophenylsulphate accounting for approximately 30% of the dose. Of the dose, approximately 10% was excreted via para-hydroxylation and the resulting defluorinated metabolites were N-acetylated and excreted as sulphate (major), glucuronide (minor) and N-acetyl-cysteinyl (minor) conjugates of 4-acetamidophenol (paracetamol). 3. The major route of metabolism of 1-[13C]-4-FAA was N-deacetylation and the metabolites excreted in the urine were qualitatively identical to 4-FA. The paracetamol metabolites produced via para-hydroxylation were also a product of N-deacetylation and reacetylation, as the [13C]-label was not retained. 4. These studies demonstrate the value of [13C]-labelling in understanding the contribution of N-acetylation, and futile deacetylation-reacetylation reactions, in aniline metabolism. In addition, this work sheds new light on the metabolic lability of certain aromatic fluorine substituents.

Acetaminophen↗
PubMed1999

Quantitative structure-metabolism relationships (QSMR) using computational chemistry: pattern recognition analysis and statistical prediction of phase II conjugation reactions of substituted benzoic acids in the rat.

1. Quantitative relationships between molecular physico-chemical properties of 22 substituted benzoic acids and the extent of excretion of their metabolites in rat urine have been investigated using computational chemistry and multivariate statistics. 2. A data set of 34 theoretically derived physico-chemical descriptors calculated was used to classify the benzoic acids according to their predominant urinary metabolic fate. 3. Quantitative structure-metabolism relationships were obtained by linear regression using combinations of physico-chemical descriptors allowing the prediction of % urinary excretion of glycine (r = 0.73) and glucuronide conjugates (r = 0.82) and % urinary excretion of the parent compound (r = 0.91).

Animals↗
PubMed1999

Quantitative studies on the urinary metabolic fate of 2-chloro-4-trifluoromethylaniline in the rat using 19F-NMR spectroscopy and directly coupled HPLC-NMR-MS.

1. The metabolism and urinary excretion of 2-chloro-4-trifluoromethylaniline has been studied in the rat using 19F-NMR spectroscopy and directly coupled HPLC-NMR-MS methods. The compound was dosed to three male Sprague-Dawley rats (50 mg kg(-1) i.p.) and urine collected over 0-8, 8-24 and 24-48 h post-dosing. 2. A total urinary recovery of 56.3+/-2.2% of the dose was achieved up to 48 h after dosing. The major metabolite in the urine was identified as 2-amino-3-chloro-5-trifluoromethylphenylsulphate accounting for a total of 33.5+/-2.2% of the dose. 3. Further metabolites detected and characterized included 2-chloro-4-trifluoromethylphenylhydroxylamine glucuronide (13.2+/-0.5% of the dose), 2-amino-3-chloro-5-trifluoromethylphenylglucuronide (3.8+/-0.4% of the dose) and 2-chloro-4-trifluoromethylaniline-N-glucuronide (3.6+/-0.1% of the dose). Several minor metabolites were also found and identified, including 2-chloro-4-trifluoromethylphenylsulphamate, which together accounted for 2.1+/-0.4% of the dose. 4. Directly coupled HPLC-NMR-MS and 19F-NMR spectroscopy is shown to provide an efficient approach for the unequivocal and rapid determination of the quantitative urinary metabolic fate and excretion balance of a fluorinated xenobiotic without the necessity for specific radiolabelling.

Aniline Compounds↗
PubMed1999

Quantitative structure-toxicity relationships for chlorophenols to bioluminescent lux-marked bacteria using atom-based semi-empirical molecular-orbital descriptors.

Literature data on the toxicity of chlorophenols for three luminescent bacteria (Vibrio fischeri, and the lux-marked Pseudomonas fluorescens 10586s pUCD607 and Burkholderia spp. RASC c2 (Tn4431)) have been analyzed in relation to a set of computed molecular physico-chemical properties. The quantitative structure-toxicity relationships of the compounds in each species showed marked differences when based upon semi-empirical molecular-orbital molecular and atom based properties. For mono-, di- and tri-chlorophenols multiple linear regression analysis of V. fischeri toxicity showed a good correlation with the solvent accessible surface area and the charge on the oxygen atom. This correlation successfully predicted the toxicity of the heavily chlorinated phenols, suggesting in V. fischeri only one overall mechanism is present for all chlorophenols. Good correlations were also found for RASC c2 with molecular properties, such as the surface area and the nucleophilic super-delocalizability of the oxygen. In contrast the best QSTR for P. fluorescens contained the 2nd order connectivity index and ELUMO suggesting a different, more reactive mechanism. Cross-species correlations were examined, and between V. fischeri and RASC c2 the inclusion of the minimum value of the nucleophilic susceptibility on the ring carbons produced good results. Poorer correlations were found with P. fluorescens highlighting the relative similarity of V. fischeri and RASC c2, in contrast to that of P. fluorescens.

Burkholderia↗
PubMed1999

Degradation of 4-fluorobiphenyl by mycorrhizal fungi as determined by (19)F nuclear magnetic resonance spectroscopy and (14)C radiolabelling analysis.

The pathways of biotransformation of 4-fluorobiphenyl (4FBP) by the ectomycorrhizal fungus Tylospora fibrilosa and several other mycorrhizal fungi were investigated by using (19)F nuclear magnetic resonance (NMR) spectroscopy in combination with (14)C radioisotope-detected high-performance liquid chromatography ((14)C-HPLC). Under the conditions used in this study T. fibrillosa and some other species degraded 4FBP. (14)C-HPLC profiles indicated that there were four major biotransformation products, whereas (19)F NMR showed that there were six major fluorine-containing products. We confirmed that 4-fluorobiphen-4'-ol and 4-fluorobiphen-3'-ol were two of the major products formed, but no other products were conclusively identified. There was no evidence for the expected biotransformation pathway (namely, meta cleavage of the less halogenated ring), as none of the expected products of this route were found. To the best of our knowledge, this is the first report describing intermediates formed during mycorrhizal degradation of halogenated biphenyls.

Basidiomycota↗
PubMed1999

Investigation of the quantitative metabolic fate and urinary excretion of 3-methyl-4-trifluoromethylaniline and 3-methyl-4-trifluoromethylacetanilide in the rat.

The urinary metabolites of 3-methyl-4-trifluoromethylaniline in the rat were characterized and quantified using a combination of (19)F NMR, HPLC-NMR ((1)H and (19)F), and HPLC-mass spectrometry techniques. The major routes of metabolism were amine N-acetylation and methyl group C-oxidation to the benzyl alcohol (with subsequent glucuronide conjugation) and further to the corresponding benzoic acid derivative. Quantitatively only a small proportion of the urinary metabolites contained the free amino group, and these were products of ortho-hydroxylation (2 and 6 position) with additional conjugation to form the ether sulfates and glucuronides. An N-glucuronide of the parent compound was also identified. 3-Methyl-4-trifluoromethylacetanilide ((13)C-labeled in the acetyl group) gave virtually the same overall metabolite profile as 3-methyl-4-trifluoromethylaniline; however, a significant level of futile N-deacetylation and reacetylation occurred as ca. 50% of the excreted N-acetylated major metabolites contained no (13)C-label at the acetyl, having been replaced by an endogenous (12)C-acetyl source. This level of futile deacetylation is the highest yet reported for a substituted aniline/acetanilide and indicates a high degree of electronic activation of the amino group toward the acetyltransferase enzymes in vivo.

Anilides↗