Biomedical subjects

J K Nicholson

Publications and source records attributed to J K Nicholson.

At least 55 records · Page 3Linked to original sources

PubMed2001

Application of directly coupled HPLC NMR to separation and characterization of lipoproteins from human serum.

Disorders in lipoprotein metabolism are critical in the etiology of several disease states such as coronary heart disease and atherosclerosis. Thus, there is considerable interest in the development of novel methods for the analysis of lipoprotein complexes. We report here a simple chromatographic method for the separation of high-density lipoprotein, low-density lipoprotein, and very low-density lipoprotein from intact serum or plasma. The separation was achieved using a hydroxyapatite column and elution with pH 7.4 phosphate buffer with 100-microL injections of whole plasma. Coelution of HDL with plasma proteins such as albumin occurred, and this clearly limits quantitation of that species by HPLC peak integration. We also show, for the first time, the application of directly coupled HPLC 1H NMR spectroscopy to confirm the identification of the three major lipoproteins. The full chromatographic run time was 90 min with stopped-flow 600-MHz NMR spectra of each lipoprotein being collected using 128 scans, in 7 min. The 1H NMR chemical shifts of lipid signals were identical to conventional NMR spectra of freshly prepared lipoprotein standards, confirming that the lipoproteins were not degraded by the HPLC separation and that their gross supramolecular organization was intact.

Chromatography, High Pressure Liquid↗

PubMed2001

High-resolution (1)H NMR and magic angle spinning NMR spectroscopic investigation of the biochemical effects of 2-bromoethanamine in intact renal and hepatic tissue.

The metabolic consequences of xenobiotic-induced toxicity were investigated using high-resolution magic angle spinning (MAS) NMR spectroscopy of intact tissue. Renal papillary necrosis (RPN) was induced in Sprague-Dawley rats (n = 12) via a single i.p. dose of 250 mg/kg 2-bromoethanamine (BEA) hydrobromide. At 2, 4, 6, and 24 h after treatment with BEA, three animals were killed and tissue samples were obtained from liver, renal cortex, and renal medulla. Tissue samples were also removed at 2 and 24 h from matched controls (n = 6). (1)H MAS NMR spectroscopic techniques were used to analyze samples of intact tissue ( approximately 10 mg). Decreased levels of nonperturbing renal osmolytes (glycerophosphocholine, betaine, and myo-inositol) were observed in the renal papilla of BEA-treated animals at 6 and 24 h postdose (p.d.), concomitant with a relative increase in the tissue concentration of creatine. Increased levels of glutaric acid were found in all tissues studied in BEA-treated animals at 4 and 6 h p.d., indicating the inhibition of mitochondrial fatty acyl CoA dehydrogenases and mitochondrial dysfunction. Increased levels of trimethylamine-N-oxide occurred in the renal cortex at 6 h p.d. Changes in the metabolite profile of liver included an increase in the relative concentrations of triglycerides, lysine, and leucine. The novel application of (1)H MAS NMR to the biochemical analysis of intact tissues following a toxic insult highlights the potential of this technique as a toxicological probe in providing a direct link between urinary biomarkers of toxicity and histopathological evaluation of toxicological lesions.

Animals↗

PubMed2001

Abnormal lipid profile of dystrophic cardiac tissue as demonstrated by one- and two-dimensional magic-angle spinning (1)H NMR spectroscopy.

Dystrophin, a protein associated with sarcolemma and cell membranes, is not expressed in sufferers of Duchenne muscular dystrophy (DMD), or in the mdx mouse. DMD is a fatal disorder, with a significant proportion of fatalities associated with cardiac failure ( approximately 40% having dilated cardiomyopathy and >90% clinically significant cardiac defects at death). In this study, the metabolic composition of intact dystrophic cardiac tissue was investigated using high-resolution magic-angle spinning (HRMAS) (1)H NMR spectroscopy with both 1- and 2D pulse sequences coupled with pattern recognition (PR). While conventional solvent presaturation spectra indicated increases in CH(2) chain length in lipids, PR analysis of correlation spectroscopy (COSY) spectra demonstrated that this was also accompanied by an increase in concentration of lactate or threonine along with a relative decrease in CH = CHCH(2)CO groups in these lipids. To investigate the physical environment of these lipids, T(2)- and diffusion-weighted (1)H MAS NMR spectra were acquired on whole-tissue samples. The relatively increased lipid signal intensity in dystrophic tissue was due to an increase in molecules with long T(2) and short diffusion rates. The use of a range of pulse programs allowed the direct probing of the biochemical environment in which the lipid infiltration occurred, and by coupling the experiments to PR the significance of lipid infiltration and accumulation was also assessed.

Animals↗

PubMed2001

Recovery of underwater resonances by magnetization transferred NMR spectroscopy (RECUR-NMR).

A method for detecting small molecule NMR resonances under a water peak in biological samples is presented. After high-efficiency solvent suppression using double WATERGATE, either a TOCSY- or ROESY-based coherence transfer sequence is applied to reestablish the resonances close to, or under, water through magnetization transfer using scalar or dipolar coupling, respectively. The use of the TOCSY and ROESY methods ensures an in-phase magnetization transfer, which makes the new approach readily extended for the measurement of transverse relaxation times, internuclear ROEs, and ROE buildup rates. An extension of the new approach for J-resolved spectroscopy is also presented and tested using a sample of human blood plasma.

Blood Glucose↗

PubMed2001

LC-1H NMR used for determination of the elution order of S-naproxen glucuronide isomers in two isocratic reversed-phase LC-systems.

The reactive metabolite S-naproxen-beta-1-O-acyl glucuronide was purified from human urine using solid phase extraction (SPE) and preparative HPLC. The structure was confirmed by 600 MHz 1H NMR. Directly coupled 600 MHz HPLC-1H NMR was used to assign the peaks in chromatograms obtained when analysing a sample containing S-naproxen aglycone and the 1-, 2-, 3-, and 4-isomers of S-naproxen-beta-1-O-acyl glucuronide in two simple isocratic reversed phase HPLC-systems. Using mobile phase 1 (50 mM formate buffer pH 5.75/acetonitrile 75:25 v/v) the elution order was: 4-O-acyl isomers, beta-1-O-acyl glucuronide, 3-O-acyl isomers, 2-O-acyl isomers, and S-naproxen aglycone. Using mobile phase II (25 mM potassium phosphate pH 7.40/acetonitrile 80:20 v/v) the elution order was: alpha/beta-4-O-acyl isomers, S-naproxen aglycone, beta-1-O-acyl glucuronide, 3-O-acyl isomers, and alpha/beta-2-O-acyl isomers. In both systems the elution order for the 2-, 3- and 4-O-acyl isomers corresponded with previously published results for 2-, 3-, and 4-fluorobenzoic acid glucuronide isomers determined by reversed phase HPLC-1H NMR (U.G. Sidelmann, S.H. Hansen, C. Gavaghan, A.W. Nicholls, H.A.J. Carless, J.C. Lindon, I.D. Wilson, J.K. Nicholson, J. Chromatogr. B Biomed. Appl. 685 (1996) 113-122]. The alpha-1-O-acyl isomer was found to be present at approximately 3% of the initial S-naproxen-beta-1-O-acyl glucuronide concentration in the glucuronide isomer mixture after 6 h of incubation at pH 7.40 and 37 degrees C. In both HPLC systems it eluted just before the beta-1-O-acyl glucuronide well separated from other isomers. Investigators should consider the possible formation of a alpha-1-O-acyl isomer when studying glucuronide reactivity and degradation.

Chromatography, High Pressure Liquid↗

PubMed2001

Directly coupled HPLC-NMR and HPLC-MS approaches for the rapid characterisation of drug metabolites in urine: application to the human metabolism of naproxen.

High resolution nuclear magnetic resonance (NMR) spectroscopy is a very powerful tool for the structural identification of xenobiotic metabolites in complex biological matrices such as plasma, urine and bile. However, these fluids are dominated by thousands of signals resulting from endogenous metabolites and it is advantageous when investigating drug metabolites in such matrices to simplify the spectra by including a separation step in the experiment by directly-coupling HPLC and NMR. Naproxen (6-methoxy-alpha-methyl-2-naphthyl acetic acid) is administered as the S-enantiomer and is metabolised in vivo to form its demethylated metabolite which is subsequently conjugated with beta-D-glucuronic acid as well as with sulfate. Naproxen is also metabolised by phase II metabolism directly to form a glycine conjugate as well as a glucuronic acid conjugate at the carboxyl group. In the present investigation, the metabolism of naproxen was investigated in urine samples with a very simple sample preparation using a combination of directly-coupled HPLC-1H NMR spectroscopy and HPLC-mass spectrometry (MS). A buffer system was developed which allows the same chromatographic method to be used for the HPLC-NMR as well as the HPLC-MS analysis. The combination of these methods is complementary in information content since the NMR spectra provide evidence to distinguish isomers such as the type of glucuronides formed, and the HPLC-MS data allow identification of molecules containing NMR-silent fragments such as occur in the sulfate ester.

Buffers↗

PubMed2001

Metabonomic characterization of genetic variations in toxicological and metabolic responses using probabilistic neural networks.

Current emphasis on efficient screening of novel therapeutic agents in toxicological studies has resulted in the evaluation of novel analytical technologies, including genomic (transcriptomic) and proteomic approaches. We have shown that high-resolution 1H NMR spectroscopy of biofluids and tissues coupled with appropriate chemometric analysis can also provide complementary data for use in in vivo toxicological screening of drugs. Metabonomics concerns the quantitative analysis of the dynamic multiparametric metabolic response of living systems to pathophysiological stimuli or genetic modification [Nicholson, J. K., Lindon, J. C., and Holmes, E. (1999) Xenobiotica 11, 1181-1189]. In this study, we have used 1H NMR spectroscopy to characterize the time-related changes in the urinary metabolite profiles of laboratory rats treated with 13 model toxins and drugs which predominantly target liver or kidney. These 1H NMR spectra were data-reduced and subsequently analyzed using a probabilistic neural network (PNN) approach. The methods encompassed a database of 1310 samples, of which 583 comprised a training set for the neural network, with the remaining 727 (independent cases) employed as a test set for validation. Using these techniques, the 13 classes of toxicity, together with the variations associated with strain, were distinguishable to >90%. Analysis of the 1H NMR spectral data by multilayer perceptron networks and principal components analysis gave a similar but less accurate classification than PNN analysis. This study has highlighted the value of probabilistic neural networks in developing accurate NMR-based metabonomic models for the prediction of xenobiotic-induced toxicity in experimental animals and indicates possible future uses in accelerated drug discovery programs. Furthermore, the sensitivity of this tool to strain differences may prove to be useful in investigating the genetic variation of metabolic responses and for assessing the validity of specific animal models.

Animals↗

PubMed2001

Metabonomic investigations into hydrazine toxicity in the rat.

The systemic biochemical effects of oral hydrazine administration (dosed at 75, 90, and 120 mg/kg) have been investigated in male Han Wistar rats using metabonomic analysis of (1)H NMR spectra of urine and plasma, conventional clinical chemistry, and liver histopathology. Plasma samples were collected both pre- and 24 h postdose, while urine was collected predose and daily over a 7 day postdose period. (1)H NMR spectra of the biofluids were analyzed visually and via pattern recognition using principal component analysis. The latter showed that there was a dose-dependent biochemical effect of hydrazine treatment on the levels of a range of low molecular weight compounds in urine and plasma, which was correlated with the severity of the hydrazine induced liver lesions. In plasma, increases in the levels of free glycine, alanine, isoleucine, valine, lysine, arginine, tyrosine, citrulline, 3-D-hydroxybutyrate, creatine, histidine, and threonine were observed. Urinary excretion of hippurate, citrate, succinate, 2-oxoglutarate, trimethylamine-N-oxide, fumarate and creatinine were decreased following hydrazine dosing, whereas taurine, creatine, threonine, N-methylnicotinic acid, tyrosine, beta-alanine, citrulline, Nalpha-acetylcitrulline and argininosuccinate excretion was increased. Moreover, the most notable effect was the appearance in urine and plasma of 2-aminoadipate, which has previously been shown to lead to neurological effects in rats. High urinary levels of 2-aminoadipate may explain the hitherto poorly understood neurological effects of hydrazine. Metabonomic analysis of high-resolution (1)H NMR spectra of biofluids has provided a means of monitoring the progression of toxicity and recovery, while also allowing the identification of novel biomarkers of development and regression of the lesion.

Administration, Oral↗

PubMed2001

HPLC/1H NMR spectroscopic studies of the reactive alpha-1-O-acyl isomer formed during acyl migration of S-naproxen beta-1-O-acyl glucuronide.

A widely held view in drug metabolism and pharmacokinetic studies is that the initial 1-isomer to 2-isomer step in the intramolecular acyl migration of drug ester glucuronides is irreversible, and that alpha-1-O-acyl isomers do not occur under physiological conditions. We investigated this hypothesis using high-performance liquid chromatography directly coupled to proton nuclear magnetic resonance spectroscopy (HPLC/1H NMR) and mass spectrometry (LC/MS) to probe the migration reactions of S-naproxen beta-1-O-acyl glucuronide, in phosphate buffer at pH 7.4, 37 degrees C. We report the first direct observation of the alpha-1-O-acyl isomer of a drug ester glucuronide (S-naproxen) formed in a biosystem via the facile acyl migration of the corresponding pure beta-1-O-acyl glucuronide. The unequivocal identification of the reactive product was achieved using stopped-flow one-dimensional HPLC/1H NMR and two-dimensional 1H-1H total correlation spectroscopy (1H-1H TOCSY). Parallel LC/ion-trap mass spectrometry yielded the confirmatory glucuronide masses. Moreover, "dynamic" stopped-flow HPLC/1H NMR experiments revealed transacylation of the isolated alpha-1-O-acyl isomer to a mixture of alpha/beta-2-O-acyl isomers; the reverse reaction from the isolated alpha/beta-2-O-acyl isomers to the alpha-1-O-acyl isomer was also clearly demonstrated. This application of "dynamic" stopped-flow HPLC/1H NMR allows key kinetic data to be obtained on a reactive metabolite that would otherwise be difficult to follow by conventional HPLC and NMR methods where sample preparation and off-line separations are necessary. These data challenge the widely held view that the alpha-1-O-acyl isomers of drug ester glucuronides do not occur under physiological conditions. Furthermore, the similar formation of alpha-1-O-acyl isomers from zomepirac and diflunisal beta-1-O-acyl glucuronides has recently been confirmed (Corcoran et al., unpublished results). Such reactions are also likely to be widespread for other drugs that form ester glucuronides in biological systems. Ultimately, the presence of significant quantities of the kinetically labile alpha-1-O-acyl glucuronide isomer may also have toxicological implications in terms of reactivity toward cellular proteins.

Acylation↗

PubMed2001

NMR and pattern recognition studies on the time-related metabolic effects of alpha-naphthylisothiocyanate on liver, urine, and plasma in the rat: an integrative metabonomic approach.

We present here a novel integrative metabonomic approach to probe toxic effects of drugs in experimental animals using alpha-naphthylisothiocyanate (ANIT) as a model hepatotoxicant. Male Han-Wistar rats were dosed with ANIT (150 mg/kg, n = 25), and plasma and liver samples were collected for NMR and magic-angle spinning (MAS) NMR spectroscopy at 3, 7, 24, 31, and 168 h postdosing. Urine was collected continuously for 3 days prior to dosing and up to 168 h postdose. Histopathology and plasma clinical chemistry was also performed at all time points. Liver samples were analyzed either intact by 600 MHz 1H MAS NMR techniques or using high resolution (liquid state) 1H NMR of water-acetonitrile extracts. These data were related to sequential 1H NMR measurements in urine and plasma using pattern recognition methods. 1D 1H NMR spectra were data-reduced and analyzed using principal components analysis (PCA) to show the time-dependent biochemical variations induced by ANIT toxicity. From the eigenvector loadings of the PCA, those regions of the 1H NMR spectra and hence the combinations of endogenous metabolites marking the main phase of the toxic episode were identified. The ANIT-induced biochemical manifestations included a hepatic lipidosis associated with hyperlipidaemia; hyperglycaemia and glycosuria; increased urinary excretion of taurine and creatine; a shift in energy metabolism characterized by increased plasma ketone bodies with reduced urinary excretion of tricarboxylic acid cycle intermediates and raised hepatic bile acids leading to bile aciduria. The integration of metabolic data derived from several sources gives a holistic approach to the study of time-related toxic effects in the intact system and enables the characterization of key metabolic effects during the development and recovery from a toxic lesion.

1-Naphthylisothiocyanate↗

PubMed2001

Metabolic profiling of chronic cadmium exposure in the rat.

A confounding problem with studying the effects of environmental exposure to contaminants in wild populations is that analytical techniques are invasive, particularly where the physiological effects of the toxin are assessed. In this study, a metabonomic approach to investigate the biochemical effects of chronic oral exposure to environmentally realistic doses of CdCl2 (low, 8 mg/kg; high, 40 mg/kg) is presented. 1H NMR spectra of urine from exposed animals were analyzed using pattern recognition methods to identify biomarkers for a 94 day exposure period. Creatinuria and both increased excretion and complexation of citrate was detected after 19 days of exposure in both exposure groups. This was accompanied by a decrease in plasma Ca2+/Mg2+ ratio in blood plasma after 94 days. Post mortem, magic angle spinning (MAS) 1H NMR spectroscopy was used alongside conventional analytical techniques to investigate intact tissue directly. According to atomic absorption spectroscopy, kidney tissue accumulated 26.8 +/- 2.5 microg of Cd2+/g dry wt (low) and 75.9 +/- 4.3 microg of Cd2+/g dry wt (high). Using high-resolution MAS 1H NMR spectroscopy altered lipid content was detected in kidneys from animals exposed to Cd2+. However, unlike acute exposure, no testicular damage was evident. This systemic approach to metabolism demonstrated the different physiological effects of chronic subacute compared with an acute exposure to Cd2+.

Administration, Oral↗

PubMed2001

Investigation of water environments in a C18 bonded silica phase using 1H magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopy.

High resolution 1H magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectra have been obtained on typical C18 bonded silicas used in chromatographic solid-phase extraction separations. It has been shown for the first time that water molecules distributed in distinct physico-chemical environments within the chromatographic system can be detected directly using a simple 1H MAS NMR measurement. The resonances assigned to water protons in differing physico-chemical environments have distinct chemical shifts, line widths, relaxation times (T1 and T2) and also exhibit temperature dependent coalescence behaviour. This novel MAS approach may lead to a better understanding of the environments of other analytes in mixtures during such separations.

Journal Article↗

PubMed2001

The potential of 19F NMR spectroscopy for rapid screening of cell cultures for models of mammalian drug metabolism.

The use of microbial cultures as a complementary model for mammalian drug metabolism has been well established previously. Here is a preliminary investigation into the potential of 19F NMR spectroscopy as a rapid screening tool to quantify the biotransformations of fluorine-containing model drugs. Biotransformations of three model drugs in 48 taxonomically diverse organisms were measured by acquiring 19F NMR spectra at 376 MHz. The presence of fluorine in the molecules allowed rapid, simultaneous detection of over 20 biotransformation products without sample pretreatment, chromatography, mass spectrometric techniques or the use of radiolabelled substrates. The detection limit at 376 MHz using 5 mm NMR tubes was ca. 0.3 microg ml(-1) using a typical analysis time of 20 min per sample. With the recent advent of flow injection NMR technology, analysis time of 5 min could be achieved with less sample. This approach may be used to develop fast small-scale microbial screens for the biosynthesis of metabolite standards and production of novel drug analogues, whilst also having a role in reducing animal experiments needed to identify animal and human metabolites of fluorinated xenobiotics.

Animals↗

PubMed2001

High-resolution magic angle spinning 1H-NMR spectroscopy studies on the renal biochemistry in the bank vole (Clethrionomys glareolus) and the effects of arsenic (As3+) toxicity.

1. High-resolution magic angle spinning (MAS) 1H-NMR spectroscopy was used to study renal metabolism and the toxicity of As3+, a common environmental contaminant, in the bank vole (Clethrionomys glareolus), a wild species of rodent. 2. Following a 14-day exposure to an environmentally relevant dose of As2O3 (28 mg kg(-1) feed), voles displayed tissue damage at autopsy. MAS 1H spectra indicated abnormal lipid profiles in these samples. 3. Tissue necrosis was also evident from measurements of the apparent diffusion coefficient of water in the intact tissue using MAS 1H diffusion-weighted spectroscopy, its first application to toxicology. 4. Comparison of renal tissue from the wood mouse (Apodemus sylvaticus) exposed to identical exposure levels of As3+ suggested that the bank vole is particularly vulnerable to As3+ toxicity.

Alanine↗

PubMed2001

S-naproxen-beta-1-O-acyl glucuronide degradation kinetic studies by stopped-flow high-performance liquid chromatography-1H NMR and high-performance liquid chromatography-UV.

Acyl-migrated isomers of drug beta-1-O-acyl glucuronides have been implicated in drug toxicity because they can bind to proteins. The acyl migration and hydrolysis of S-naproxen-beta-1-O-acyl glucuronide (S-nap-g) was followed by dynamic stopped-flow HPLC-1H NMR and HPLC methods. Nine first order rate constants in the chemical equilibrium between six species (S-nap-g, its alpha/beta-2-O-acyl, alpha/beta-3-O-acyl, alpha/beta-4-O-acyl, and alpha-1-O-acyl-migration isomers, and S-naproxen aglycone) were determined by HPLC-UV studies in 25 mM potassium phosphate buffer, pH 7.40, 25 mM potassium phosphate buffer in D2O pD 7.40, and 25 mM potassium phosphate buffer in D2O pD 7.40/MeCN 80:20 v/v (HPLC-1H NMR mobile phase). In the 25 mM potassium phosphate buffer (pH 7.40) the acyl-migration rate constants (h(-1)) were 0.18 (S-nap-g-alpha/beta-2-O-acyl isomer), 0.23 (alpha/beta-2-O-acyl-alpha-1-O-acyl), 2.6 (alpha-1-O-acyl-alpha/beta-2-O-acyl), 0.12 (alpha/beta-2-O-acyl-alpha/beta-3-O-acyl), 0.048 (alpha/beta-3-O-acyl-alpha/beta-2-O-acyl), 0.059 (alpha/beta-3-O-acyl-alpha/beta-4-O-acyl), and 0.085 (alpha/beta-4-O-acyl-alpha/beta-3-O-acyl). The hydrolysis rate constants (h(-1)) were 0.025 (hydrolysis of S-nap-g) and 0.0058 (hydrolysis of all acyl-migrated isomers). D2O and MeCN decreased the magnitude of all nine kinetic rate constants by up to 80%. The kinetic rate constants for the degradation of S-nap-g in the mobile phase used for HPLC-1H NMR determined using HPLC-UV could predict the results obtained by the dynamic stopped-flow HPLC-1H NMR experiments of the individual acyl-migrated isomers. It is therefore recommended that beta-1-O-acyl glucuronide degradation kinetics be investigated by HPLC-UV methods once the identification and elution order of the isomers have been established by HPLC-1H NMR.

Acetonitriles↗

PubMed2001

An update on the use of flow cytometry in HIV infection and AIDS.

Flow cytometry has played an invaluable role in recent advances made against HIV and AIDS, and there is every reason to expect it will continue to do so. Newer-generation machines, using three- and four-color panels, make measurements of lymphocyte subsets ever more accurate and potentially cost-effective. Similarly, recent single-platform techniques promise even more efficiencies, freeing CD4 enumeration from parallel determination of the CBC. In other developments, quantitative reporting of immunophenotype may well change the way subsets traditionally defined by qualitative determinations are viewed. The most promising initial candidates for such changes are surrogates of immune activation, such as CD38 expression on CD8 cells. But technical demonstration of reproducibility and reliability, and clinical trial evidence of utility--especially as measured against such established laboratory parameters as CD4 cell and HIV RNA determinations--are needed before a change from qualitative to quantitative immunophenotype measurement can be widely accepted. Although too soon to tell if they will emerge from the research arena into the clinical arena, tetramer binding, intracellular cytokine detection, and assays of cell division are rapidly advancing understanding of HIV-disease, and indeed understanding of all of human immunology. Some of these assays provide often subtly different information, and together they will help determine the most important surrogate measures of clinical immunity. Identifying such surrogates is critical to developing a rational HIV vaccine. The power of flow cytometry is its ability simultaneously to analyze data in four, five, six, or more dimensions. For longitudinal experiments, time adds yet another dimension. Humans struggle to think past three dimensions; interpreting results from experimental permutations and combinations that expand geometrically with each additional parameter studied requires considerable effort. Ultimately computer programs might come to our aid, but for now we have to live with the information overload if we hope to gain insight into the workings of complex biologic systems. In a world of limited resources, and with limited capacity to conceptualize in multiple dimensions, insight, inspiration, and perhaps a little luck are needed to ask the right questions and use the right assays if the recent string of advances against AIDS is to continue.

Acquired Immunodeficiency Syndrome↗

PubMed2000

The biochemical profile of rat testicular tissue as measured by magic angle spinning 1H NMR spectroscopy.

The testis is the principal organ of male fertility, responsible for the production of spermatozoa and their maturation into sperm. However, the underlying biochemistry of the testis is relatively understudied. The fluidic and homogeneous nature of the testis makes it an ideal organ for high resolution magic angle spinning (MAS) 1H NMR spectroscopy. In this study we have catalogued the low molecular weight metabolites. The testis contains large amounts of creatine, of which a substantial proportion was shown to be extracellular using bipolar gradients to measure apparent diffusion coefficients. The tissue also contained relatively high amounts of uridine.

Alanine↗

PubMed2000

An NMR-based metabonomic approach to investigate the biochemical consequences of genetic strain differences: application to the C57BL10J and Alpk:ApfCD mouse.

As the human genome sequencing projects near completion, there is an active search for technologies that can provide insights into the genetic basis for physiological variation and interpreting gene expression in terms of phenotype at the whole organism level in order to understand the pathophysiology of disease. We present a novel metabonomic approach to the investigation of genetic influences on metabolic balance and metabolite excretion patterns in two phenotypically normal mouse models (C57BL10J and Alpk:ApfCD). Chemometric techniques were applied to optimise recovery of biochemical information from complex (1)H NMR urine spectra and to determine metabolic biomarker differences between the two strains. Differences were observed in tricarboxylic acid cycle intermediates and methylamine pathway activity. We suggest here a new 'metabotype' concept, which will be of value in relating quantitative physiological and biochemical data to both phenotypic and genetic variation in animals and man.

Amino Acids↗