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1.
Sci Rep ; 9(1): 13067, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31506554

RESUMO

Inborn errors of metabolism (IEMs) are rare diseases produced by the accumulation of abnormal amounts of metabolites, toxic to the newborn. When not detected on time, they can lead to irreversible physiological and psychological sequels or even demise. Metabolomics has emerged as an efficient and powerful tool for IEM detection in newborns, children, and adults with late onset. In here, we screened urine samples from a large set of neonates (470 individuals) from a homogeneous population (Basque Country), for the identification of congenital metabolic diseases using NMR spectroscopy. Absolute quantification allowed to derive a probability function for up to 66 metabolites that adequately describes their normal concentration ranges in newborns from the Basque Country. The absence of another 84 metabolites, considered abnormal, was routinely verified in the healthy newborn population and confirmed for all but 2 samples, of which one showed toxic concentrations of metabolites associated to ketosis and the other one a high trimethylamine concentration that strongly suggested an episode of trimethylaminuria. Thus, a non-invasive and readily accessible urine sample contains enough information to assess the potential existence of a substantial number (>70) of IEMs in newborns, using a single, automated and standardized 1H- NMR-based analysis.

2.
Magn Reson Chem ; 57(9): 579-588, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-30680787

RESUMO

Both the German and European organic food markets are growing fast, and there is also a rising demand for organic chicken eggs. Consumers are willing to pay higher prices for organic eggs produced in an animal-appropriate environment considering animal welfare. Strict labelling requirements do not prevent chicken eggs from being a subject of food fraud. Conventionally produced (barn/free-range) eggs can easily be mislabeled as organic eggs. Especially because the demand for organically produced chicken eggs is likely to exceed supply in the future, mislabeling appears to be a realistic scenario. Therefore, there is a need for analytical methods that are suitable to classify eggs as being either conventionally or organically produced. Nuclear magnetic resonance (NMR) spectroscopy in combination with multivariate data analysis is a suitable tool to screen eggs according to the different systems of husbandry. Sample preparation is based on a fat extraction method, which was optimised for application to freeze-dried egg yolk. Samples were analysed using typical q-NMR parameters. A nontargeted approach was used for the analysis of the 1 H NMR data. Principal component analysis (PCA) was applied followed by a linear discriminant analysis (PCA-LDA) and Monte Carlo cross-validation. In total, 344 chicken eggs (214 barn/free-range eggs and 130 eggs from organic farms), most of them originating from Germany, were used to build and validate the prediction model. The results showed that the prediction model allowed for the correct classification of about 93% of the organic eggs.


Assuntos
Ovos/análise , Análise de Alimentos/métodos , Alimentos Orgânicos/análise , Espectroscopia de Ressonância Magnética/métodos , Animais , Galinhas , Análise Discriminante , Gema de Ovo/química , Qualidade dos Alimentos , Alemanha , Método de Monte Carlo , Análise Multivariada , Agricultura Orgânica
3.
Anal Chem ; 90(20): 11962-11971, 2018 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-30211542

RESUMO

We report an extensive 600 MHz NMR trial of quantitative lipoprotein and small-molecule measurements in human blood serum and plasma. Five centers with eleven 600 MHz NMR spectrometers were used to analyze 98 samples including 20 quality controls (QCs), 37 commercially sourced, paired serum and plasma samples, and two National Institute of Science and Technology (NIST) reference material 1951c replicates. Samples were analyzed using rigorous protocols for sample preparation and experimental acquisition. A commercial lipoprotein subclass analysis was used to quantify 105 lipoprotein subclasses and 24 low molecular weight metabolites from the NMR spectra. For all spectrometers, the instrument specific variance in measuring internal QCs was lower than the percentage described by the National Cholesterol Education Program (NCEP) criteria for lipid testing [triglycerides <2.7%; cholesterol <2.8%; low-density lipoprotein (LDL) cholesterol <2.8%; high-density lipoprotein (HDL) cholesterol <2.3%], showing exceptional reproducibility for direct quantitation of lipoproteins in both matrixes. The average relative standard deviations (RSDs) for the 105 lipoprotein parameters in the 11 instruments were 4.6% and 3.9% for the two NIST samples, whereas they were 38% and 40% for the 37 commercially sourced plasmas and sera, respectively, showing negligible analytical compared to biological variation. The coefficient of variance (CV) obtained for the quantification of the small molecules across the 11 spectrometers was below 15% for 20 out of the 24 metabolites analyzed. This study provides further evidence of the suitability of NMR for high-throughput lipoprotein subcomponent analysis and small-molecule quantitation with the exceptional required reproducibility for clinical and other regulatory settings.


Assuntos
Lipoproteínas/sangue , Ressonância Magnética Nuclear Biomolecular , Humanos , Laboratórios , Lipoproteínas/metabolismo , Peso Molecular , Prótons , Controle de Qualidade
4.
Nat Commun ; 8(1): 1662, 2017 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-29162796

RESUMO

The NMR chemical shifts of a substance in a complex mixture strongly depend on the composition of the mixture itself, as many weak interactions occur that are hardly predictable. Chemical shift variability is the major obstacle to automatically assigning, and subsequently quantitating, metabolite signals in body fluids, particularly urine. Here we demonstrate that the chemical shifts of signals in urine are actually predictable. This is achieved by constructing ca. 4000 artificial mixtures where the concentrations of 52 most abundant urine metabolites-including 11 inorganic ions-are varied, to sparsely but efficiently populate an N-dimensional concentration matrix. A strong relationship is established between the concentration matrix and the chemical shift matrix, so that chemical shifts of > 90 metabolite signals can be accurately predicted in real urine samples. The concentrations of the invisible inorganic ions are also accurately predicted, along with those of albumin and of several other abundant urine components.


Assuntos
Urina/química , Adulto , Idoso , Feminino , Humanos , Espectroscopia de Ressonância Magnética , Masculino , Metabolômica , Pessoa de Meia-Idade , Adulto Jovem
5.
Anal Chem ; 89(15): 8004-8012, 2017 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-28692288

RESUMO

Lipoprotein profiling of human blood by 1H nuclear magnetic resonance (NMR) spectroscopy is a rapid and promising approach to monitor health and disease states in medicine and nutrition. However, lack of standardization of measurement protocols has prevented the use of NMR-based lipoprotein profiling in metastudies. In this study, a standardized NMR measurement protocol was applied in a ring test performed across three different laboratories in Europe on plasma and serum samples from 28 individuals. Data was evaluated in terms of (i) spectral differences, (ii) differences in LPD predictions obtained using an existing prediction model, and (iii) agreement of predictions with cholesterol concentrations in high- and low-density lipoproteins (HDL and LDL) particles measured by standardized clinical assays. ANOVA-simultaneous component analysis (ASCA) of the ring test spectral ensemble that contains methylene and methyl peaks (1.4-0.6 ppm) showed that 97.99% of the variance in the data is related to subject, 1.62% to sample type (serum or plasma), and 0.39% to laboratory. This interlaboratory variation is in fact smaller than the maximum acceptable intralaboratory variation on quality control samples. It is also shown that the reproducibility between laboratories is good enough for the LPD predictions to be exchangeable when the standardized NMR measurement protocol is followed. With the successful implementation of this protocol, which results in reproducible prediction of lipoprotein distributions across laboratories, a step is taken toward bringing NMR more into scope of prognostic and diagnostic biomarkers, reducing the need for less efficient methods such as ultracentrifugation or high-performance liquid chromatography (HPLC).


Assuntos
Lipoproteínas HDL/sangue , Lipoproteínas LDL/sangue , Espectroscopia de Prótons por Ressonância Magnética , Adulto , Feminino , Humanos , Laboratórios/normas , Análise dos Mínimos Quadrados , Lipoproteínas VLDL/sangue , Gravidez , Análise de Componente Principal , Espectroscopia de Prótons por Ressonância Magnética/normas , Adulto Jovem
6.
J Proteome Res ; 15(1): 311-25, 2016 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-26566167

RESUMO

This work assesses the urinary metabolite signature of prematurity in newborns by nuclear magnetic resonance (NMR) spectroscopy, while establishing the role of possible confounders and signature specificity, through comparison to other disorders. Gender and delivery mode are shown to impact importantly on newborn urine composition, their analysis pointing out at specific metabolite variations requiring consideration in unmatched subject groups. Premature newborns are, however, characterized by a stronger signature of varying metabolites, suggestive of disturbances in nucleotide metabolism, lung surfactants biosynthesis and renal function, along with enhancement of tricarboxylic acid (TCA) cycle activity, fatty acids oxidation, and oxidative stress. Comparison with other abnormal conditions (respiratory depression episode, large for gestational age, malformations, jaundice and premature rupture of membranes) reveals that such signature seems to be largely specific of preterm newborns, showing that NMR metabolomics can retrieve particular disorder effects, as well as general stress effects. These results provide valuable novel information on the metabolic impact of prematurity, contributing to the better understanding of its effects on the newborn's state of health.


Assuntos
Nascimento Prematuro/urina , Síndrome do Desconforto Respiratório do Recém-Nascido/urina , Adolescente , Adulto , Biomarcadores/urina , Estudos de Casos e Controles , Feminino , Transtornos do Crescimento/urina , Humanos , Recém-Nascido , Masculino , Idade Materna , Metaboloma , Gravidez , Urinálise/métodos , Adulto Jovem
7.
J Nat Prod ; 78(5): 977-86, 2015 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-25946005

RESUMO

Isatis tinctoria is an ancient dye and medicinal plant with potent anti-inflammatory and antiallergic properties. Metabolic differences were investigated by NMR spectroscopy of accessions from different origins that were grown under identical conditions on experimental plots. For these accessions, metabolite profiles at different harvesting dates were analyzed, and single and repeatedly harvested plants were compared. Leaf samples were shock-frozen in liquid N2 immediately after being harvested, freeze-dried, and cryomilled prior to extraction. Extracts were prepared by pressurized liquid extraction with ethyl acetate and 70% aqueous methanol. NMR spectra were analyzed using a combination of different methods of multivariate data analysis such as principal component analysis (PCA), canonical analysis (CA), and k-nearest neighbor concept (k-NN). Accessions and harvesting dates were well separated in the PCA/CA/k-NN analysis in both extracts. Pairwise statistical total correlation spectroscopy (STOCSY) revealed unsaturated fatty acids, porphyrins, carbohydrates, indole derivatives, isoprenoids, phenylpropanoids, and minor aromatic compounds as the cause of these differences. In addition, the metabolite profile was affected by the repeated harvest regime, causing a decrease of 1,5-anhydroglucitol, sucrose, unsaturated fatty acids, porphyrins, isoprenoids, and a flavonoid.


Assuntos
Isatis/química , Metabolômica , Plantas Medicinais/química , Desoxiglucose/análise , Ácidos Graxos Insaturados/análise , Flavonoides/análise , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Folhas de Planta/química , Porfirinas/análise , Sacarose/análise , Terpenos/análise
8.
Anal Chem ; 86(19): 9887-94, 2014 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-25180432

RESUMO

Proton nuclear magnetic resonance (NMR)-based metabolic phenotyping of urine and blood plasma/serum samples provides important prognostic and diagnostic information and permits monitoring of disease progression in an objective manner. Much effort has been made in recent years to develop NMR instrumentation and technology to allow the acquisition of data in an effective, reproducible, and high-throughput approach that allows the study of general population samples from epidemiological collections for biomarkers of disease risk. The challenge remains to develop highly reproducible methods and standardized protocols that minimize technical or experimental bias, allowing realistic interlaboratory comparisons of subtle biomarker information. Here we present a detailed set of updated protocols that carefully consider major experimental conditions, including sample preparation, spectrometer parameters, NMR pulse sequences, throughput, reproducibility, quality control, and resolution. These results provide an experimental platform that facilitates NMR spectroscopy usage across different large cohorts of biofluid samples, enabling integration of global metabolic profiling that is a prerequisite for personalized healthcare.


Assuntos
Espectroscopia de Ressonância Magnética/normas , Metaboloma , Metabolômica/normas , Prótons , Manejo de Espécimes/normas , Biomarcadores/sangue , Biomarcadores/urina , Humanos , Espectroscopia de Ressonância Magnética/instrumentação , Metabolômica/instrumentação , Controle de Qualidade , Reprodutibilidade dos Testes
9.
JIMD Rep ; 16: 101-11, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25012580

RESUMO

Approximately 1 in 400 neonates in Turkey is affected by inherited metabolic diseases. This high prevalence is at least in part due to consanguineous marriages. Standard screening in Turkey now covers only three metabolic diseases (phenylketonuria, congenital hypothyroidism, and biotinidase deficiency). Once symptoms have developed, tandem-MS can be used, although this currently covers only up to 40 metabolites. NMR potentially offers a rapid and versatile alternative.We conducted a multi-center clinical study in 14 clinical centers in Turkey. Urine samples from 989 neonates were collected and investigated by using NMR spectroscopy in two different laboratories. The primary objective of the present study was to explore the range of variation of concentration and chemical shifts of specific metabolites without clinically relevant findings that can be detected in the urine of Turkish neonates. The secondary objective was the integration of the results from a healthy reference population of neonates into an NMR database, for routine and completely automatic screening of congenital metabolic diseases.Both targeted and untargeted analyses were performed on the data. Targeted analysis was aimed at 65 metabolites. Limits of detection and quantitation were determined by generating urine spectra, in which known concentrations of the analytes were added electronically as well as by real spiking. Untargeted analysis involved analysis of the whole spectrum for abnormal features, using statistical procedures, including principal component analysis. Outliers were eliminated by model building. Untargeted analysis was used to detect known and unknown compounds and jaundice, proteinuria, and acidemia. The results will be used to establish a database to detect pathological concentration ranges and for routine screening.

10.
Anal Chim Acta ; 833: 29-39, 2014 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-24909771

RESUMO

It is known that (1)H NMR spectroscopy represents a good tool for predicting the grape variety, the geographical origin, and the year of vintage of wine. In the present study we have shown that classification models can be improved when (1)H NMR profiles are fused with stable isotope (SNIF-NMR, (18)O, (13)C) data. Variable selection based on clustering of latent variables was performed on (1)H NMR data. Afterwards, the combined data of 718 wine samples from Germany were analyzed using linear discriminant analysis (LDA), partial least squares-discriminant analysis (PLS-DA), factorial discriminant analysis (FDA) and independent components analysis (ICA). Moreover, several specialized multiblock methods (common components and specific weights analysis (ComDim), consensus PCA and consensus PLS-DA) were applied to the data. The best improvement in comparison with (1)H NMR data was obtained for prediction of the geographical origin (up to 100% for the fused data, whereas stable isotope data resulted only in 60-70% correct prediction and (1)H NMR data alone in 82-89% respectively). Certain enhancement was obtained also for the year of vintage (from 88 to 97% for (1)H NMR to 99% for the fused data), whereas in case of grape varieties improved models were not obtained. The combination of (1)H NMR data with stable isotope data improves efficiency of classification models for geographical origin and vintage of wine and can be potentially used for other food products as well.


Assuntos
Vinho/análise , Espectroscopia de Prótons por Ressonância Magnética
11.
J Agric Food Chem ; 61(23): 5610-9, 2013 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-23682581

RESUMO

The authenticity, the grape variety, the geographical origin, and the year of vintage of wines produced in Germany were investigated by (1)H NMR spectroscopy in combination with several steps of multivariate data analysis including principal component analysis (PCA), linear discrimination analysis (LDA), and multivariate analysis of variance (MANOVA) together with cross-validation (CV) embedded in a Monte Carlo resampling approach (MC) and others. A total of about 600 wines were selected and carefully collected from five wine-growing areas in the southern and southwestern parts of Germany. Simultaneous saturation of the resonances of water and ethanol by application of a low-power eight-frequency band irradiation using shaped pulses allowed for high receiver gain settings and hence optimized signal-to-noise ratios. Correct prediction of classification of the grape varieties of Pinot noir, Lemberger, Pinot blanc/Pinot gris, Müller-Thurgau, Riesling, and Gewürztraminer of 95% in the wine panel was achieved. The classification of the vintage of all analyzed wines resulted in correct predictions of 97 and 96%, respectively, for vintage 2008 (n = 318) and 2009 (n = 265). The geographic origin of all wines from the largest German wine-producing regions, Rheinpfalz, Rheinhessen, Mosel, Baden, and Württemberg, could be predicted 89% correctly on average. Each NMR spectrum could be regarded as the individual "fingerprint" of a wine sample, which includes information about variety, origin, vintage, physiological state, technological treatment, and others.


Assuntos
Espectroscopia de Ressonância Magnética/métodos , Vitis/química , Vinho/análise , Análise Discriminante , Geografia , Alemanha , Análise Multivariada , Vitis/classificação , Vinho/estatística & dados numéricos
12.
Anal Chem ; 85(12): 5801-9, 2013 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-23718684

RESUMO

Metabolism is essential to understand human health. To characterize human metabolism, a high-resolution read-out of the metabolic status under various physiological conditions, either in health or disease, is needed. Metabolomics offers an unprecedented approach for generating system-specific biochemical definitions of a human phenotype through the capture of a variety of metabolites in a single measurement. The emergence of large cohorts in clinical studies increases the demand of technologies able to analyze a large number of measurements, in an automated fashion, in the most robust way. NMR is an established metabolomics tool for obtaining metabolic phenotypes. Here, we describe the analysis of NMR-based urinary profiles for metabolic studies, challenged to a large human study (3007 samples). This method includes the acquisition of nuclear Overhauser effect spectroscopy one-dimensional and J-resolved two-dimensional (J-Res-2D) (1)H NMR spectra obtained on a 600 MHz spectrometer, equipped with a 120 µL flow probe, coupled to a flow-injection analysis system, in full automation under the control of a sampler manager. Samples were acquired at a throughput of ~20 (or 40 when J-Res-2D is included) min/sample. The associated technical analysis error over the full series of analysis is 12%, which demonstrates the robustness of the method. With the aim to describe an overall metabolomics workflow, the quantification of 36 metabolites, mainly related to central carbon metabolism and gut microbial host cometabolism, was obtained, as well as multivariate data analysis of the full spectral profiles. The metabolic read-outs generated using our analytical workflow can therefore be considered for further pathway modeling and/or biological interpretation.


Assuntos
Automação Laboratorial/métodos , Espectroscopia de Ressonância Magnética/métodos , Metaboloma/fisiologia , Urinálise/métodos , Adulto , Idoso , Automação Laboratorial/normas , Feminino , Análise de Injeção de Fluxo/métodos , Análise de Injeção de Fluxo/normas , Humanos , Espectroscopia de Ressonância Magnética/normas , Masculino , Pessoa de Meia-Idade , Urinálise/normas
13.
J Magn Reson ; 228: 81-94, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23357430

RESUMO

Metabolites and their concentrations are direct reporters on body biochemistry. Thanks to technical developments metabolic profiling of body fluids, such as blood plasma, by for instance NMR has in the past decade become increasingly accurate enabling successful clinical diagnostics. Human Serum Albumin (HSA) is the main plasma protein (∼60% of all plasma protein) and responsible for the transport of endogenous (e.g. fatty acids) and exogenous metabolites, which it achieves thanks to its multiple binding sites and its flexibility. HSA has been extensively studied with regard to its binding of drugs (exogenous metabolites), but only to a lesser extent with regard to its binding of endogenous (non-fatty acid) metabolites. To obtain correct NMR measured metabolic profiles of blood plasma and/or potentially extract information on HSA and fatty acids content, it is necessary to characterize these endogenous metabolite/plasma protein interactions. Here, we investigate these metabolite-HSA interactions in blood plasma and blood plasma mimics. The latter contain the roughly twenty metabolites routinely detected by NMR (also most abundant) in normal relative concentrations with fatted or non-fatted HSA added or not. First, we find that chemical shift changes are small and seen only for a few of the metabolites. In contrast, a significant number of the metabolites display reduced resonance integrals and reduced free concentrations in the presence of HSA or fatted HSA. For slow-exchange (or strong) interactions, NMR resonance integrals report the free metabolite concentration, while for fast exchange (weak binding) the chemical shift reports on the binding. Hence, these metabolites bind strongly to HSA and/or fatted HSA, but to a limited degree because for most metabolites their concentration is smaller than the HSA concentration. Most interestingly, fatty acids decrease the metabolite-HSA binding quite significantly for most of the interacting metabolites. We further find that competition between the metabolites for binding is absent for most of these metabolites. These mappings in plasma mimics may thus open new opportunities for improved metabolic profiling of blood plasma. For instance, correct metabolite concentrations can be determined for the non-interacting metabolites and/or concentration corrections made for interacting metabolites. Secondly, the interacting metabolites could be used to act as reporters on HSA and fatty acid concentration in plasma, and thus potentially act as biomarker in diagnostic studies of trauma or cardiovascular diseases. Finally, we find in the blood plasma mimics that after ultrafiltration, commonly used to remove the protein from plasma, the measured concentration equals the total metabolite concentration, except for the strongest binding metabolite citrate.


Assuntos
Proteínas Sanguíneas/metabolismo , Ácidos Graxos/metabolismo , Espectroscopia de Ressonância Magnética/métodos , Plasma/metabolismo , Albumina Sérica/metabolismo , Humanos , Ligação Proteica
14.
Magn Reson Chem ; 49(11): 734-9, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22002683

RESUMO

The 400 MHz (1)H NMR analysis of alcoholic beverages using standard pulse programs lacks the necessary sensitivity to detect minor constituents such as methanol, acetaldehyde or ethyl acetate. This study investigates the application of a shaped pulse sequence during the relaxation delay to suppress the eight (1)H NMR frequencies of water and ethanol (the OH singlet of both water and ethanol, as well as the CH(2) quartet and CH(3) triplet of ethanol). The sequence of reference measurement for frequency determination followed by the suppression experiment is controlled by a macro in the acquisition software so that a measurement under full automation is possible (12 min per sample total time). Additionally, sample preparation was optimized to avoid precipitation, which is facilitated by 1:1 dilution with ethanol and pH 7.4 buffer. Compared with the standard water presaturation pulse program, the eightfold suppression allowed a significantly higher setting of receiver gain without receiver overflow, which significantly increased the signal-to-noise ratio by an average factor of 10. The signal intensities increased by a factor of 20. The resulting limits of detection (below 1 g/hl of pure alcohol) now allow the control of legal requirements for minor compounds in alcoholic beverages.


Assuntos
Bebidas Alcoólicas/análise , Etanol/química , Água/química , Etanol/análise , Espectroscopia de Ressonância Magnética , Prótons , Água/análise
15.
J Proteome Res ; 10(9): 4165-76, 2011 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-21744784

RESUMO

When small B lymphocytes bind antigen in the context of suitable signals, a profound geno-proteomic metamorphosis is activated that generates antibody-secreting cells. To study the metabolic changes associated with this differentiation program, we compared the exometabolome of differentiating murine B lymphoma cells and primary B cells by monodimensional proton nuclear magnetic resonance spectroscopy and mass spectrometry coupled to liquid chromatography. Principal component analysis, a multivariate statistical analysis, highlighted metabolic hallmarks of the sequential differentiation phases discriminating between the proliferation and antibody secreting phases and revealing novel metabolic pathways. During proliferation, lactate production increased together with consumption of essential amino acids; massive Ig secretion was paralleled by alanine and glutamate production, glutamine being used as carbon and energy sources. Notably, ethanol and 5'-methylthioadenosine were produced during the last phase of protein secretion and the proliferative burst, respectively. Our metabolomics results are in agreement with previous genoproteomics studies. Thus, metabolic profiling of extracellular medium is a useful tool to characterize the functional state of differentiating B cells and to identify novel underlying metabolic pathways.


Assuntos
Linfócitos B/citologia , Linfócitos B/metabolismo , Diferenciação Celular/fisiologia , Metaboloma/fisiologia , Metabolômica/métodos , Plasmócitos/citologia , Plasmócitos/metabolismo , Animais , Linhagem Celular Tumoral , Linfoma de Células B/patologia , Espectrometria de Massas , Camundongos , Ressonância Magnética Nuclear Biomolecular , Análise de Componente Principal , Pegadas de Proteínas
16.
Environ Sci Technol ; 45(11): 4710-7, 2011 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-21542577

RESUMO

Dissolved organic matter (DOM) is ubiquitous in aquatic ecosystems and is derived from various inputs that control its turnover. Glaciers and ice sheets are the second largest water reservoir in the global hydrologic cycle, but little is known about glacial DOM composition or contributions to biogeochemical cycling. Here we employ SPR-W5-WATERGATE (1)H NMR spectroscopy to elucidate and quantify the chemical structures of DOM constituents in Antarctic glacial ice as they exist in their natural state (average DOC of 8 mg/L) without isolation or preconcentration. This Antarctic glacial DOM is predominantly composed of a mixture of small recognizable molecules differing from DOM in marine, lacustrine, and other terrestrial environments. The major constituents detected in three distinct types of glacial ice include lactic and formic acid, free amino acids, and a mixture of simple sugars and amino sugars with concentrations that vary between ice types. The detection of free amino acid and amino sugar monomer components of peptidoglycan within the ice suggests that Antarctic glacial DOM likely originates from in situ microbial activity. As these constituents are normally considered to be biologically labile (fast cycling) in nonglacial environments, accelerated glacier melt and runoff may result in a flux of nutrients into adjacent ecosystems.


Assuntos
Camada de Gelo/química , Compostos Orgânicos/análise , Regiões Antárticas , Espectroscopia de Ressonância Magnética
17.
J Proteome Res ; 9(1): 319-32, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19908917

RESUMO

This work aims at characterizing the metabolic profile of human lung cancer, to gain new insights into tumor metabolism and to identify possible biomarkers with potential diagnostic value in the future. Paired samples of tumor and noninvolved adjacent tissues from 12 lung tumors have been directly analyzed by (1)H HRMAS NMR (500/600 MHz) enabling, for the first time to our knowledge, the identification of over 50 compounds. The effect of temperature on tissue stability during acquisition time has also been investigated, demonstrating that analysis should be performed within less than two hours at low temperature (277 K), to minimize glycerophosphocholine (GPC) and phosphocholine (PC) conversion to choline and reduce variations in some amino acids. The application of Principal Component Analysis (PCA) and Hierarchical Cluster Analysis (HCA) to the standard 1D (1)H spectra resulted in good separation between tumor and control samples, showing that inherently different metabolic signatures characterize the two tissue types. On the basis of spectral integration measurements, lactate, PC, and GPC were found to be elevated in tumors, while glucose, myo-inositol, inosine/adenosine, and acetate were reduced. These results show the valuable potential of HRMAS NMR-metabonomics for investigating the metabolic phenotype of lung cancer.


Assuntos
Neoplasias Pulmonares/metabolismo , Ressonância Magnética Nuclear Biomolecular/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Análise por Conglomerados , Feminino , Humanos , Masculino , Metaboloma , Pessoa de Meia-Idade , Análise Multivariada , Análise de Componente Principal
18.
Magn Reson Chem ; 47 Suppl 1: S130-7, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19899106

RESUMO

Nuclear magnetic resonance (NMR) spectroscopy is rapidly gaining importance in mixture analysis, originally driven by the pharmaceutical and nowadays also by clinical applications within metabonomics. Quality control of food-related material has very similar requirements, as it also deals with mixtures, and many of the compounds found in body fluids are analyzed as well. NMR allows analysis in two ways within one experiment: namely, targeted and untargeted. Targeted stands for the safe identification and consequent quantification of individual compounds, whereas untargeted means the detection of all deviations visible by NMR using statistical analysis based on normality models. Very important is the stability and reproducibility of the NMR instrumentation used, and this means inherent minimized system internal variance. NMR is especially suited for such requirements, as it allows detection of the smallest concentration changes of many metabolites simultaneously. High-throughput flow-injection NMR as the basis for fruit juice screening allows low cost per sample and delivers substantially more relevant information than any other method and is probably the only method to produce such results.


Assuntos
Bebidas/normas , Frutas/química , Glucose/química , Ácido Láctico/química , Magnésio/química , Espectroscopia de Ressonância Magnética , Malatos/química , Potássio/química , Controle de Qualidade
19.
J Proteome Res ; 8(9): 4264-71, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19527021

RESUMO

Differences between individual phenotypes are due both to differences in genotype and to exposure to different environmental factors. A fundamental contribution to the definition of the individual phenotype for clinical and therapeutic applications would come from a deeper understanding of the metabolic phenotype. The existence of unique individual metabolic phenotypes has been hypothesized, but the experimental evidence has been only recently collected. Analysis of individual phenotypes over the timescale of years shows that the metabolic phenotypes are largely invariant. The present work also supports the idea that the individual metabolic phenotype can also be considered a metagenomic entity that is strongly affected by both gut microbiome and host metabolic phenotype, the latter defined by both genetic and environmental contributions.


Assuntos
Metabolômica/métodos , Fenótipo , Proteínas/metabolismo , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Ressonância Magnética Nuclear Biomolecular/métodos , Mapeamento de Peptídeos/métodos , Análise de Componente Principal , Proteínas/análise , Proteinúria/urina , Fatores de Tempo
20.
Mol Biosyst ; 5(2): 180-90, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19156264

RESUMO

The first application of high field NMR spectroscopy (800 MHz for (1)H observation) to human hepatic bile (as opposed to gall bladder bile) is reported. The bile sample used for detailed investigation was from a donor liver with mild fat infiltration, collected during organ retrieval prior to transplantation. In addition, to focus on the detection of bile acids in particular, a bile extract was analysed by 800 MHz (1)H NMR spectroscopy, HPLC-NMR/MS and UPLC-MS. In the whole bile sample, 40 compounds have been assigned with the aid of two-dimensional (1)H-(1)H TOCSY and (1)H-(13)C HSQC spectra. These include phosphatidylcholine, 14 amino acids, 10 organic acids, 4 carbohydrates and polyols (glucose, glucuronate, glycerol and myo-inositol), choline, phosphocholine, betaine, trimethylamine-N-oxide and other small molecules. An initial NMR-based assessment of the concentration range of some key metabolites has been made. Some observed chemical shifts differ from expected database values, probably due to a difference in bulk diamagnetic susceptibility. The NMR spectra of the whole extract gave identification of the major bile acids (cholic, deoxycholic and chenodeoxycholic), but the glycine and taurine conjugates of a given bile acid could not be distinguished. However, this was achieved by HPLC-NMR/MS, which enabled the separation and identification of ten conjugated bile acids with relative abundances varying from approximately 0.1% (taurolithocholic acid) to 34.0% (glycocholic acid), of which, only the five most abundant acids could be detected by NMR, including the isomers glycodeoxycholic acid and glycochenodeoxycholic acid, which are difficult to distinguish by conventional LC-MS analysis. In a separate experiment, the use of UPLC-MS allowed the detection and identification of 13 bile acids. This work has shown the complementary potential of NMR spectroscopy, MS and hyphenated NMR/MS for elucidating the complex metabolic profile of human hepatic bile. This will be useful baseline information in ongoing studies of liver excretory function and organ transplantation.


Assuntos
Ácidos e Sais Biliares/química , Bile/metabolismo , Cromatografia Líquida de Alta Pressão/métodos , Fígado/metabolismo , Espectroscopia de Ressonância Magnética/métodos , Espectrometria de Massas/métodos , Cromatografia Líquida/métodos , Fígado Gorduroso/metabolismo , Humanos , Metaboloma , Modelos Químicos , Reprodutibilidade dos Testes
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