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13C-formylation for improved nuclear magnetic resonance profiling of amino metabolites in biofluids.

ABSTRACT: An increased interest in metabolite profiling is driving the need for improved analytical techniques with greater performance for a variety of important applications. Despite their limited sensitivity, nuclear magnetic resonance (NMR) methods are attractive because of their simplicity, reproducibility, quantitative nature, and wide applicability. The use of chemoselective isotopic tags has the potential to advance the application of NMR for analyzing metabolites in complex biofluids by allowing detection of metabolites down to the low micromoalr level with high resolution and specificity. Here, we report a new (13)C-tagging method using (13)C-formic acid that delivers high sensitivity, good quantitation, and excellent resolution for (1)H-(13)C 2D NMR profiling of amino metabolites. High reproducibility (coefficient of variation (CV) = 2%) was observed for metabolites in urine with concentrations down to 10 microM. As amino compounds comprise an important class of metabolites and small molecules of biological roles, this new method therefore should be amenable to a variety of applications.

SUBMITTER: Ye T

PROVIDER: S-EPMC2837761 | biostudies-literature | 2010 Mar

REPOSITORIES: biostudies-literature

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13C-formylation for improved nuclear magnetic resonance profiling of amino metabolites in biofluids.

Ye Tao T Zhang Shucha S Mo Huaping H Tayyari Fariba F Gowda G A Nagana GA Raftery Daniel D

Analytical chemistry 20100301 6

An increased interest in metabolite profiling is driving the need for improved analytical techniques with greater performance for a variety of important applications. Despite their limited sensitivity, nuclear magnetic resonance (NMR) methods are attractive because of their simplicity, reproducibility, quantitative nature, and wide applicability. The use of chemoselective isotopic tags has the potential to advance the application of NMR for analyzing metabolites in complex biofluids by allowing ...[more]

PMID: 20180538

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Project description:Bifidobacteria are widely used as probiotics in several commercial products; however, to date there is little knowledge about their carbohydrate metabolic pathways. In this work, we studied the metabolism of glucose and lactose in the widely used probiotic strain Bifidobacterium animalis subsp. lactis BB-12 by in vivo (13)C nuclear magnetic resonance (NMR) spectroscopy. The metabolism of [1-(13)C]glucose was characterized in cells grown in glucose as the sole carbon source. Moreover, the metabolism of lactose specifically labeled with (13)C on carbon 1 of the glucose or the galactose moiety was determined in suspensions of cells grown in lactose. These experiments allowed the quantification of some intermediate and end products of the metabolic pathways, as well as determination of the consumption rate of carbon sources. Additionally, the labeling patterns in metabolites derived from the metabolism of glucose specifically labeled with (13)C on carbon 1, 2, or 3 in cells grown in glucose or lactose specifically labeled in carbon 1 of the glucose moiety ([1-(13)Cglucose]lactose), lactose specifically labeled in carbon 1 of the galactose moiety ([1-(13)Cgalactose]lactose), and [1-(13)C]glucose in lactose-grown cells were determined in cell extracts by (13)C NMR. The NMR analysis showed that the recovery of carbon was fully compatible with the fructose 6-phosphate, or bifid, shunt. The activity of lactate dehydrogenase, acetate kinase, fructose 6-phosphate phosphoketolase, and pyruvate formate lyase differed significantly between glucose and lactose cultures. The transcriptional analysis of several putative glucose and lactose transporters showed a significant induction of Balat_0475 in the presence of lactose, suggesting a role for this protein as a lactose permease. This report provides the first in vivo experimental evidence of the metabolic flux distribution in the catabolic pathway of glucose and lactose in bifidobacteria and shows that the bifid shunt is the only pathway involved in energy recruitment from these two sugars. On the basis of our experimental results, a model of sugar metabolism in B. animalis subsp. lactis is proposed.

Project description:BackgroundOur aim was to analyze and compare the characteristics and differences of blood metabolites between lymphangioleiomyomatosis (LAM) patients and healthy controls, in order to find biomarkers that can be used for the diagnosis and classification of LAM.MethodsBetween January 2020 to January 2022, 61 eligible LAM patients [51 sporadic LAM (S-LAM) and 10 tuberous sclerosis complex LAM (TSC-LAM)] from the First Affiliated Hospital of Guangzhou Medical University and 30 healthy controls were enrolled. Blood samples were taken for nuclear magnetic resonance (NMR) detection. Data analysis was performed by the umbrella program, and Wilcoxon analysis was used for comparisons between groups. The difference indicators were modeled by logistic regression. Diagnostic accuracy of the best predictive parameters was evaluated by the area under the receiver operating characteristic (ROC) curve (AUC), and the sensitivity and specificity were calculated.ResultsThe indexes differed between LAM patients and healthy controls, S-LAM patients and healthy controls, and between TSC-LAM patients and healthy controls. There were two different metabolic indexes between S-LAM and TSC-LAM patients. After logistic regression modeling and ROC analysis, methionine (AUC =0.929, sensitivity =73.8%, specificity =100%, cut-off value =0.011 mmol/L) and acetic acid (AUC =0.966, sensitivity =95.1%, specificity =90%, cut-off value =0.006 mmol/L) had the highest diagnostic efficiency in LAM patients, and could be used to distinguish between affected and healthy people. Methionine was significantly associated with pneumothorax (P<0.05), and creatinine was significantly correlated with hysteromyoma (P<0.05).ConclusionsMethionine and acetic acid in the plasma of LAM patients are potential biomarkers. Methionine was also associated with pneumothorax in LAM patients. Also, acetone and creatinine were promising metabolic markers to distinguish S-LAM from TSC-LAM. NMR as a new non-invasive diagnostic method had a good discriminatory power for LAM.

Dataset Information

13C-formylation for improved nuclear magnetic resonance profiling of amino metabolites in biofluids.

Publications

13C-formylation for improved nuclear magnetic resonance profiling of amino metabolites in biofluids.

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