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Tomoyoshi Soga

Publications and source records attributed to Tomoyoshi Soga.

18 recordsLinked to original sources
PubMed2025

2,3-Pyridinedicarboxylate Is Associated with Shorter Recurrence-Free Survival in Patients with Hypopharyngeal Squamous Cell Carcinoma.

UNLABELLED: Introduction: Metabolites are associated with the biology of cancer; however, no metabolites related to prognosis have been identified in head and neck cancer. This study aimed to identify metabolites associated with prognosis in patients with hypopharyngeal squamous cell carcinoma (HPSCC). METHODS: Fifty-two patients who underwent surgery for HPSCC were included and randomly divided into test and validation cohorts of 26 patients each for further metabolome analysis using capillary electrophoresis/mass spectrometry on tumor and non-tumor tissues of the hypopharynx. Twenty-two patients who received adjuvant therapy after surgery were included. The receiver operating characteristic (ROC) and univariate and multivariate analyses were used to explore the relationship between recurrence-free survival (RFS), clinicopathological factors, and differentiated metabolites. RESULTS: ROC analysis revealed six metabolites significantly associated with RFS in both cohorts, and multivariate analysis indicated that 2,3-pyridinedicarboxylate was a significantly independent poorer prognostic factor in the cohorts including patients with HPSCC without any adjuvant therapies (p = 0.017). CONCLUSION: 2,3-Pyridinedicarboxylate, involved in NAD+ metabolism and genomic stability, suggests the possibility of developing molecular-targeted drugs for the production of metabolites related to prognosis. This study identifies novel prognostic metabolites and their associated metabolic pathways in HPSCC, highlighting potential therapeutic targets for treatment. .

Humans↗
PubMed2007

Capillary electrophoresis-mass spectrometry for metabolomics.

A new approach for the comprehensive and quantitative analysis of charged metabolites by capillary electrophoresis-mass spectrometry (CE-MS) is described. Metabolites are first separated by CE based on charge and size, and then selectively detected using MS by monitoring ions over a large range of m/z values. This technique enables the simultaneous determination of over 1000 charged species, and it can readily be applied to various types of biological samples originating from bacteria, plants, mammals, body fluids, and others. This chapter highlights detailed practical procedures for using this technology.

Animals↗
PubMed2006

MathDAMP: a package for differential analysis of metabolite profiles.

BACKGROUND: With the advent of metabolomics as a powerful tool for both functional and biomarker discovery, the identification of specific differences between complex metabolite profiles is becoming a major challenge in the data analysis pipeline. The task remains difficult, given the datasets' size, complexity, and common shifts in migration (elution/retention) times between samples analyzed by hyphenated mass spectrometry methods. RESULTS: We present a Mathematica (Wolfram Research, Inc.) package MathDAMP (Mathematica package for Differential Analysis of Metabolite Profiles), which highlights differences between raw datasets acquired by hyphenated mass spectrometry methods by applying arithmetic operations to all corresponding signal intensities on a datapoint-by-datapoint basis. Peak identification and integration is thus bypassed and the results are displayed graphically. To facilitate direct comparisons, the raw datasets are automatically preprocessed and normalized in terms of both migration times and signal intensities. A combination of dynamic programming and global optimization is used for the alignment of the datasets along the migration time dimension. The processed datasets and the results of direct comparisons between them are visualized using density plots (axes represent migration time and m/z values while peaks appear as color-coded spots) providing an intuitive overall view. Various forms of comparisons and statistical tests can be applied to highlight subtle differences. Overlaid electropherograms (chromatograms) corresponding to the vicinities of the candidate differences from any result may be generated in a descending order of significance for visual confirmation. Additionally, a standard library table (a list of m/z values and migration times for known compounds) may be aligned and overlaid on the plots to allow easier identification of metabolites. CONCLUSION: Our tool facilitates the visualization and identification of differences between complex metabolite profiles according to various criteria in an automated fashion and is useful for data-driven discovery of biomarkers and functional genomics.

Biomarkers↗
PubMed2006

HybGFS: a hybrid method for genome-fingerprint scanning.

BACKGROUND: Protein identification based on mass spectrometry (MS) has previously been performed using peptide mass fingerprinting (PMF) or tandem MS (MS/MS) database searching. However, these methods cannot identify proteins that are not already listed in existing databases. Moreover, the alternative approach of de novo sequencing requires costly equipment and the interpretation of complex MS/MS spectra. Thus, there is a need for novel high-throughput protein-identification methods that are independent of existing predefined protein databases. RESULTS: Here, we present a hybrid method for genome-fingerprint scanning, known as HybGFS. This technique combines genome sequence-based peptide MS/MS ion searching with liquid-chromatography elution-time (LC-ET) prediction, to improve the reliability of identification. The hybrid method allows the simultaneous identification and mapping of proteins without a priori information about their coding sequences. The current study used standard LC-MS/MS data to query an in silico-generated six-reading-frame translation and the enzymatic digest of an entire genome. Used in conjunction with precursor/product ion-mass searching, the LC-ETs increased confidence in the peptide-identification process and reduced the number of false-positive matches. The power of this method was demonstrated using recombinant proteins from the Escherichia coli K12 strain. CONCLUSION: The novel hybrid method described in this study will be useful for the large-scale experimental confirmation of genome coding sequences, without the need for transcriptome-level expression analysis or costly MS database searching.

Algorithms↗
PubMed2006

Evaluation of metabolic alteration in transgenic rice overexpressing dihydroflavonol-4-reductase.

BACKGROUND AND AIMS: Previous studies have shown that transgenic rice plants overexpressing YK1, which possesses dihydroflavonol-4-reductase (DFR) activity, showed biotic and abiotic stress tolerance. High throughput profiles of metabolites have also been shown in such transgenic plants by Fourier transform ion cyclotron mass spectrometry. In this study, capillary electrophoresis mass spectrometry analysis (CE/MS) was employed to identify precise metabolites such as organic acids, amino acids and sugars. METHODS: Using CE/MS, we analysed several metabolites of glycolysis, the tricarboxylic acid (TCA) cycle and the pentose phosphate pathway. In addition, the concentrations of sugars and ion were quantified. KEY RESULTS: In YK1 (DFR)-overexpressing plants, the concentrations of cis-aconitate, isocitrate and 2-oxoglutarate were higher in leaves, whereas those of fructose-1,6-bisphosphate and glyceraldehyde-3-phosphate were lower in roots. In seeds, the amounts of free amino acids and metals were altered, whereas sugars in seeds were kept constant. In YK1 calli, an approx. 3-fold increase in glutathione was observed, whereas the activities of glutathione peroxidase and glutathione reductase were concomitantly increased. CONCLUSIONS: The overexpression of YK1 (DFR) was associated with slight changes in the amounts of several metabolites analysed in whole plants, whilst glutathione derivatives were substantially increased in suspension-cultured cells.

Alcohol Oxidoreductases↗
PubMed2006

Differential metabolomics reveals ophthalmic acid as an oxidative stress biomarker indicating hepatic glutathione consumption.

Metabolomics is an emerging tool that can be used to gain insights into cellular and physiological responses. Here we present a metabolome differential display method based on capillary electrophoresis time-of-flight mass spectrometry to profile liver metabolites following acetaminophen-induced hepatotoxicity. We globally detected 1,859 peaks in mouse liver extracts and highlighted multiple changes in metabolite levels, including an activation of the ophthalmate biosynthesis pathway. We confirmed that ophthalmate was synthesized from 2-aminobutyrate through consecutive reactions with gamma-glutamylcysteine and glutathione synthetase. Changes in ophthalmate level in mouse serum and liver extracts were closely correlated and ophthalmate levels increased significantly in conjunction with glutathione consumption. Overall, our results provide a broad picture of hepatic metabolite changes following acetaminophen treatment. In addition, we specifically found that serum ophthalmate is a sensitive indicator of hepatic GSH depletion, and may be a new biomarker for oxidative stress. Our method can thus pinpoint specific metabolite changes and provide insights into the perturbation of metabolic pathways on a large scale and serve as a powerful new tool for discovering low molecular weight biomarkers.

Animals↗
PubMed2006

Metabolomics approach for enzyme discovery.

The search for novel enzymes is an important but difficult task in functional genomics. Here, we present a systematic method based on in vitro assays in combination with metabolite profiling to discover novel enzymatic activities. A complex mixture of metabolites is incubated with purified candidate proteins and the reaction mixture is subsequently profiled by capillary electrophoresis electrospray ionization mass spectrometry (CE-MS). Specific changes in the metabolite composition can directly suggest the presence of an enzymatic activity while subsequent identification of the compounds whose level changed specifically can pinpoint the actual substrate(s) and product(s) of the reaction. We first evaluated the method using several Escherichia coli metabolic enzymes and then applied it to the functional screening of uncharacterized proteins. In this manner, YbhA and YbiV proteins were found to display both phosphotransferase and phosphatase activity toward different sugars/sugar phosphates. Our approach should be broadly applicable and useful for enzyme discovery in any system.

Electrophoresis, Capillary↗
PubMed2006

Prediction of liquid chromatographic retention times of peptides generated by protease digestion of the Escherichia coli proteome using artificial neural networks.

We developed a computational method to predict the retention times of peptides in HPLC using artificial neural networks (ANN). We performed stepwise multiple linear regressions and selected for ANN input amino acids that significantly affected the LC retention time. Unlike conventional linear models, the trained ANN accurately predicted the retention time of peptides containing up to 50 amino acid residues. In 834 peptides, there was a strong correlation (R2 = 0.928) between measured and predicted retention times. We demonstrated the utility of our method by the prediction of the retention time of 121,273 peptides resulting from LysC-digestion of the Escherichia coli proteome. Our approach is useful for the proteome-wide characterization of peptides and the identification of unknown peptide peaks obtained in proteome analysis.

Chromatography, High Pressure Liquid↗
PubMed2005

Large-scale prediction of cationic metabolite identity and migration time in capillary electrophoresis mass spectrometry using artificial neural networks.

We developed a computational technique to assist in the large-scale identification of charged metabolites. The electrophoretic mobility of metabolites in capillary electrophoresis-mass spectrometry (CE-MS) was predicted from their structure, using an ensemble of artificial neural networks (ANNs). Comparison between relative migration times of 241 various cations measured by CE-MS and predicted by a trained ANN ensemble produced a correlation coefficient of 0.931. When we used our technique to characterize all metabolites listed in the KEGG ligand database, the correct compounds among the top three candidates were predicted in 78.0% of cases. We suggest that this approach can be used for the prediction of the migration time of any cation and that it represents a powerful method for the identification of uncharacterized CE-MS peaks in metabolome analysis.

Cations↗
PubMed2005

Conductivity detection in capillary zone electrophoresis: inspection by PeakMaster.

A simple rule stating that the signal in conductivity detection in capillary zone electrophoresis is proportional to the difference between the analyte mobility and mobility of the background electrolyte (BGE) co-ion is valid only for systems with fully ionized electrolytes. In zone electrophoresis systems with weak electrolytes both conductivity signal and electromigration dispersion of analyte peaks depend on the conductivity and pH effects. This allows optimization of the composition of BGEs to give a good conductivity signal of analytes while still keeping electromigration dispersion near zero, regardless of the injected amount of sample. The demands to achieve minimum electromigration dispersion and high sensitivity in conductivity detection can be accomplished at the same time. PeakMaster software is used for inspection of BGEs commonly used for separation of sugars (carbohydrates, saccharides) at highly alkaline pH. It is shown that the terms direct and indirect conductivity detection are misleading and should not be used.

Electric Conductivity↗
PubMed2004

Qualitative and quantitative analysis of amino acids by capillary electrophoresis-electrospray ionization-tandem mass spectrometry.

We describe a method to identify and quantify amino acids using capillary electrophoresis-electrospray ionization-triple-quadrupole tandem mass spectrometry (CE-ESI-MS/MS). Amino acids, including physiological amino acids, were first separated by CE under acidic pH conditions and then detected by MS/MS. To efficiently introduce the whole sample into the capillary, no electrical potential was applied to the electrospray probe until running electrophoresis. The position of the electrosprayer with respect to the MS capillary entrance drastically affected sensitivity and generation of cluster ions. MS/MS with multiple reaction monitoring (MRM) detection was performed to obtain sufficient selectivity and sensitivity. Under optimized CE-MS/MS conditions, the minimum detectable levels for 32 free amino acids normally found in proteins and other physiological amino acids were between 0.1 and 14 micromol/L with pressure injection of 50 mbar for 3 s (3 nL) at a signal-to-noise ratio of 3. For most amino acids, this constitutes a severalfold increase in sensitivity compared to CE-MS. The relative standard deviations (% RSD) for all amino acids were better than 0.4% for migration times and between 1.4% and 8.6% for peak areas (n = 10). Since amino acids exhibited characteristic MS/MS spectra, this approach is useful for the simultaneous, selective, quantitative, and reproducible analysis of amino acids in physiological and biological samples that contain various kinds of matrices. The power of the method was demonstrated by analyzing amino acids in human urine.

Amino Acids↗
PubMed2004

Application of capillary electrophoresis-mass spectrometry to synthetic in vitro glycolysis studies.

We propose an approach designed to reconstitute a metabolic pathway composed of multistep biochemical reactions, rather than to dissect the individual reactions that make up the pathway. A synthetic in vitro glycolysis was reconstructed from ten purified Escherichia coli (E. coli) enzymes to obtain a better understanding of the regulation of sequential enzymatic reactions. The key to the success of this approach is the ability to perform direct and simultaneous determination of the diverse metabolic intermediates in the pathway by capillary electrophoresis-mass spectrometry. We observed that the pathway is regulated by a delicate balance between the changing metabolite concentrations and behaves like a natural biological oscillating network that has hitherto not been reported for E. coli glycolysis. The end-product, pyruvate, was periodically synthesized from glucose at an overall efficiency of 30%, corresponding to an average of 90% conversion efficiency for each of the ten steps involved. This approach is likely useful for the synthesis of natural products requiring complex sequential biocatalytic reactions.

Catalysis↗
PubMed2004

Simultaneous determination of the main metabolites in rice leaves using capillary electrophoresis mass spectrometry and capillary electrophoresis diode array detection.

The study of the metabolomics of primary metabolites using conventional chemical analyses requires a high-throughput method. Chemical derivatizations are a prerequisite for gas-chromatographic separation, and a large sample quantity is needed for liquid-chromatographic separation and nuclear magnetic resonance detection systems. Recently, we have developed a capillary electrophoresis-mass spectrometry (CE-MS) technology that can simultaneously quantify a large number of primary metabolites, using only a small quantity of samples, and without any chemical derivatizations. Parallel use of a capillary electrophoresis-diode array detector (CE-DAD) system further enables almost all water-soluble intracellular metabolites to be analyzed. We demonstrate, with rice leaves, a simple and rapid method of sample preparation for CE analysis; using this method, we have successfully measured the levels of 88 main metabolites involved in glycolysis, the tricarboxylic acid cycle, the pentose phosphate pathway, photorespiration, and amino acid biosynthesis.

Electrophoresis, Capillary↗
PubMed2002

Pressure-assisted capillary electrophoresis electrospray ionization mass spectrometry for analysis of multivalent anions.

We describe a method, based on pressure-assisted capillary electrophoresis coupled to electrospray ionization mass spectrometry (PACE/ESI-MS), that allows the simultaneous and quantitative analysis of multivalent anions, such as citrate isomers, nucleotides, nicotinamide-adenine dinucleotides, and flavin adenine dinucleotide, and coenzyme A (CoA) compounds. Key to the analysis was using a noncharged polymer, poly(dimethylsiloxane), coated to the inner surface of the capillary to prevent anionic species from adsorbing onto the capillary wall. It was also necessary to drive a constant liquid flow toward the MS by applying air pressure to the inlet capillary during electrophoresis to maintain a conductive liquid junction between the capillary and the electrospray needle. Although theoretical plates were inferior to those obtained by CE/ESI-MS using a cationic polymer-coated capillary, the PACE/ESI-MS method improved reproducibility and sensitivity of these anions. Eighteen anions were separated by PACE and selectively detected by a quadrupole mass spectrometer with a sheath-flow electrospray ionization interface. The relative standard deviations (n = 6) of the method were better than 0.6% for migration times and between 1.4% and 6.2% for peak areas. The detection limits for these species were between 0.4 and 3.7 micromol/L with pressure injection of 50 mbar for 30 s (30 nL), that is, mass detection limits calculated in the range from 12 to 110 fmol at a signal-to-noise ratio of 3. The utility of the method was demonstrated by analysis of citrate isomers, nucleotides, dinucleotides, and CoA compounds extracted from Bacillus subtilis cells.

Anions↗
PubMed2002

Simultaneous determination of anionic intermediates for Bacillus subtilis metabolic pathways by capillary electrophoresis electrospray ionization mass spectrometry.

A method for simultaneous determination of anionic metabolites based on capillary electrophoresis (CE) coupled to electrospray ionization mass spectrometry is described. To prevent current drop by the system, electroosmotic flow (EOF) reversal by using a cationic polymer-coated capillary was indispensable. A mixture containing 32 standards including carboxylic acids, phosphorylated carboxylic acids, phosphorylated saccharides, nucleotides, and nicotinamide and flavin adenine coenzymes of glycolysis and the tricarboxylic acid cycle pathways were separated by CE and selectively detected by a quadrupole mass spectrometer with a sheath-flow electrospray ionization interface. Key to the analysis was EOF reversal using a cationic polymer-coated capillary and an electrolyte system consisting of 50 mM ammonium acetate, pH 9.0. The relative standard deviations of the method were better than 0.4% for migration times and between 0.9% and 5.4% for peak areas. The concentration detection limits for these metabolites were between 0.3 and 6.7 micromol/L with pressure injection of 50 mbar for 30 s (30 nL); i.e., mass detection limits ranged from 9 to 200 fmol, at a signal-to-noise ratio of 3. This method was applied to the comprehensive analysis of metabolic intermediates extracted from Bacillus subtilis, and 27 anionic metabolites could be directly detected and quantified.

Anions↗
PubMedDate not supplied

Quantitative metabolome analysis using capillary electrophoresis mass spectrometry.

A new approach for the comprehensive and quantitative analysis of charged metabolites by capillary electrophoresis mass spectrometry (CE-MS) is proposed. Metabolites are first separated by CE based on charge and size and then selectively detected using MS by monitoring over a large range of m/z values. This method enabled the determination of 352 metabolic standards and its utility was demonstrated in the analysis of 1692 metabolites from Bacillus subtilis extracts, revealing significant changes in metabolites during B. subtilis sporulation.

Anions↗