Menopause is associated with increased abdominal fat and increased risk of developing diabetes and cardiovascular disease.
Objectives
The present study evaluated the plasma metabolic response in relation to insulin sensitivity after weight loss via diet intervention.
Methods
This work includes two studies; i) Ten women on a 5 weeks Paleolithic-type diet (PD, 30 energy percent (E%) protein, 40 E% fat, 30 E% carbohydrates), ii) 55 women on 6 months of either PD or Nordic Nutrition Recommendations diet (NNR, 15 E% protein, 30 E% fat, and 55 E% carbohydrates). Plasma metabolic profiles were acquired at baseline and post diet using gas chromatography time-of-flight/mass spectrometry and investigated in relation to insulin sensitivity using multivariate bioinformatics.
Results
Both the PD and NNR diet resulted in significant weight loss, reduced waist circumference, improved serum lipid profiles, and improved insulin sensitivity. We detected a baseline metabolic profile that correlated significantly with insulin sensitivity, and of which components increased significantly in the PD group compared to NNR. Specifically, a significant increase in myo-inositol (MI), a second messenger of insulin action, and β-hydroxybutyric acid (β-HB) increased while dihomo-gamma-linoleic acid (DGLA) decreased in PD compared to NNR, which correlated with improved insulin sensitivity. We also detected a significant decrease in tyrosine and tryptophan, potential markers of insulin resistance when elevated in the circulation, with the PD but not the NNR.
Conclusions
Using metabolomics, we detected changes in the plasma metabolite profiles associated with weight loss in postmenopausal women by different diets. The metabolic profiles following 6 months of PD were linked to beneficial effects on insulin sensitivity compared to NNR.
Boiling ethanol extraction is a frequently used method for metabolomics studies of biological samples. However, the stability of several central carbon metabolites, including nucleotide triphosphates, and the influence of the cellular matrix on their degradation have not been addressed.
Objectives
To study how a complex cellular matrix extracted from yeast (Saccharomyces cerevisiae) may affect the degradation profiles of nucleotide triphosphates extracted under boiling ethanol conditions.
Methods
We present a double-labelling LC–MS approach with a 13C-labeled yeast cellular extract as complex surrogate matrix, and 13C15N-labeled nucleotides as internal standards, to study the effect of the yeast matrix on the degradation of nucleotide triphosphates.
Results
While nucleotide triphosphates were degraded to the corresponding diphosphates in pure solutions, degradation was prevented in the presence of the yeast matrix under typical boiling ethanol extraction conditions.
Conclusions
Extraction of biological samples under boiling ethanol extraction conditions that rapidly inactivate enzyme activity are suitable for labile central energy metabolites such as nucleotide triphosphates due to the stabilizing effect of the yeast matrix. The basis of this phenomenon requires further study.
Stable isotopic labeling experiments are powerful tools to study metabolic pathways, to follow tracers and fluxes in biotic and abiotic transformations and to elucidate molecules involved in metal complexing.
Objective
To introduce a software tool for the identification of isotopologues from mass spectrometry data.
Methods
DeltaMS relies on XCMS peak detection and X13CMS isotopologue grouping and then analyses data for specific isotope ratios and the relative error of these ratios. It provides pipelines for recognition of isotope patterns in three experiment types commonly used in isotopic labeling studies: (1) search for isotope signatures with a specific mass shift and intensity ratio in one sample set, (2) analyze two sample sets for a specific mass shift and, optionally, the isotope ratio, whereby one sample set is isotope-labeled, and one is not, (3) analyze isotope-guided perturbation experiments with a setup described in X13CMS.
Results
To illustrate the versatility of DeltaMS, we analyze data sets from case-studies that commonly pose challenges in evaluation of natural isotopes or isotopic signatures in labeling experiment. In these examples, the untargeted detection of sulfur, bromine and artificial metal isotopic patterns is enabled by the automated search for specific isotopes or isotope signatures.
Conclusion
DeltaMS provides a platform for the identification of (pre-defined) isotopologues in MS data from single samples or comparative metabolomics data sets.
The chemical sensitivity of urine metabolomics analysis is greatly compromised due to the large amounts of inorganic salts in urine (NaCl, KCl), which are detrimental to analytical instrumentation, e.g. chromatographic columns or mass spectrometers. Traditional desalting approaches applied to urine pretreatment suffer from the chemical losses, which reduce the information depth of analysis.
Objectives
We aimed to test a simple approach for the simultaneous preconcentration and desalting of organic solutes in urine based on the collection of induced bursting bubble aerosols above the surface of urine samples.
Method
Bursting bubbles were generated at ambient conditions by feeding gas through an air diffuser at the bottom of diluted (200 times in ultrapure water) urine solution (50–500 mL). Collected aerosols were analyzed by the direct-infusion electrospray ionization mass spectrometry (ESI–MS).
Results
The simultaneous preconcentration (ca. 6–12 fold) and desalting (ca. six–tenfold) of organic solutes in urine was achieved by the bursting bubble sample pretreatment, which allowed ca. three-times higher number of identified urine metabolites by high-resolution MS analysis. No chemical losses due to bubbling were observed. The increased degree of MS data clustering was demonstrated on the principal component analysis of data sets from the urine of healthy people and from the urine people with renal insufficiency. At least ten times higher sensitivity of trace drug detection in urine was demonstrated for clenbuterol and salbutamol.
Conclusion
Our results indicate the high versatility of bubble bursting as a simple pretreatment approach to enhance the chemical depth and sensitivity of urine analysis. The approach could be attractive for personalized medicine as well as for the diagnostics of renal disorders of different etiology (diabetic nephropathy, chronic renal failure, transplant-associated complications, oncological disorders).
Graphical Abstract
Urine desalting and preconcentration in bursting bubbles.
High-fat and high-carbohydrate diets cause a number of metabolic disorders in mammals. However, little is known about metabolomic changes caused by dietary imbalances in fish.
Objectives
The objective of this study was to assess the impacts of high-fat diet (HFD), high-carbohydrate diet (HCD) and high-fat-high-carbohydrate diet (HFHCD) on metabolites in a farmed cyprinid fish Megalobrama amblycephala.
Methods
We have employed the 1H NMR-based metabolomic approach to measure the concentrations of metabolites in plasma and liver of four different diet groups: HFD, HCD, HFHCD and control. Multivariate statistical analyses were used to determine significantly changed metabolites between all group-pairs.
Results
All three test diets have affected metabolic profiles, phenotypes and clinical chemistry. High-fat diets (HFD, HFHCD) resulted in a higher average weight than HCD, but high-carbohydrate diets (HCD, HFHCD) caused signs of liver damage. HCD has resulted in elevated metabolites in energy pathways, leading to further disturbances in creatine pathway. Excess of carbohydrate and lipid metabolism products in the HFHCD group appears to have caused “congestion” of the TCA cycle, causing a significant decline in the numbers of amino acids entering the cycle, which in turn resulted in elevated levels of seven amino acids in this group. Gut microbiota metabolites (TMA) exhibited a strong positive correlation with the carbohydrate content and a negative correlation with the fat content in diets.
Conclusion
These results provide an important insight into the diet-affected metabolic disorders that often lead to financial losses in the aquaculture of Megalobrama amblycephala.
Chromatography with mass spectrometry (MS) is a technique of choice for metabolomic analysis of plant extracts. Single dimension gas chromatography (1DGC) with MS leads to poorly resolved metabolites of complex Eucalyptus spp. leaf oil secondary metabolites and consequently limited metabolic coverage of secondary compounds. Multidimensional chromatography with high resolution MS can contribute to advances in this field.
Objectives
Deeper insight into metabolite composition and variation for Eucalyptus spp. leaf oils through systematic untargeted metabolic profiling using comprehensive two-dimensional GC (GC?×?GC) with high resolution time-of-flight MS (accTOFMS), using generalised processes for metabolite identification.
Methods
GC?×?GC separation used cryogenic modulation, with standard length polar first dimension and short fast analysis non-polar 2D columns. Compound tentative identification incorporated 1D and 2D retention information, retention indices, mass spectrum matching, and accurate mass MS data. Global metabolic profiles were interpreted through 2D contour plots and chemometric analysis.
Results
Strategies for metabolite screening and identification using GC?×?GC-accTOFMS were proposed. Considerably more components are detected and recognised than for 1DGC. Structured 2D molecular composition chromatographic patterns aid identification. ca. 400 metabolites were detected, 183 compounds were identified or tentatively identified, representing between 50.8–90.0% of the total ion count, comprising various chemical families. PCA revealed discriminating metabolites, allowing chemotaxonomic classification of species.
Conclusion
Expansion of metabolic coverage by using GC?×?GC-accTOFMS, and detailed 2D metabolic fingerprints of E. polybractea, E. citriodora, E. radiata and E. globulus leaf oils were established. This high resolution analytical platform, and identification strategy can be adapted to metabolic analysis of other plant extracts.
Graphical abstract
Phytoconstituents of four Australian eucalypt leaf oils were profiled using high resolution GC?×?GC-accurate mass TOFMS. Two-dimensional plots illustrated significant expansion of metabolic coverage. PCA discriminated metabolites of the eucalypts.
Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.
Objectives
In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.
Methods
The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.
Results
A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.
Conclusion
The workflow generated repeatable and informative fingerprints for robust metabolome characterization.
To express a Δ6-desaturase gene and produce gamma-linolenic acid (GLA) and stearidonic acid (SDA) in prokaryotic expression system (Escherichia coli), and analyze its substrate specificity in the omega-3 fatty acid biosynthetic pathway.
Results
Full-length ORF (1448 bp) of Δ6Des-Iso was isolated from Isochrysis sp. and characterized using multiple sequence alignment, phylogenetic analysis, transmembrane domain, and protein tertiary structure. Δ6Des-Iso is a front-end desaturase consisting of three conserved histidine domains and a cytochrome b5 domain. Δ6Des-Iso was cloned and expressed in E. coli with the production of GLA and SDA. Recombinant E. coli utilized 27 and 8% of exogenously supplied alpha-linolenic acid (ALA) and linoleic acid (LA) to produce 6.3% of SDA and 2.3% of GLA, respectively, suggesting that isolated Δ6Des-Iso is specific to the omega-3 pathway.
Conclusion
For the first time production of GLA and SDA in a prokaryotic system was achieved.
Data sharing is being increasingly required by journals and has been heralded as a solution to the ‘replication crisis’.
Objectives
(i) Review data sharing policies of journals publishing the most metabolomics papers associated with open data and (ii) compare these journals’ policies to those that publish the most metabolomics papers.
Methods
A PubMed search was used to identify metabolomics papers. Metabolomics data repositories were manually searched for linked publications.
Results
Journals that support data sharing are not necessarily those with the most papers associated to open metabolomics data.
Conclusion
Further efforts are required to improve data sharing in metabolomics.
Patients with chronic fatigue syndrome and those with orthostatic intolerance share many symptoms, yet questions exist as to whether CFS patients have physiological evidence of orthostatic intolerance.
Objective
To determine if some CFS patients have increased rates of orthostatic hypotension, hypertension, tachycardia, or hypocapnia relative to age-matched controls.
Design
Assess blood pressure, heart rate, respiratory rate, end tidal CO2 and visual analog scales for orthostatic symptoms when supine and when standing for 8 minutes without moving legs.
Setting
Referral practice and research center.
Participants
60 women and 15 men with CFS and 36 women and 4 men serving as age matched controls with analyses confined to 62 patients and 35 controls showing either normal orthostatic testing or a physiological abnormal test.
CFS patients had higher rates of abnormal tests than controls (53% vs 20%, p < .002), but rates of orthostatic tachycardia, orthostatic hypotension, and orthostatic hypertension did not differ significantly between patients and controls (11.3% vs 5.7%, 6.5% vs 2.9%, 19.4% vs 11.4%, respectively). In contrast, rates of orthostatic hypocapnia were significantly higher in CFS than in controls (20.6% vs 2.9%, p < .02). This CFS group reported significantly more feelings of illness and shortness of breath than either controls or CFS patients with normal physiological tests.
Conclusion
A substantial number of CFS patients have orthostatic intolerance in the form of orthostatic hypocapnia. This allows subgrouping of patients with CFS and thus reduces patient pool heterogeneity engendered by use of a clinical case definition.
Adoption of automatic profiling tools for 1H-NMR-based metabolomic studies still lags behind other approaches in the absence of the flexibility and interactivity necessary to adapt to the properties of study data sets of complex matrices.
Objectives
To provide an open source tool that fully integrates these needs and enables the reproducibility of the profiling process.
Methods
rDolphin incorporates novel techniques to optimize exploratory analysis, metabolite identification, and validation of profiling output quality.
Results
The information and quality achieved in two public datasets of complex matrices are maximized.
Conclusion
rDolphin is an open-source R package (http://github.com/danielcanueto/rDolphin) able to provide the best balance between accuracy, reproducibility and ease of use.
Since blood is in contact with all tissues in the body and is considered to dynamically reflect the body’s pathophysiological status, serum metabolomics changes are important and have diagnostic value in early cancer detection.
Objectives
In this prospective study, we investigated the application of metabolomics to differentiate subjects with incident breast cancer (BC) from subjects who remained free of cancer during a mean follow-up period of 7 years with the aim of identifying valuable biomarkers for BC.
Methods
Baseline serum samples from 84 female subjects with incident BC (BC group) and 88 cancer-free female subjects (control group) were used. Metabolic alterations associated with BC were investigated via metabolomics analysis of the baseline serum samples using ultra-performance liquid chromatography-linear-trap quadrupole-Orbitrap mass spectrometry.
Results
A total of 57 metabolites were identified through the metabolic analysis. Among them, 20 metabolite levels were significantly higher and 22 metabolite levels were significantly lower in the BC group than in the control group at baseline. Ten metabolic pathways, including amino acid metabolism, arachidonic acid (AA) metabolism, fatty acid metabolism, linoleic acid metabolism, and retinol metabolism, showed significant differences between the BC group and the control group. Logistic regression revealed that the incidence of BC was affected by leucine, AA, prostaglandin (PG)J2, PGE2, and γ-linolenic acid (GLA).
Conclusions
This prospective study showed the clinical relevance of dysregulation of various metabolisms on the incidence of BC. Additionally, leucine, AA, PGJ2, PGE2, and GLA were identified as independent variables affecting the incidence of BC.
Recurrent pregnancy loss (RPL) is a heterogeneous condition and thrombophilias have been considered as a probable cause.
OBJECTIVE
The aim of this study was to investigate the prevalence of the coagulation factor XIII Val34Leu polymorphism among women with unexplained RPL.
METHODS
A total of 140 women with a history of unexplained RPL and 100 age-matched healthy fertile women were recruited. The presence of FXIII Val34Leu polymorphism among the cases and controls was investigated using PCR-RFLP method.
RESULTS
Genotype analyses of the subjects revealed that the patients had a significantly higher prevalence of V/L and L/L than the controls (P<0.05): 33.5% vs. 15%, and 9.2% vs. 2%, respectively.
CONCLUSION
These results indicate a significant association between FXIII Val34Leu polymorphism and unexplained RPL in the Iranian patient.
Allograft rejection is still an important complication after kidney transplantation. Currently, monitoring of these patients mostly relies on the measurement of serum creatinine and clinical evaluation. The gold standard for diagnosing allograft rejection, i.e. performing a renal biopsy is invasive and expensive. So far no adequate biomarkers are available for routine use.
Objectives
We aimed to develop a urine metabolite constellation that is characteristic for acute renal allograft rejection.
Methods
NMR-Spectroscopy was applied to a training cohort of transplant recipients with and without acute rejection.
Results
We obtained a metabolite constellation of four metabolites that shows promising performance to detect renal allograft rejection in the cohorts used (AUC of 0.72 and 0.74, respectively).
Conclusion
A metabolite constellation was defined with the potential for further development of an in-vitro diagnostic test that can support physicians in their clinical assessment of a kidney transplant patient.
Multimodal nanomaterials are useful for providing enhanced diagnostic information simultaneously for a variety of in vivo imaging methods. According to our research findings, these multimodal nanomaterials offer promising applications for cancer therapy.
Results
Melanin nanoparticles can be used as a platform imaging material and they can be simply produced by complexation with various imaging active ions. They are capable of specifically targeting epidermal growth factor receptor (EGFR)-expressing cancer cells by being anchored with a specific antibody. Ion-doped melanin nanoparticles were found to have high bioavailability with long-term stability in solution, without any cytotoxicity in both in vitro and in vivo systems.
Conclusion
By combining different imaging modalities with melanin particles, we can use the complexes to obtain faster diagnoses by computed tomography deep-body imaging and greater detailed pathological diagnostic information by magnetic resonance imaging. The ion-doped melanin nanoparticles also have applications for radio-diagnostic treatment and radio imaging-guided surgery, warranting further proof of concept experimental.
Quantification of tetrahydrofolates (THFs), important metabolites in the Wood–Ljungdahl pathway (WLP) of acetogens, is challenging given their sensitivity to oxygen.
Objective
To develop a simple anaerobic protocol to enable reliable THFs quantification from bioreactors.
Methods
Anaerobic cultures were mixed with anaerobic acetonitrile for extraction. Targeted LC–MS/MS was used for quantification.
Results
Tetrahydrofolates can only be quantified if sampled anaerobically. THF levels showed a strong correlation to acetyl-CoA, the end product of the WLP.
Conclusion
Our method is useful for relative quantification of THFs across different growth conditions. Absolute quantification of THFs requires the use of labelled standards.
We aimed to identify new high-yield dextranase strains and study the catalytic potential of dextranase from the strain in industrial applications.
Results
Dextranase-producing strains were screened from soil samples, and a potential strain was identified as Chaetomium globosum according to its phenotype, biochemical characteristics, and rDNA analysis. Crude dextranase was purified to reach 10.97-fold specific activity and 18.7% recovery. The molecular weight of the enzyme was 53 kDa with an optimum temperature and pH of 60 °C and 5.5, respectively. Enzyme activity was stable at pH 4.0–7.0 and displayed sufficient thermal stability at temperatures?<?50 °C. Mn2+ (10 mM) enhanced dextranase activity by 134.44%. The enzyme was identified as an endodextranase. It displayed very high hydrolytic affinity toward high-molecular weight dextran T2000, reaching 97.9% hydrolysis within 15 min at 2 U/mL.
Conclusion
Collectively, these results suggest that Chaetomium globosum shows higher production and specificity of dextranase than that from other reported strains. These findings may offer new insights into the potential of dextranase in the sugar, medical, and food industries.
Aqueous–methanol mixtures have successfully been applied to extract a broad range of metabolites from plant tissue. However, a certain amount of material remains insoluble.
Objectives
To enlarge the metabolic compendium, two ionic liquids were selected to extract the methanol insoluble part of trunk from Betula pendula.
Methods
The extracted compounds were analyzed by LC/MS and GC/MS.
Results
The results show that 1-butyl-3-methylimidazolium acetate (IL-Ac) predominantly resulted in fatty acids, whereas 1-ethyl-3-methylimidazolium tosylate (IL-Tos) mostly yielded phenolic structures. Interestingly, bark yielded more ionic liquid soluble metabolites compared to interior wood.
Conclusion
From this one can conclude that the application of ionic liquids may expand the metabolic snapshot.
Concerning NMR-based metabolomics, 1D spectra processing often requires an expert eye for disentangling the intertwined peaks.
Objectives
The objective of NMRProcFlow is to assist the expert in this task in the best way without requirement of programming skills.
Methods
NMRProcFlow was developed to be a graphical and interactive 1D NMR (1H & 13C) spectra processing tool.
Results
NMRProcFlow (http://nmrprocflow.org), dedicated to metabolic fingerprinting and targeted metabolomics, covers all spectra processing steps including baseline correction, chemical shift calibration and alignment.
Conclusion
Biologists and NMR spectroscopists can easily interact and develop synergies by visualizing the NMR spectra along with their corresponding experimental-factor levels, thus setting a bridge between experimental design and subsequent statistical analyses.
Swine dysentery caused by Brachyspira hyodysenteriae is a production limiting disease in pig farming. Currently antimicrobial therapy is the only treatment and control method available.
Objective
The aim of this study was to characterize the metabolic response of porcine colon explants to infection by B. hyodysenteriae.
Methods
Porcine colon explants exposed to B. hyodysenteriae were analyzed for histopathological, metabolic and pro-inflammatory gene expression changes.
Results
Significant epithelial necrosis, increased levels of l-citrulline and IL-1α were observed on explants infected with B. hyodysenteriae.
Conclusions
The spirochete induces necrosis in vitro likely through an inflammatory process mediated by IL-1α and NO.
