Tag Archives: metabolomics

Using nuclear magnetic resonance urine metabolomics to develop a prediction model of early stages of renal disease in subjects with type 2 diabetes

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J Ricardo Lucio-Gutiérrez, Paula Cordero-Pérez, Iris C Farías-Navarro, Ramiro Tijerina-Marquez, Concepción Sánchez-Martínez, José Luis Ávila-Velázquez, Pedro A García-Hernández, Homero Náñez-Terreros, Jordi Coello-Bonilla, Míriam Pérez-Trujillo, Teodor Parella, Liliana Torres-González, Noemí H Waksman-Minsky, Alma L Saucedo

Journal of Pharmaceutical and Biomedical Analysis, 2022, 19, 114885

Abstract

Type 2 diabetes mellitus (DM2) is a multimorbidity, long-term condition, and one of the worldwide leading causes of chronic kidney disease (CKD) –a silent disease, usually detected when non-reversible renal damage have already occurred. New strategies and more effective laboratory methods are needed for more opportune diagnosis of DM2-CKD. This study comprises clinical parameters and nuclear magnetic resonance (NMR)-based urine metabolomics data from 60 individuals (20–65 years old, 67.7% females), sorted in 5 experimental groups (healthy subjects; diabetic patients without any clinical sign of CKD; and patients with mild, moderate, and severe DM2-CKD), according to KDIGO. DM2-CKD produces a continuous variation of the urine metabolome, characterized by an increase/decrement of a group of metabolites that can be used to monitor CKD progression (trigonelline, hippurate, phenylalanine, glycolate, dimethylamine, alanine, 2-hydroxybutyrate, lactate, and citrate). NMR profiles were used to obtain a statistical model, based on partial least squares analysis (PLS-DA) to discriminate among groups. The PLS-DA model yielded good validation parameters (sensitivity, specificity, and area under the curve (AUC) of the receiver operating characteristic curve (ROC) plot: 0.692, 0.778 and 0.912, respectively) and, thus, it can differentiate between subjects with DM2-CKD in early stages, from subjects with a mild or severe condition. This metabolic signature exhibits a molecular variation associated to DM2-CKD, and data suggests it can be used to predict risk of DM2-CKD in patients without clinical signs of renal disease, offering a new alternative to current diagnosis methods.

Fatty Acids and Metabolomic Composition of Follicular Fluid Collected from Environments Associated with Good and Poor Oocyte Competence in Goats

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Dolors Izquierdo, Montserrat Roura, Míriam Pérez-Trujillo, Sandra Soto-Heras, María-Teresa Paramio

International Journal of Molecular Sciences 2022, 23(8), 4141

Abstract

In goats, embryo oocyte competence is affected by follicle size regardless the age of the females. In previous studies we have found differences in blastocyst development between oocytes coming of small (<3 mm) and large follicles (>3 mm) in prepubertal (1–2 months-old) goats. Oocyte competence and Follicular Fluid (FF) composition changes throughout follicle growth. The aim of this study was to analyze Fatty Acids (FAs) composition and metabolomic profiles of FF recovered from small and large follicles of prepubertal goats and follicles of adult goats. FAs were analyzed by chromatography and metabolites by 1H-Nuclear Magnetic Resonance (1H-NMR) Spectrometry. The results showed important differences between adult and prepubertal follicles: (a) the presence of α,β-glucose in adult and no detection in prepubertal; (b) lactate, -N-(CH3)3 groups and inositol were higher in prepubertal (c) the percentage of Linolenic Acid, Total Saturated Fatty Acids and n-3 PUFAs were higher in adults; and (d) the percentage of Linoleic Acid, total MUFAs, PUFAs, n-6 PUFAs and n-6 PUFAs: n-3 PUFAs ratio were higher in prepubertal goats. Not significant differences were found in follicle size of prepubertal goats, despite the differences in oocyte competence for in vitro embryo production.

Representative 1H NMR spectra of follicular fluid samples of (a) adult and (b) prepubertal goats. Spectra were acquired at 298.0 K and at a magnetic field of 600 MHz, with suppression of the residual water signal.

(a) PCA scores plot (PC1-PC2) from 1H NMR spectral data of follicular fluid samples of prepubertal (n = 16; blue dots) and adult (n = 40; black dots) goats. (b) PCA heat map loadings plot (PC1-PC2) with some discriminant variables assigned.

SeRMN contribution to SMASH Small Molecule NMR Conference

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Kumar Motiram-Corral is presenting at SMASH 2021 Conference a talk entitled In situ Enantiospecific Detection of Multiple Metabolites in Mixtures using NMR Spectroscopy, related to some of our recent research work. The presentation will be 1st of September in the section “Unveiling the Unknown – New Methods in Structure Elucidation“.

L. T. Kuhn, K. Motiram-Corral, T. J. Athersuch, T. Parella, M. Pérez-Trujillo, Angew. Chem. Int. Ed. 59 (2020) 23615.

Metabolomics and transcriptomics to decipher molecular mechanisms underlying ectomycorrhizal root colonization of an oak tree

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M. Sebastiana, A. Gargallo-Garriga, J. Sardans, M. Pérez-Trujillo, F. Monteiro, A. Figueiredo, M. Maia, R. Nascimento, M. Sousa Silva, A. N. Ferreira, C. Cordeiro, A. P. Marques, L. Sousa, R. Malhó & J. Peñuelas

Scientific Reports volume 11, Article number: 8576 (2021). https://doi.org/10.1038/s41598-021-87886-5

Mycorrhizas are known to have a positive impact on plant growth and ability to resist major biotic and abiotic stresses. However, the metabolic alterations underlying mycorrhizal symbiosis are still understudied. By using metabolomics and transcriptomics approaches, cork oak roots colonized by the ectomycorrhizal fungus Pisolithus tinctorius were compared with non-colonized roots. Results from this global metabolomics analysis suggest decreases in root metabolites which are common components of exudates, and in compounds related to root external protective layers which could facilitate plant-fungal contact and enhance symbiosis. Root metabolic pathways involved in defense against stress were induced in ectomycorrhizal roots that could be involved in a plant mechanism to avoid uncontrolled growth of the fungal symbiont in the root apoplast. Several of the identified symbiosis-specific metabolites, such as GABA, may help to understand how ectomycorrhizal fungi such as P. tinctorius benefit their host plants.

SeRMN contribution at EUROMAR 2021 Conference

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Some of our recent research work was presented at the European NMR meeting Euromar 2021 that was going to take place at Portoroz (Slovenia), but which was finally virtual from the 5th to the 8th of July 2021.

· Míriam Pérez-Trujillo presented the talk In situ Enantiospecific Detection of Multiple Metabolites in Mixtures using NMR Spectroscopy in the “Metabolomics” session. In this talk our last advances in enantiodifferentiation using NMR were shown and discussed.

To date, the enantiospecific analysis of mixtures necessarily requires prior separation of the individual components. The simultaneous enantiospecific detection of multiple chiral molecules in a mixture represents a major challenge, which would lead to a significantly better understanding of the underlying biological processes; e.g. via enantiospecifically analyzing metabolites in their native environment. Here, we report on the first in situ enantiospecific detection of a thirty-nine-component mixture. As a proof of concept, eighteen essential amino acids (AAs) at physiological concentrations were simultaneously enantiospecifically detected using NMR spectroscopy and a chiral solvating agent. This work represents a first step towards the simultaneous multicomponent enantiospecific analysis of complex mixtures, a capability that will have substantial impact on metabolism studies, metabolic phenotyping, chemical reaction monitoring, and many other fields where complex mixtures containing chiral molecules require efficient characterization.

L. T. Kuhn, K. Motiram-Corral, T. J. Athersuch, T. Parella, M. Pérez-Trujillo, Angew. Chem. Int. Ed. 59 (2020) 23615.

SeRMN contributions at 10th GERMN biennial /9th IberAmerican/7th Iberian NMR Meeting

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Some of the SeRMN staff has presented our recent research work at the biannual Spanish and IberAmerican NMR meeting, 10th GERMN biennial /9th IberAmerican/7th Iberian NMR Meeting. This year it was a virtual meeting taking place from 26 to 29 April 2021.

Pau Nolis presented an oral communication entitled “Reducing experimental time using Multiple Fid Acquisition“. P. Nolis, K. Motiram-Corral, M. Pérez-Trujillo, T. Parella.

Speeding-up NMR molecular analysis is an important research field which has been continuously advancing since NMR early days. The relevant benefits are clear and evident: i) reduce analysis time per sample => reduce analysis cost; ii) gain spectrometer time to analyze new samples => improve spectrometer efficiency. Multiple FID Acquisition (MFA) strategy consists in the design of NMR pulse sequence experiments accommodating N acquisition windows, each registering different relevant structural information. This strategy is faster
than perform a traditional sequential acquisition of N separated experiments. Several design strategies and practical experiments will be shown and discussed.

Míriam Pérez-Trujillo presented an oral communication entitled “Simultaneous Enantiospecific Detection of Multiple Metabolites in Mixtures using NMR Spectroscopy“. L. T. Kuhn, K. Motiram-Corral, T. J. Athersuch, T. Parella, M. Pérez-Trujillo.

Chirality plays a fundamental role in nature, but its detection and quantification still face many limitations. To date, the enantiospecific analysis of mixtures necessarily requires prior separation of the individual components. The simultaneous enantiospecific detection of multiple chiral molecules in a mixture represents a major challenge, which would lead to a
significantly better understanding of the underlying biological processes; e.g. via enantiospecifically analyzing metabolites in their native environment. Here, we report on the first in situ enantiospecific detection of a thirty-ninecomponent mixture. As a proof of concept, eighteen essential amino acids (AAs) at physiological concentrations were simultaneously enantiospecifically detected using NMR spectroscopy and a chiral solvating agent. This work
represents a first step towards the simultaneous multicomponent enantiospecific analysis of complex mixtures, a capability that will have substantial impact on metabolism studies, metabolic phenotyping, chemical reaction monitoring, and many other fields where complex mixtures containing chiral molecules require efficient characterization.

Special Issue: NMR-Based Metabolomics

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Special Issue: NMR-Based Metabolomics, by Míriam Pérez-Trujillo* and Toby J. Athersuch*

Molecules2021, 26(11), 3283; https://doi.org/10.3390/molecules26113283

This article belongs to the Special Issue NMR-Based Metabolomics

Nuclear magnetic resonance (NMR) spectroscopy remains one of the core analytical platforms for metabolomics, providing complementary chemical information to others, such as mass spectrometry, and offering particular advantages in some areas of research on account of its inherent robustness, reproducibility, and phenomenal dynamic range. While routine experimental protocols for profiling and related statistical analysis pipelines have been established, they often present considerable challenges to the analyst, including spectral overlap, accurate and precise quantification, and chemical shift variation. Consequently, there is still much activity across all areas of NMR spectroscopic analysis in relation to metabolomics. Furthermore, there remain many biological systems and sample types that have not been extensively explored using NMR spectroscopy-based metabolomics.In this Special Issue, several advances in methodology, and new applications in the field of NMR-based metabolomics, have been presented. In addition, the SI includes authoritative review articles focused on the state-of-the-art of quantitative NMR spectroscopy in biomedical metabolomics applications, and novel applications in the agri-food sector.

Metabolomic fingerprinting of pig seminal plasma identifies in vivo fertility biomarkers

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PREPRINT VERSION AT Journal of Animal Science and Biotechnology

Yentel Mateo-Otero, Pol Fernández-López, Ariadna Delgado-Bermúdez, Pau Nolis, Jordi Roca, Jordi Miró, Isabel Barranco, Marc Yeste

Background

Metabolomic approaches, which include the study of low molecular weight molecules, is an emerging -omics technology useful for the identification of biomarkers. In this field, nuclear magnetic resonance (NMR) spectroscopy approach has already been used to uncover (in)fertility biomarkers in the seminal plasma (SP) of several mammalian species. However, NMR studies profiling SP metabolome to uncover in vivo fertility biomarkers are yet to be carried out in pigs. Thus, this study aimed to evaluate the putative relationship between the presence/concentration of SP-metabolites and in vivo fertility outcomes. To this end, 24 entire ejaculates (three ejaculates per boar) were collected from artificial insemination (AI)-boars throughout a year (one ejaculate every four months). Immediately after collection, ejaculates were centrifuged (1,500×g for 10 min twice) to obtain SP-samples and were stored (− 80°C) for subsequent metabolomic analysis by NMR spectroscopy. Fertility outcomes from 1,525 inseminations were recorded over a year, including farrowing rate, litter size, stillbirths per litter and the duration of pregnancy. These data were corrected to isolate the direct boar effect on each in vivo fertility parameter using a multivariate statistical model.

Results

A total of 24 metabolites were identified and quantified in all SP-samples. ROC curve analysis showed that lactate levels in SP had discriminative capacity for farrowing rate (area under the curve (AUC) = 0.764; P < 0.05) while carnitine (AUC = 0.847), hypotaurine (AUC = 0.819), sn-glycero-3-phosphocholine (AUC = 0.833), glutamate (AUC = 0.799) and glucose (AUC = 0.750) had it for litter size (P < 0.05). Similarly, citrate (AUC = 0.743), creatine (AUC = 0.812), phenylalanine (AUC = 0.750), tyrosine (AUC = 0.753) and malonate (AUC = 0.868) levels had discriminative capacity for stillbirths per litter (P < 0.05); and malonate (AUC = 0.767) and fumarate (AUC = 0.868) concentrations for gestation length (P < 0.05).

Conclusions

Considering these results, the assessment of selected SP-metabolites in ejaculates through NMR spectroscopy could be considered as a promising non-invasive tool to predict in vivo fertility outcomes in pigs. Moreover, supplementing AI-doses with specific metabolites should also be contemplated as a way to improve their fertility potential.

Simultaneous Enantiospecific Detection of Multiple Compounds in Mixtures using NMR Spectroscopy

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Simultaneous Enantiospecific Detection of Multiple Compounds in Mixtures using NMR Spectroscopy, by Lars T. Kuhn, Kumar Motiram-Corral, Toby J. Athersuch, Teodor Parella, Míriam Pérez-Trujillo*

Angew. Chem. Int. Ed., 2020 / doi:10.1002/anie.202011727

Chirality plays a fundamental role in nature, but its detection and quantification still face many limitations. To date, the enantiospecific analysis of mixtures necessarily requires prior separation of the individual components. The simultaneous enantiospecific detection of multiple chiral molecules in a mixture represents a major challenge, which would lead to a significantly better understanding of the underlying biological processes; e.g. via enantiospecifically analysing metabolites in their native environment. Here, we report on the first in situ enantiospecific detection of a thirty‐nine‐component mixture. As a proof of concept, eighteen essential amino acids at physiological concentrations were simultaneously enantiospecifically detected using NMR spectroscopy and a chiral solvating agent. This work represents a first step towards the simultaneous multicomponent enantiospecific analysis of complex mixtures, a capability that will have substantial impact on metabolism studies, metabolic phenotyping, chemical reaction monitoring, and many other fields where complex mixtures containing chiral molecules require efficient characterisation.

Simultaneous enantiospecific detection of a mixture of amino acids by NMR spectroscopy

This work has been selected to be presented as a talk at 2021 scientific conferences:

· 42nd FGMR (German Chemical Society, Magnetic Resonance Section) Annual Discussion Meeting – Virtual, Sep 27 to Oct 1.

· SMASH- Small Molecule NMR Conference 2021 – Virtual, Aug 30 to Sep 2.

· Euromar 2021 Conference – Virtual, 5 to 8 July.

· 10th GERMN (Spanish NMR group of the Real Sociedad Española de Química) biennial & 9th IberoAmerican NMR Meeting – Virtual, 26 to 19 April.

31P-NMR Metabolomics Revealed Species-Specific Use of Phosphorous in Trees

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31P-NMR Metabolomics Revealed Species-Specific Use of Phosphorous in Trees of a French Guiana Rainforest, by Gargallo-Garriga, A.; Sardans, J.; Llusià, J.; Peguero, G.; Asensio, D.; Ogaya, R.; Urbina, I.; Langenhove, L.V.; Verryckt, L.T.; Courtois, E.A.; Stahl, C.; Grau, O.; Urban, O.; Janssens, I.A.; Nolis, P.; Pérez-Trujillo, M.; Parella, T.; Peñuelas, J.  Molecules 2020, 25, 3960. https://doi.org/10.3390/molecules25173960

Productivity of tropical lowland moist forests is often limited by availability and functional allocation of phosphorus (P) that drives competition among tree species and becomes a key factor in determining forestall community diversity. We used non-target 31P-NMR metabolic profiling to study the foliar P-metabolism of trees of a French Guiana rainforest. The objective was to test the hypotheses that P-use is species-specific, and that species diversity relates to species P-use and concentrations of P-containing compounds, including inorganic phosphates, orthophosphate monoesters and diesters, phosphonates and organic polyphosphates. We found that tree species explained the 59% of variance in 31P-NMR metabolite profiling of leaves. A principal component analysis showed that tree species were separated along PC 1 and PC 2 of detected P-containing compounds, which represented a continuum going from high concentrations of metabolites related to non-active P and P-storage, low total P concentrations and high N:P ratios, to high concentrations of P-containing metabolites related to energy and anabolic metabolism, high total P concentrations and low N:P ratios. These results highlight the species-specific use of P and the existence of species-specific P-use niches that are driven by the distinct species-specific position in a continuum in the P-allocation from P-storage compounds to P-containing molecules related to energy and anabolic metabolism.

This article belongs to the Special Issue:

https://www.mdpi.com/journal/molecules/special_issues/nmr_metabolomics