Diel plant water use and competitive soil cation exchange interact to enhance NH<Subscript>4</Subscript><Superscript>+</Superscript> and K<Superscript>+</Superscript> availability in the rhizosphere

Aims

Hydro-biogeochemical processes in the rhizosphere regulate nutrient and water availability, and thus ecosystem productivity. We hypothesized that two such processes often neglected in rhizosphere models — diel plant water use and competitive cation exchange — could interact to enhance availability of K⁺ and NH₄ ⁺, both high-demand nutrients.

Methods

A rhizosphere model with competitive cation exchange was used to investigate how diel plant water use (i.e., daytime transpiration coupled with no nighttime water use, with nighttime root water release, and with nighttime transpiration) affects competitive ion interactions and availability of K⁺ and NH₄ ⁺.

Results

Competitive cation exchange enabled low-demand cations that accumulate against roots (Ca²⁺, Mg²⁺, Na⁺) to desorb NH₄ ⁺ and K⁺ from soil, generating non-monotonic dissolved concentration profiles (i.e. ‘hotspots’ 0.1–1 cm from the root). Cation accumulation and competitive desorption increased with net root water uptake. Daytime transpiration rate controlled diel variation in NH₄ ⁺ and K⁺ aqueous mass, nighttime water use controlled spatial locations of ‘hotspots’, and day-to-night differences in water use controlled diel differences in ‘hotspot’ concentrations.

Conclusions

Diel plant water use and competitive cation exchange enhanced NH₄ ⁺ and K⁺ availability and influenced rhizosphere concentration dynamics. Demonstrated responses have implications for understanding rhizosphere nutrient cycling and plant nutrient uptake.

     

<Superscript>13</Superscript>C-assisted metabolic flux analysis to investigate heterotrophic and mixotrophic metabolism in <Emphasis Type="Italic">Cupriavidus necator</Emphasis> H16

Introduction

Cupriavidus necator H16 is a gram-negative bacterium, capable of lithoautotrophic growth by utilizing hydrogen as an energy source and fixing carbon dioxide (CO₂) through Calvin–Benson–Bassham (CBB) cycle. The potential to utilize synthesis gas (Syngas) and the prospects of rerouting carbon from polyhydroxybutyrate synthesis to value-added compounds makes C. necator an excellent chassis for industrial application.

Objectives

In the context of lack of sufficient quantitative information of the metabolic pathways and to advance in rational metabolic engineering for optimized product synthesis in C. necator H16, we carried out a metabolic flux analysis based on steady-state ¹³C-labelling.

Methods

In this study, steady-state carbon labelling experiments, using either d-[1-¹³C]fructose or [1,2-¹³C]glycerol, were undertaken to investigate the carbon flux through the central carbon metabolism in C. necator H16 under heterotrophic and mixotrophic growth conditions, respectively.

Results

We found that the CBB cycle is active even under heterotrophic condition, and growth is indeed mixotrophic. While Entner–Doudoroff (ED) pathway is shown to be the major route for sugar degradation, tricarboxylic acid (TCA) cycle is highly active in mixotrophic condition. Enhanced flux is observed in reductive pentose phosphate pathway (redPPP) under the mixotrophic condition to supplement the precursor requirement for CBB cycle. The flux distribution was compared to the mRNA abundance of genes encoding enzymes involved in key enzymatic reactions of the central carbon metabolism.

Conclusion

This study leads the way to establishing ¹³C-based quantitative fluxomics for rational pathway engineering in C. necator H16.

     

Automated metabolite identification from biological fluid <Superscript>1</Superscript>H NMR spectra

Introduction

Metabolite identification in biological samples using Nuclear Magnetic Resonance (NMR) spectra is a challenging task due to the complexity of the biological matrices.

Objectives

This paper introduces a new, automated computational scheme for the identification of metabolites in 1D ¹H NMR spectra based on the Human Metabolome Database.

Methods

The methodological scheme comprises of the sequential application of preprocessing, data reduction, metabolite screening and combination selection.

Results

The proposed scheme has been tested on the 1D ¹H NMR spectra of: (a) an amino acid mixture, (b) a serum sample spiked with the amino acid mixture, (c) 20 blood serum, (d) 20 human amniotic fluid samples, (e) 160 serum samples from publicly available database. The methodological scheme was compared against widely used software tools, exhibiting good performance in terms of correct assignment of the metabolites.

Conclusions

This new robust scheme accomplishes to automatically identify peak resonances in ¹H-NMR spectra with high accuracy and less human intervention with a wide range of applications in metabolic profiling.

     

<Superscript>13</Superscript>C metabolomics reveals widespread change in carbon fate during coral bleaching

Introduction

Rising seawater temperatures are threatening the persistence of coral reefs; where above critical thresholds, thermal stress results in a breakdown of the coral-dinoflagellate symbiosis and the loss of algal symbionts (coral bleaching). As symbiont-derived organic products typically form a major portion of host energy budgets, this has major implications for the fitness and persistence of symbiotic corals.

Objectives

We aimed to determine change in autotrophic carbon fate within individual compounds and downstream metabolic pathways in a coral symbiosis exposed to varying degrees of thermal stress and bleaching.

Methods

We applied gas chromatography–mass spectrometry coupled to a stable isotope tracer (¹³C), to track change in autotrophic carbon fate, in symbiont and host individually, following exposure to elevated water temperature.

Results

Thermal stress resulted in partner-specific changes in carbon fate, which progressed with heat stress duration. We detected modifications to carbohydrate and fatty acid metabolism, lipogenesis, and homeostatic responses to thermal, oxidative and osmotic stress. Despite pronounced photodamage, remaining in hospite symbionts continued to produce organic products de novo and translocate to the coral host. However as bleaching progressed, we observed minimal ¹³C enrichment of symbiont long-chain fatty acids, also reflected in ¹³C enrichment of host fatty acid pools.

Conclusion

These data have major implications for our understanding of coral symbiosis function during bleaching. Our findings suggest that during early stage bleaching, remaining symbionts continue to effectively translocate a variety of organic products to the host, however under prolonged thermal stress there is likely a reduction in the quality of these products.

     

Characterization of ankylosing spondylitis and rheumatoid arthritis using <Superscript>1</Superscript>H NMR-based metabolomics of human fecal extracts

Introduction

The differences in fecal metabolome between ankylosing spondylitis (AS)/rheumatoid arthritis (RA) patients and healthy individuals could be the reason for an autoimmune disorder.

Objectives

The study explored the fecal metabolome difference between AS/RA patients and healthy controls to clarify human immune disturbance.

Methods

Fecal samples from 109 individuals (healthy controls 34, AS 40, and RA 35) were analyzed by ¹H NMR spectroscopy. Data were analyzed with principal component analysis (PCA) and orthogonal projection to latent structure discriminant (OPLS-DA) analysis.

Results

Significant differences in the fecal metabolic profiles could distinguish AS/RA patients from healthy controls but could not distinguish between AS and RA patients. The significantly decreased metabolites in AS/RA patients were butyrate, propionate, methionine, and hypoxanthine. Significantly increased metabolites in AS/RA patients were taurine, methanol, fumarate, and tryptophan.

Conclusion

The metabolome variations in feces indicated AS and RA were two homologous diseases that could not be distinguished by ¹H NMR metabolomics.

     

Modelling the acid/base <Superscript>1</Superscript>H NMR chemical shift limits of metabolites in human urine

Introduction

Despite the use of buffering agents the ¹H NMR spectra of biofluid samples in metabolic profiling investigations typically suffer from extensive peak frequency shifting between spectra. These chemical shift changes are mainly due to differences in pH and divalent metal ion concentrations between the samples. This frequency shifting results in a correspondence problem: it can be hard to register the same peak as belonging to the same molecule across multiple samples. The problem is especially acute for urine, which can have a wide range of ionic concentrations between different samples.

Objectives

To investigate the acid, base and metal ion dependent ¹H NMR chemical shift variations and limits of the main metabolites in a complex biological mixture.

Methods

Urine samples from five different individuals were collected and pooled, and pre-treated with Chelex-100 ion exchange resin. Urine samples were either treated with either HCl or NaOH, or were supplemented with various concentrations of CaCl₂, MgCl₂, NaCl or KCl, and their ¹H NMR spectra were acquired.

Results

Nonlinear fitting was used to derive acid dissociation constants and acid and base chemical shift limits for peaks from 33 identified metabolites. Peak pH titration curves for a further 65 unidentified peaks were also obtained for future reference. Furthermore, the peak variations induced by the main metal ions present in urine, Na⁺, K⁺, Ca²⁺ and Mg²⁺, were also measured.

Conclusion

These data will be a valuable resource for ¹H NMR metabolite profiling experiments and for the development of automated metabolite alignment and identification algorithms for ¹H NMR spectra.

     

Automation of chemical assignment for identifying molecular formula of S-containing metabolites by combining metabolomics and chemoinformatics with <Superscript>34</Superscript>S labeling

Introduction

Sulfur-containing metabolites (S-metabolites) in organisms including plants have unique benefits to humans. So far, few analytical methods have explored such metabolites.

Objectives

We aimed to develop an automatic chemically assigning platform by metabolomics and chemoinformatics with ³⁴S labeling to identify the molecular formula of S-metabolites.

Methods

Direct infusion analysis using Fourier transform ion cyclotron resonance-mass spectrometry provided ultra-high-resolution data including clearly separated isotopic ions—¹⁵N, ³⁴S, ¹⁸O, and ¹³C₂—in the flower, silique, leaf, stem, and root of non-labeled and ³⁴S-labeled Arabidopsis thaliana. Chemoinformatic analysis assigned several elemental compositions of S-metabolites to the acquired S-containing monoisotopic ions using mass accuracy and peak resolution in the non-labeled metabolome data. Possible elemental compositions were characterized on the basis of diagnostic scores of the exact mass and isotopic ion pattern, and a database search. By comparing elemental compositions assigned to the ³⁴S-labeled data with those assigned to the non-labeled data, the elemental composition of S-metabolites were determined. The determined elemental compositions were surveyed using the in-house database, which stores molecular formulae downloaded from metabolome databases.

Results

We identified 35 molecular formulae for known S-metabolites and characterized 72 for unknown. Chemoinformatics required around 1.5 min to analyze a pair of the non-labeled and ³⁴S-labeled data of the organ.

Conclusion

In this study, we developed an automation platform for automatically identifying the presence of S-metabolites. We identified the molecular formula of known S-metabolites, which are accessible in free databases, together with that of unknown. This analytical method did not focus on identifying the structure of S-metabolites, but on the automatic identification of their molecular formula.

     

Anti-hyperglycemic effect of loquat leaf extract is associated with the redistribution of glucose carbon to its metabolites: a <Superscript>13</Superscript>C-tracing study in HepG2 cells

Introduction

Loquat leaf extract (LLE) is commonly used in China for a variety of ailments including diabetes. Several recent reports implicate LLE and a sesquiterpene glycoside, one of its components, as being an anti-hyperglycemic agent. However, the underlying mechanism of action of this anti-hyperglycemic agent has not been reported.

Objective

We have conducted a tracer-based metabolomics study to investigate the effects of sesquiterpene and loquat extract on the balance of flux of central glucose metabolism in HepG2 cells and to compare with those of “insulin sensitizers”, metformin and rosiglitazone.

Methods

Human hepatoma HepG2 cells in confluence culture were incubated in Dulbecco’s modified Eagle’s medium containing 50% [1, 2 ¹³C₂]-glucose in the presence of rosiglitazone, metformin, LLE or pure sesquiterpene. Cells were harvested in 48 h. Mass isotopomers of metabolites (glycogen, ribose, deoxyribose, glutamate and palmitate) were determined.

Results

¹³C labeling in metabolic intermediates were summarized in a mass isotopomer matrix. Treatment with loquat extract/sesquiterpene, metformin and rosiglitazone each produced distinctive mass isotopomer patterns reflecting disparate effects on the contribution of glucose to various metabolites production, and on several metabolic flux ratios. The overall effect of LLE and sesquiterpene on glucose metabolism is clearly different from those of the known “insulin sensitizers”.

Conclusion

Our study demonstrates the utility of isotopomer matrix in summarizing metabolic actions of LLE on the balance of fluxes occurring within the central glucose metabolism in HepG2 cells. ¹³C carbon tracing (tracer-based metabolomics) is a useful systems biology tool to elucidate glucose metabolic pathways affected by diabetes and its treatment.

     

<Superscript>123</Superscript>I-ADAM SPET imaging of serotonin transporter in patients with epilepsy and comorbid depression

Background

Purpose of the study was to investigate alterations in midbrain serotonin transporter (SERT) binding in patients with epilepsy and symptoms of depression compared to patients with epilepsy with no symptoms of depression.

Methods

We studied 12 patients with epilepsy (7 patients had focal and 5 had generalized epilepsy syndromes). The presence of self-reported symptoms of depression was assessed using Beck Depression Inventory (BDI) and the Emotional State Questionnaire (EST-Q). The binding potential of the SERT was assessed by performing brain single photon emission tomography (SPET) using the SERT radioligand 2-((2-((dimethylamino)methyl)phenyl)thio)-5-(123)iodophenylamine (¹²³I-ADAM).

Results

Seven patients had BDI and EST-Q subscale scores greater than 11 points, which was interpreted as the presence of symptoms of depression. We found that ¹²³I-ADAM binding was not significantly different between patients with epilepsy with and without symptoms of depression. In addition, ¹²³I-ADAM binding did not show a significant correlation to either BDI or EST-Q depression subscale scores and did not differ between patients with focal vs. generalized epilepsy.

Conclusion

The results of our study failed to demonstrate alterations of SERT binding properties in patients with epilepsy with or without symptoms of depression.

     

Measurement of the <Superscript>15</Superscript>N/<Superscript>14</Superscript>N ratio of phenylalanine in fermentation matrix by isotope ratio mass spectrometry

Objectives

To determine the origin of ¹⁵N-labeled phenylalanine in microbial metabolic flux analysis using ¹⁵N as a tracer, a method for measuring phenylalanine δ¹⁵N using HPLC coupled with elemental analysis-isotope ratio mass spectrometry (EA-IRMS) was developed.

Results

The original source of the ¹⁵N-labeled phenylalanine was determined using this new method that consists of three steps: optimization of the HPLC conditions, evaluation of the isotope fractionation effects, and evaluation of the effect of pre-processing on the phenylalanine nitrogen stable isotope. In addition, the use of a ¹⁵N-labeled inorganic nitrogen source, rather than ¹⁵N-labeled amino acids, was explored using this method.

Conclusions

The method described here can also be applied to the analysis of metabolic flux.

     

G-CSF/anti-G-CSF antibody complexes drive the potent recovery and expansion of CD11b<Superscript>+</Superscript>Gr-1<Superscript>+</Superscript> myeloid cells without compromising CD8<Superscript>+</Superscript> T cell immune responses

Background

Administration of recombinant G-CSF following cytoreductive therapy enhances the recovery of myeloid cells, minimizing the risk of opportunistic infection. Free G-CSF, however, is expensive, exhibits a short half-life, and has poor biological activity in vivo.

Methods

We evaluated whether the biological activity of G-CSF could be improved by pre-association with anti-G-CSF mAb prior to injection into mice.

Results

We find that the efficacy of G-CSF therapy can be enhanced more than 100-fold by pre-association of G-CSF with an anti-G-CSF monoclonal antibody (mAb). Compared with G-CSF alone, administration of G-CSF/anti-G-CSF mAb complexes induced the potent expansion of CD11b⁺Gr-1⁺ myeloid cells in mice with or without concomitant cytoreductive treatment including radiation or chemotherapy. Despite driving the dramatic expansion of myeloid cells, in vivo antigen-specific CD8⁺ T cell immune responses were not compromised. Furthermore, injection of G-CSF/anti-G-CSF mAb complexes heightened protective immunity to bacterial infection. As a measure of clinical value, we also found that antibody complexes improved G-CSF biological activity much more significantly than pegylation.

Conclusions

Our findings provide the first evidence that antibody cytokine complexes can effectively expand myeloid cells, and furthermore, that G-CSF/anti-G-CSF mAb complexes may provide an improved method for the administration of recombinant G-CSF.

     

Pathway-specific metabolome analysis with <Superscript>18</Superscript>O<Subscript>2</Subscript>-labeled <Emphasis Type="Italic">Medicago truncatula</Emphasis> via a mass spectrometry-based approach

Introduction

Oxygen from carbon dioxide, water or molecular oxygen, depending on the responsible enzyme, can lead to a large variety of metabolites through chemical modification.

Objectives

Pathway-specific labeling using isotopic molecular oxygen (¹⁸O₂) makes it possible to determine the origin of oxygen atoms in metabolites and the presence of biosynthetic enzymes (e.g., oxygenases). In this study, we established the basis of ¹⁸O₂-metabolome analysis.

Methods

¹⁸O₂ labeled whole Medicago truncatula seedlings were prepared using ¹⁸O₂-air and an economical sealed-glass bottle system. Metabolites were analyzed using high-accuracy and high-resolution mass spectrometry. Identification of the metabolite was confirmed by NMR following UHPLC–solid-phase extraction (SPE).

Results

A total of 511 peaks labeled by ¹⁸O₂ from shoot and 343 peaks from root were annotated by untargeted metabolome analysis. Additionally, we identified a new flavonoid, apigenin 4′-O-[2′-O-coumaroyl-glucuronopyranosyl-(1–2)-O-glucuronopyranoside], that was labeled by ¹⁸O₂. To the best of our knowledge, this is the first report of apigenin 4′-glucuronide in M. truncatula. Using MSⁿ analysis, we estimated that ¹⁸O atoms were specifically incorporated in apigenin, the coumaroyl group, and glucuronic acid. For apigenin, an ¹⁸O atom was incorporated in the 4′-hydroxy group. Thus, non-specific incorporation of an ¹⁸O atom by recycling during one month of labeling is unlikely compared with the more specific oxygenase-catalyzing reaction.

Conclusion

Our finding indicated that ¹⁸O₂ labeling was effective not only for the mining of unknown metabolites which were biosynthesized by oxygenase-related pathway but also for the identification of metabolites whose oxygen atoms were derived from oxygenase activity.

     

Accumulation of Fascin<Superscript>+</Superscript> cells during experimental autoimmune neuritis

Background

Experimental autoimmune neuritis (EAN) is a well-known animal model of human demyelinating polyneuropathies and is characterized by inflammation and demyelination in the peripheral nervous system. Fascin is an evolutionarily highly conserved cytoskeletal protein of 55 kDa containing two actin binding domains that cross-link filamentous actin to hexagonal bundles.

Methods

Here we have studied by immunohistochemistry the spatiotemporal accumulation of Fascin?+?cells in sciatic nerves of EAN rats.

Results

A robust accumulation of Fascin?+?cell was observed in the peripheral nervous system of EAN which was correlated with the severity of neurological signs in EAN.

Conclusion

Our results suggest a pathological role of Fascin in EAN.

Virtual slides

The virtual slides for this article can be found here: http://www.diagnosticphatology.diagnomx.eu/vs/6734593451114811

     

<Superscript>1</Superscript>H NMR metabolic profiling of gastric cancer patients with lymph node metastasis

Introduction

Gastric cancer (GC) is a malignant tumor worldwide. As primary pathway for metastasis, the lymphatic system is an important prognostic factor for GC patients. Although the metabolic changes of gastric cancer have been investigated in extensive studies, little effort focused on the metabolic profiling of lymph node metastasis (LNM)-positive or negative GC patients.

Objectives

We performed ¹H NMR spectrum of GC tissue samples with and without LNM to identify novel potential metabolic biomarkers in the process of LNM of GC.

Methods

¹H NMR-based untargeted metabolomics approach combined with multivariate statistical analyses were used to study the metabolic profiling of tissue samples from LNM-positive GC patients (n?=?40), LNM-negative GC patients (n?=?40) and normal controls (n?=?40).

Results

There was a clear separation between GC patients and normal controls, and 33 differential metabolites were identified in the study. Moreover, GC patients were also well-classified according to LNM-positive or negative. Totally eight distinguishing metabolites were selected in the metabolic profiling of GC patients with LNM-positive or negative, suggesting the metabolic dysfunction in the process of LNM. According to further validation and analysis, especially BCAAs metabolism (leucine, isoleucine, valine), GSH and betaine may be as potential factors of diagnose and prognosis of GC patients with or without LNM.

Conclusion

To our knowledge, this is the first metabolomics study focusing on LNM of GC. The identified distinguishing metabolites showed a promising application on clinical diagnose and therapy prediction, and understanding the mechanism underlying the carcinogenesis, invasion and metastasis of GC.

     

Investigation of species and environmental effects on rhubarb roots metabolome using <Superscript>1</Superscript>H NMR combined with high performance thin layer chromatography

Introduction

The pharmacological activities of medicinal plants are reported to be due to a wide range of metabolites, therein, the concentrations of which are greatly affected by many genetic and/or environmental factors. In this context, a metabolomics approach has been applied to reveal these relationships. The investigation of such complex networks that involve the correlation between multiple biotic and abiotic factors and the metabolome, requires the input of information acquired by more than one analytical platform. Thus, development of new metabolomics techniques or hyphenations is continuously needed.

Objectives

Feasibility of high performance thin-layer chromatography (HPTLC) were investigated as a supplementary tool for medicinal plants metabolomics supporting ¹H nuclear magnetic resonance (¹H NMR) spectroscopy.

Method

The overall metabolic difference of plant material collected from two species (Rheum palmatum and Rheum tanguticum) in different geographical locations and altitudes were analyzed by ¹H NMR- and HPTLC-based metabolic profiling. Both NMR and HPTLC data were submitted to multivariate data analysis including principal component analysis and orthogonal partial least square analysis.

Results

The NMR and HPTLC profiles showed that while chemical variations of rhubarb are in some degree affected by all the factors tested in this study, the most influential factor was altitude of growth. The metabolites responsible for altitude differentiation were chrysophanol, emodin and sennoside A, whereas aloe emodin, catechin, and rhein were the key species-specific markers.

Conclusion

These results demonstrated the potential of HTPLC as a supporting tool for metabolomics due to its high profiling capacity of targeted metabolic groups and preparative capability.

     

Characterizing alternative feeds for rainbow trout (<Emphasis Type="Italic">O. mykiss</Emphasis>) by <Superscript>1</Superscript>H NMR metabolomics

Introduction

Fish feed formulations are constantly evolving to improve the quality of diets for farmed fish and to ensure the sustainability of the aquaculture sector. Nowadays, insect, microalgae and yeast are feedstuff candidates for new feeds. However, the characterization of aquafeed is still based on proximate and targeted analyses which may not be sufficient to assess feed quality.

Objectives

Our aim was to highlight the soluble compounds that specifically differ between selected plant-based feeds complemented with alternative feedstuffs and discuss their origin and potential for fish nutrition.

Methods

A growth trial was carried out to evaluate growth performances and feed conversion ratios of fish fed plant-based, commercial, insect, spirulina and yeast feeds. ¹H NMR metabolomics profiling of each feed was performed using a CPMG sequence on polar extracts. Spectra were processed, and data were analyzed using multivariate and univariate analyses to compare alternative feeds to a plant-based feed.

Results

Fish fed insect or yeast feed showed the best growth performances associated with the lowest feed conversion ratios compared to plant-based feed. Soluble compound ¹H NMR profiles of insect and spirulina alternative feeds differed significantly from the plant-based one that clustered with yeast feed. In insect and spirulina feeds, specific differences compared to plant-based feed concerned glycerol and 3-hydroxybutyrate, respectively.

Conclusion

This strategy based on compositional differences between plant-based and alternative feeds can be useful for detecting compounds unsuspected until now that could impact fish metabolism.

     

rDolphin: a GUI R package for proficient automatic profiling of 1D <Superscript>1</Superscript>H-NMR spectra of study datasets

Introduction

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.

     

A multidimensional <Superscript>1</Superscript>H NMR lipidomics workflow to address chemical food safety issues

Introduction

Although it is still at a very early stage compared to its mass spectrometry (MS) counterpart, proton nuclear magnetic resonance (NMR) lipidomics is worth being investigated as an original and complementary solution for lipidomics. Dedicated sample preparation protocols and adapted data acquisition methods have to be developed to set up an NMR lipidomics workflow; in particular, the considerable overlap observed for lipid signals on 1D spectra may hamper its applicability.

Objectives

The study describes the development of a complete proton NMR lipidomics workflow for application to serum fingerprinting. It includes the assessment of fast 2D NMR strategies, which, besides reducing signal overlap by spreading the signals along a second dimension, offer compatibility with the high-throughput requirements of food quality characterization.

Method

The robustness of the developed sample preparation protocol is assessed in terms of repeatability and ability to provide informative fingerprints; further, different NMR acquisition schemes—including classical 1D, fast 2D based on non-uniform sampling or ultrafast schemes—are evaluated and compared. Finally, as a proof of concept, the developed workflow is applied to characterize lipid profiles disruption in serum from β-agonists diet fed pigs.

Results

Our results show the ability of the workflow to discriminate efficiently sample groups based on their lipidic profile, while using fast 2D NMR methods in an automated acquisition framework.

Conclusion

This work demonstrates the potential of fast multidimensional ¹H NMR—suited with an appropriate sample preparation—for lipidomics fingerprinting as well as its applicability to address chemical food safety issues.

     

Comprehensive <Superscript>1</Superscript>H NMR metabolic profiling of body fluids for differentiation of meningitis in adults

Introduction

Meningitis, a morbidly infectious central nervous system pathology is accompanied by acute inflammation of the meninges, causing raised intracranial pressure linked with serious neurological sequelae.

Objective

To observe the variation in the metabolic profile, that may occur in serum and urine along with CSF in adults using ¹H NMR spectroscopy, with an attempt of appropriate and timely treatment regimen.

Methods

The ¹H NMR-based metabolomics has been performed in 115 adult subjects for differentiating bacterial meningitis (BM) and tubercular meningitis (TBM).

Results

The discriminant function analysis (DFA) of the three bio-fluids collectively identified 3-hydroxyisovalerate, lactate, glucose, formate, valine, alanine, ketonic bodies, malonate and choline containing compounds (choline and GPC) as significant metabolites among cases versus control group. The differentiation of bacterial meningitis and tuberculous meningitis (BM vs. TBM) can be done on the basis of identification of 3-hydroxyisovalerate, isobutyrate and formate in case of CSF (with a correct classification of 78 %), alanine in serum (correct classification 60 %), valine and acetone in case of urine (correct classification 89.1 %). The NMR spectral bins based orthogonal signal correction principal component analysis score plots of significant metabolites obtained from DFA also provided group classification among cases versus control group in CSF, serum and urine samples. The variable importance in projection scores also identified similar significant metabolites as obtained from DFA, collectively in CSF, serum and urine samples, responsible for differentiation of meningitis.

Conclusion

The CSF contained metabolites which are formed during infection and inflammation, and these were also found in significant quantity in serum and urine samples.

     

Maize rhizosphere priming: field estimates using <Superscript>13</Superscript>C natural abundance

Introduction

Root-mediated changes in soil organic matter (SOM) decomposition, termed rhizosphere priming effects (RPE), play crucial roles in the global carbon (C) cycle, but their mechanisms and field relevance remain ambiguous. We hypothesize that nitrogen (N) shortages may intensify SOM decomposition in the rhizosphere because of increase of fine roots and rhizodeposition.

Methods

RPE and their dependence on N-fertilization were studied using a C₃-to-C₄ vegetation change. N-fertilized and unfertilized soil cores, with and without maize, were incubated in the field for 50 days. Soil CO₂ efflux was measured, partitioned for SOM- and root-derived CO₂, and RPE was calculated. Plant biomass, microbial biomass C (MBC) and N (MBN), and enzyme activities (β-1,4-glucosidase; N-acetylglucosaminidase; L-leucine aminopeptidase) were analyzed.

Results

Roots enhanced SOM mineralization by 35 % and 126 % with and without N, respectively. This was accompanied by higher specific root-derived CO₂ in unfertilized soils. MBC, MBN and enzyme activities increased in planted soils, indicating microbial activation, causing positive RPE. N-fertilization had minor effects on MBC and MBN, but it reduced β-1,4-glucosidase and L-leucine aminopeptidase activities under maize through lower root-exudation. In contrast, N-acetylglucosaminidase activity increased with N-fertilization in planted and unplanted soils.

Conclusions

This study showed the field relevance of RPE and confirmed that, despite higher root biomass, N availability reduces RPE by lowering root and microbial activity.