1H-NMR Metabolomic Profiling Research Articles

Tissue-Specific 1H-NMR Metabolomic Profiling in Mice with Adenine-Induced Chronic Kidney Disease.

Chronic kidney disease (CKD) results in the impaired filtration of metabolites, which may be toxic or harmful to organs/tissues. The objective of this study was to perform unbiased 1H nuclear magnetic resonance (NMR)-based metabolomics profiling of tissues from mice with CKD. Five-month-old male C57BL6J mice were placed on either a casein control diet or adenine-supplemented diet to induce CKD for 24 weeks. CKD was confirmed by significant increases in blood urea nitrogen (24.1 ± 7.7 vs. 105.3 ± 18.3 mg/dL, p < 0.0001) in adenine-fed mice. Following this chronic adenine diet, the kidney, heart, liver, and quadriceps muscles were rapidly dissected; snap-frozen in liquid nitrogen; and the metabolites were extracted. Metabolomic profiling coupled with multivariate analyses confirm clear separation in both aqueous and organic phases between control and CKD mice. Severe energetic stress and apparent impaired mitochondrial metabolism were observed in CKD kidneys evidenced by the depletion of ATP and NAD+, along with significant alterations in tricarboxylic acid (TCA) cycle intermediates. Altered amino acid metabolism was observed in all tissues, although significant differences in specific amino acids varied across tissue types. Taken together, this study provides a metabolomics fingerprint of multiple tissues from mice with and without severe CKD induced by chronic adenine feeding.

Intracellular sodium elevation reprograms cardiac metabolism

Intracellular Na elevation in the heart is a hallmark of pathologies where both acute and chronic metabolic remodelling occurs. Here, we assess whether acute (75 μM ouabain 100 nM blebbistatin) or chronic myocardial Nai load (PLM3SA mouse) are causally linked to metabolic remodelling and whether the failing heart shares a common Na-mediated metabolic ‘fingerprint’. Control (PLMWT), transgenic (PLM3SA), ouabain-treated and hypertrophied Langendorff-perfused mouse hearts are studied by 23Na, 31P, 13C NMR followed by 1H-NMR metabolomic profiling. Elevated Nai leads to common adaptive metabolic alterations preceding energetic impairment: a switch from fatty acid to carbohydrate metabolism and changes in steady-state metabolite concentrations (glycolytic, anaplerotic, Krebs cycle intermediates). Inhibition of mitochondrial Na/Ca exchanger by CGP37157 ameliorates the metabolic changes. In silico modelling indicates altered metabolic fluxes (Krebs cycle, fatty acid, carbohydrate, amino acid metabolism). Prevention of Nai overload or inhibition of Na/Camito may be a new approach to ameliorate metabolic dysregulation in heart failure.

NMR-based metabolomics reveals effect of Ganoderma boninense infection on oil palm leaf at 30 days post-infection

Basal stem rot is the major disease in oil palm industry that caused by a fungal named Ganoderma boninense (G. boninense) species. Infected palms are symptomless at the early stage of this disease which imposes difficulties in detecting the disease. Therefore, this study was carried out to obtain the 1H NMR metabolomic profiling of both non-infected and G. boninense infected oil palm leaf at 30 days post-infection (dpi). This combination has provided a rapid approach in investigating the changes in the compound variations of non-infected and G. boninense infected oil palm leaf. Non-infected and G. boninense infected oil palm leaf at 30 dpi was extracted using aqueous methanol (methanol: water, 80: 20 v/v). The crude extracts obtained were analyzed by 1H NMR-based metabolomics approach. Analysis of metabolomics data from 1H NMR was conducted by multivariate data analysis of principal component analysis (PCA). Significant differences were found between the two groups. Compared to the non-infected leaf, the G. boninense infected leaf had higher relative abundance of choline, asparagine, alanine, succinic acid, gallic acid, epicatechin, trimethylamine, N-acetylglucosamine, N-acetyltyrosine, β-sitosterol, 2,3-butanediol, lactic acid, caffeic acid, p-hydroxybenzoic acid, α-tocopherol, β-cryptoxanthin and kaempferol. The non-infected leaf showed higher level of sucrose, xylose, α-glucose, S-sulfocysteine and indole-3-acetic acid. NMR-based metabolomics applied in this study reveals that G. boninense alters a manifold of primary and secondary compounds in oil palm leaf.

1 H NMR metabolomic profiling elucidated attenuation of neurometabolic alterations by lercanidipine in MCAo model in rats.

Stroke is a leading cause of death and disability worldwide with limited therapeutic interventions. The current study explored proton nuclear magnetic resonance spectroscopy (1 H NMR)-based metabolomic approach to elucidate the effect of lercanidipine on neurometabolic alterations in transient model of ischaemic stroke in rats. In the present investigation, male Wistar rats were subjected to middle cerebral artery occlusion (MCAo) for 2h followed by reperfusion using intraluminal filament method. Rats were randomly divided into three groups as vehicle-treated sham control, vehicle-treated MCAo control and lercanidipine-treated MCAo. Vehicle or lercanidipine (0.5mg/kg, i.p.) was administered 120min post-reperfusion. The rat brain cortex tissues were isolated 24h post-MCAo and were investigated by 1 H NMR spectroscopy through perchloric extraction method. A total of 23 metabolites were altered significantly after cerebral ischaemic-reperfusion injury in MCAo control as compared to sham control rats. Lercanidipine significantly reduced the levels of valine, alanine, lactate, acetate and tyrosine, while N-acetylaspartate, glutamate, glutamine, aspartate, creatine/phosphocreatine, choline, glycerophosphorylcholine, taurine, myo-inositol and adenosine di-phosphate were elevated as compared to MCAo control. Present study illustrates effect of lercanidipine on neurometabolic alterations which might be mediated through its antioxidant, anti-inflammatory, vasodilatory and anti-apoptotic property in MCAo model of stroke.

Combined LC-MS and 1H-NMR metabolomic profiling uncovers dietary biomarkers in a cohort of healthy Northern Irish older adults:

AbstractA longstanding issue in the field of nutrition is the potential inaccuracy of methods traditionally used for dietary assessment (i.e. food diaries and food frequency questionnaires). It is possible to overcome the limitations and biases of these techniques by combining them with analytical measurements in human biofluids. Metabolomic technologies are gaining popularity as nutritional tools due to their capacity to measure metabolic responses to external stimuli, such as the ingestion of certain foods. This project performed both LC-MS and 1H-NMR metabolomic profiling on serum samples collected as part of the NICOLA study (Northern Irish Cohort for the Longitudinal Study of Aging) in order to discover novel dietary biomarkers. A dietary validation cohort (NIDAS) was incorporated within NICOLA, involving 45 males and 50 females, aged 50 years and over. Participants provided detailed dietary data (4-day food diary) and blood samples at two time-points, six months apart. Serum samples were processed on two analytical platforms. 1H-NMR spectra were acquired using a Bruker 600 MHz Ascent coupled to a TCI cryoprobe and processed using Bayesil (University of Alberta, Canada). A Waters TQ-S coupled with an Acquity I-class UPLC was used in combination with a targeted commercially available kit (AbsoluteIDQ p180 kit, Biocrates). Mass spectra obtained were processed with MetIDQ and verified using MassLynx (v4.1). Data were tested for normality, and metabolite concentrations were correlated with recorded dietary intake of each food type using SPSS. Additional tests (PCA, PLS-DA, ROC Curves) were performed on MetaboAnalyst 4.0 (University of Alberta, Canada). More than 50 statistically significant (P &lt; 0.05) food-metabolite correlations were detected, 15 of which remained significant after eliminating potential confounding from sex, age and BMI. The strongest correlations were between fruit consumption and acetic acid, and between dairy consumption and certain glycerophospholipids (e.g. LysoPC aa C20:3). Stratifying the cohort by gender yielded further correlations, including PC ae C38:2 (dairy; males), PC aa C34:4 (dairy; females), PC aa C36:4 (dairy; females) and trans-4-Hydroxyproline (meat; males). A number of potential blood-based food biomarkers were detected, many of which are gender-specific, and some are corroborated by previously published studies. However, further validation work is required. For example, biological plausibility needs to be established, and the findings need to be reproduced in other cohorts to demonstrate their applicability in larger and more diverse populations. These results contribute greatly to the ongoing efforts to discover and validate reliable nutritional biomarkers as an objective and unbiased measurement of food intake.

Evaluation of sample preparation protocols for quantitative NMR-based metabolomics.

Quantification of metabolites in biological fluids and tissues by NMR spectroscopy is challenged by the presence of abundant macromolecules and lipoproteins in samples, which give broad signals in the NMR spectra. To improve the quality of NMR spectra the different protocols for protein and lipid removal from the sample are used. This work is aimed at the evaluation of the effectiveness of various methods of purification of blood serum from proteins and lipids for 1H NMR metabolomic profiling. The advantages and limitations of different methods of the sample preparation for NMR-based quantitative metabolomics have been compared, including ultrafiltration, methanol and ethanol extractions with and without additional lipid removal, and methanol-chloroform extraction. The concentrations of 30 abundant metabolites extracted from human blood serum have been measured. It is found that ultrafiltration provides the best lipid removal, but causes significant and inhomogeneous metabolite losses. Ethanol and methanol extractions demonstrate similar performance with the minimal metabolite losses, and are ideal for fluids and tissues with low lipid content. The additional purification of alcohol extracts from lipids allows for the significant improving of NMR spectra, but causes additional metabolite losses. The methanol-chloroform extraction seems to be an optimal method for tissues with the high lipid content, providing a satisfactory lipid removal and low metabolite losses. The ultrafiltration leads to large losses of metabolites (up to 60%) and for this reason is not suitable for quantitative analysis.

Metabolomic profiles differ among unique genotypes of a threatened Caribbean coral

Global threats to reefs require urgent efforts to resolve coral attributes that affect survival in a changing environment. Genetically different individuals of the same coral species are known to exhibit different responses to the same environmental conditions. New information on coral physiology, particularly as it relates to genotype, could aid in unraveling mechanisms that facilitate coral survival in the face of stressors. Metabolomic profiling detects a large subset of metabolites in an organism, and, when linked to metabolic pathways, can provide a snapshot of an organism’s physiological state. Identifying metabolites associated with desirable, genotype-specific traits could improve coral selection for restoration and other interventions. A key step toward this goal is determining whether intraspecific variation in coral metabolite profiles can be detected for species of interest, however little information exists to illustrate such differences. To address this gap, we applied untargeted 1H-NMR and LC-MS metabolomic profiling to three genotypes of the threatened coral Acropora cervicornis. Both methods revealed distinct metabolite “fingerprints” for each genotype examined. A number of metabolites driving separation among genotypes were identified or putatively annotated. Pathway analysis suggested differences in protein synthesis among genotypes. For the first time, these data illustrate intraspecific variation in metabolomic profiles for corals in a common garden. Our results contribute to the growing body of work on coral metabolomics and suggest future work could identify specific links between phenotype and metabolite profile in corals.

NMR-Based Metabolomics Approach To Study the Influence of Different Conditions of Water Irrigation and Greenhouse Ventilation on Zucchini Crops.

This study describes the approach of 1H NMR metabolomic profiling for the differentiation of zucchini produced under different conditions of water irrigation (desalinated seawater -0.397 dS/m, 0.52 €/m3 vs groundwater -2.36 dS/m, 0.29 €/m3) and ventilation (surface area of the vent openings/greenhouse area was 15.0% for one sector and 9.8% for the other). Overall, 72 extracts of zucchini ( Cucubirta pepo L. cv Victoria) under four different conditions were regularly analyzed during the spring-summer cycle from April to July 2017. We have found that zucchini plants irrigated with desalinated seawater increased the zucchini production yield, presented fruits with higher concentration of glucose, fructose, and vitamin B3, and displayed an increased antioxidant activity. On the contrary, plant groundwater irrigation produced the increment of sucrose level that could rise the sweetness perception of the fruits. Finally, the ventilation variable produced a higher concentration of trigonelline, histidine, and phenylalanine but only on those zucchinis irrigated with groundwater.

1H-NMR metabolomic profiling reveals a distinct metabolic recovery response in shoots and roots of temporarily drought-stressed sugar beets.

Yield formation in regions with intermittent drought periods depends on the plant’s ability to recover after cessation of the stress. The present work assessed differences in metabolic recovery of leaves and roots of drought-stressed sugar beets with high temporal resolution. Plants were subjected to drought for 13 days, and rewatered for 12 days. At one to two-day intervals, plant material was harvested for untargeted 1H-NMR metabolomic profiling, targeted analyses of hexose-phosphates, starch, amino acids, nitrate and proteins, and physiological measurements including relative water content, osmotic potential, electrolyte leakage and malondialdehyde concentrations. Drought triggered changes in primary metabolism, especially increases in amino acids in both organs, but leaves and roots responded with different dynamics to rewatering. After a transient normalization of most metabolites within 8 days, a second accumulation of amino acids in leaves might indicate a stress imprint beneficial in upcoming drought events. Repair mechanisms seemed important during initial recovery and occurred at the expense of growth for at least 12 days. These results indicate that organ specific metabolic recovery responses might be related to distinct functions and concomitant disparate stress levels in above- and belowground organs. With respect to metabolism, recovery was not simply a reversal of the stress responses.

1H NMR-based metabolomic profiling for identification of metabolites in Capsicum annuum cv. mirasol infected by beet mild curly top virus (BMCTV)

Beet mild curly top virus (BMCTV) is associated with an outbreak of curly top in chili pepper, tomato and other Solanaceae species, which can cause severe crop losses. The aim of this work was to obtain the 1H NMR metabolomic profiling of both healthy chili peppers (cv. mirasol) and infected chili peppers with BMCTV in order to find chemical markers associated to the infection process. Significant differences were found between the two groups, according to principal component analysis and orthogonal projections to latent structure discriminant analysis. Compared to the asymptomatic peppers, the symptomatic fruits had higher relative abundance of fructose, isoleucine, histidine, phenylalanine and tryptophan. Contrarily, the asymptomatic samples showed greater amounts of malonate and isobutyrate. These results suggest that in diseased chili peppers there are metabolic changes related to the viral acquisition of energy for replication and capsid assembly. This is the first study describing the chemical profiling of a polar extract obtained from Capsicum annuum infected by BMCTV under open field conditions.

1H NMR-based metabolomic fingerprinting to determine metabolite levels in serrano peppers (Capsicum annum L.) grown in two different regions

Chili pepper (Capsicum annuum) is the most important and emblematic condiment in Mexican food. Serrano pepper is a variety of C. annuum that is traditionally cultivated in Mexico and commercialized in local markets. The aim of this study was to describe the 1H NMR metabolomic profiling of the aqueous phase of serrano peppers harvested from two distinct regions, in the states of Veracruz and Oaxaca, Mexico. According to the current results, aspartate citrate, lactate, leucine and sucrose were found at higher amount in the serrano peppers from Veracruz. On the other hand, acetate, formate, fumarate, malonate, phosphocholine, pyruvate and succinate showed the highest abundance in this product from Oaxaca. These are the main metabolites that distinguish one group from the other. The spectrometric method reported presently is characterized by great simplicity, robustness and reproducibility. Thus, this technique can be used for establishing reliable metabolomic fingerprints of serrano peppers grown under different environmental conditions.

1H NMR metabolomic signatures related to giant cell tumor of the bone

1H NMR metabolomic profiling for giant cell tumor of the bone.

(1)H-NMR urinary metabolomic profiling for diagnosis of gastric cancer.

Background:Metabolomics has shown promise in gastric cancer (GC) detection. This research sought to identify whether GC has a unique urinary metabolomic profile compared with benign gastric disease (BN) and healthy (HE) patients.Methods:Urine from 43 GC, 40 BN, and 40 matched HE patients was analysed using 1H nuclear magnetic resonance (1H-NMR) spectroscopy, generating 77 reproducible metabolites (QC-RSD <25%). Univariate and multivariate (MVA) statistics were employed. A parsimonious biomarker profile of GC vs HE was investigated using LASSO regularised logistic regression (LASSO-LR). Model performance was assessed using Receiver Operating Characteristic (ROC) curves.Results:GC displayed a clear discriminatory biomarker profile; the BN profile overlapped with GC and HE. LASSO-LR identified three discriminatory metabolites: 2-hydroxyisobutyrate, 3-indoxylsulfate, and alanine, which produced a discriminatory model with an area under the ROC of 0.95.Conclusions:GC patients have a distinct urinary metabolite profile. This study shows clinical potential for metabolic profiling for early GC diagnosis.

1H NMR metabolomic profiling of the blue crab (Callinectes sapidus) from the Adriatic Sea (SE Italy): A comparison with warty crab (Eriphia verrucosa), and edible crab (Cancer pagurus)

The metabolomic profile of blue crab (Callinectes sapidus) captured in the Acquatina lagoon (SE Italy) was compared to an autochthonous (Eriphia verrucosa) and to a commercial crab species (Cancer pagurus). Both lipid and aqueous extracts of raw claw muscle were analyzed by 1H NMR spectroscopy and MVA (multivariate data analysis). Aqueous extracts were characterized by a higher inter-specific discriminating power compared to lipid fractions. Specifically, higher levels of glutamate, alanine and glycine characterized the aqueous extract of C. sapidus, while homarine, lactate, betaine and taurine characterized E. verrucosa and C. pagurus. On the other hand, only the signals of monounsaturated fatty acids distinguished the lipid profiles of the three crab species. These results support the commercial exploitation and the integration of the blue crab in human diet of European countries as an healthy and valuable seafood.

Metabolomic NMR Fingerprinting to Identify and Predict Survival of Patients with Metastatic Colorectal Cancer

Earlier detection of patients with metastatic colorectal cancer (mCRC) might improve their treatment and survival outcomes. In this study, we used proton nuclear magnetic resonance ((1)H-NMR) to profile the serum metabolome in patients with mCRC and determine whether a disease signature may exist that is strong enough to predict overall survival (OS). In 153 patients with mCRC and 139 healthy subjects from three Danish hospitals, we profiled two independent sets of serum samples in a prospective phase II study. In the training set, (1)H-NMR metabolomic profiling could discriminate patients with mCRC from healthy subjects with a cross-validated accuracy of 100%. In the validation set, 96.7% of subjects were correctly classified. Patients from the training set with maximally divergent OS were chosen to construct an OS predictor. After validation, patients predicted to have short OS had significantly reduced survival (HR, 3.4; 95% confidence interval, 2.06-5.50; P = 1.33 × 10(-6)). A number of metabolites concurred with the (1)H-NMR fingerprint of mCRC, offering insights into mCRC metabolic pathways. Our findings establish that (1)H-NMR profiling of patient serum can provide a strong metabolomic signature of mCRC and that analysis of this signature may offer an independent tool to predict OS.

Metabolomic profiling of rapid cold hardening and cold shock in Drosophila melanogaster

A short exposure to a mild cold stress is sufficient to increase cold tolerance in many insects. This phenomenon, termed rapid cold hardening (RCH) expands the thermal interval that can be exploited by the insect. To investigate the possible role of altered metabolite levels during RCH, the present study used untargeted 1H NMR metabolomic profiling to examine the metabolomic response in Drosophila melanogaster during the 72 h following RCH and cold shock treatment. These findings are discussed in relation to the costs and benefits of RCH that are measured in terms of survival and reproductive output. Cold shock caused a persistent disturbance of the metabolite profile that correlated well with a delayed onset of cold shock mortality. The disruption of metabolite homeostasis was smaller following RCH, where control levels were fully recovered after 72 h. RCH improved both survival and reproductive output after a subsequent cold shock but the RCH treatment alone was associated with costs in terms of reduced survival and reproductive output. The most pronounced changes following the RCH treatment were elevated levels of glucose and trehalose. Although, it is difficult to discern if a change in a specific metabolite is linked to physiological processes of adaptive, neutral or detrimental nature we observed that the onset and magnitude of the increased sugar levels correlated tightly with the improved chill tolerance following RCH. These findings suggest a putative role of cryoprotectants during RCH which are discussed in the light of the existing literature on the mechanistic background of RCH.