1H High-resolution Magic Angle Spinning Research Articles

μHigh resolution-magic-angle spinning NMR spectroscopy for metabolic phenotyping of Caenorhabditis elegans.

Analysis of model organisms, such as the submillimeter-size Caenorhabditis elegans, plays a central role in understanding biological functions across species and in characterizing phenotypes associated with genetic mutations. In recent years, metabolic phenotyping studies of C. elegans based on (1)H high-resolution magic-angle spinning (HR-MAS) nuclear magnetic resonance (NMR) spectroscopy have relied on the observation of large populations of nematodes, requiring labor-intensive sample preparation that considerably limits high-throughput characterization of C. elegans. In this work, we open new platforms for metabolic phenotyping of C. elegans mutants. We determine rich metabolic profiles (31 metabolites identified) from samples of 12 individuals using a (1)H NMR microprobe featuring high-resolution magic-angle coil spinning (HR-MACS), a simple conversion of a standard HR-MAS probe to μHR-MAS. In addition, we characterize the metabolic variations between two different strains of C. elegans (wild-type vs slcf-1 mutant). We also acquire a NMR spectrum of a single C. elegans worm at 23.5 T. This study represents the first example of a metabolomic investigation carried out on a small number of submillimeter-size organisms, demonstrating the potential of NMR microtechnologies for metabolomics screening of small model organisms.

Metabolomics analysis uncovers that dietary restriction buffers metabolic changes associated with aging in Caenorhabditis elegans.

Dietary restriction (DR) is one of the most universal means of extending lifespan. Yet, whether and how DR specifically affects the metabolic changes associated with aging is essentially unknown. Here, we present a comprehensive and unbiased picture of the metabolic variations that take place with age at the whole organism level in Caenorhabditis elegans by using 1H high-resolution magic-angle spinning (HR-MAS) nuclear magnetic resonance (NMR) analysis of intact worms. We investigate metabolic variations potentially important for lifespan regulation by comparing the metabolic fingerprint of two previously described genetic models of DR, the long-lived eat-2(ad465) and slcf-1(tm2258) worms, as single mutants or in combination with a genetic suppressor of their lifespan phenotype. Our analysis shows that significant changes in metabolite profiles precede the major physiological decline that accompanies aging and that DR protects from some of those metabolic changes. More specifically, low phosphocholine (PCho) correlates with high life expectancy. A mutation in the tumor suppressor gene PTEN/DAF-18, which suppresses the beneficial effects of DR in both C. elegans and mammals, increases both PCho level and choline kinase expression. Furthermore, we show that choline kinase function in the intestine can regulate lifespan. This study highlights the relevance of NMR metabolomic approaches for identifying potential biomarkers of aging.

Dose-dependent influence of short-term intermittent ethanol intoxication on cerebral neurochemical changes in rats detected by ex vivo proton nuclear magnetic resonance spectroscopy

The aim of this study was to quantitatively assess the effects of short-term intermittent ethanol intoxication on cerebral metabolite changes among sham controls (CNTL), low-dose ethanol (LDE)-exposed, and high-dose ethanol (HDE)-exposed rats, which were determined with ex vivo high-resolution spectra. Eight-week-old male Wistar rats were divided into three groups. Twenty rats in the LDE (n=10) and the HDE (n=10) groups received ethanol doses of 1.5 and 2.5g/kg, respectively, through oral gavage every 8h for 4days. At the end of the 4-day intermittent ethanol exposure, one-dimensional ex vivo 500-MHz 1H nuclear magnetic resonance spectra were acquired from 30 samples of the frontal cortex region (from the three groups). Normalized total N-acetylaspartate (tNAA: NAA+NAAG [N-acetylaspartyl-glutamate]), GABA, and glutathione (GSH) levels were significantly lower in the frontal cortex of the HDE-exposed rats than that of the LDE-exposed rats. Moreover, compared to the CNTL group, the LDE rats exhibited significantly higher normalized GABA levels. The six pairs of normalized metabolite levels were positively (+) or negatively (−) correlated in the rat frontal cortex as follows: tNAA and GABA (+), tNAA and aspartate (Asp) (+), myo-Inositol (mIns) and Asp (−), mIns and alanine (+), mIns and taurine (+), and mIns and tNAA (−). Our results suggested that short-term intermittent ethanol intoxication might result in neuronal degeneration and dysfunction, changes in the rate of GABA synthesis, and oxidative stress in the rat frontal cortex. Our ex vivo 1H high-resolution magic angle spinning nuclear magnetic resonance spectroscopy results suggested some novel metabolic markers for the dose-dependent influence of short-term intermittent ethanol intoxication in the frontal cortex.

Analysis of soil organic matter at the solid–water interface by nuclear magnetic resonance spectroscopy

Environmental context Structural and conformational information on organic matter–clay complexes and whole soils was obtained using different NMR methods. The results show that organic matter interactions with clay mineral surfaces determine the accessibility of specific organic matter components at the soil–water interface. This physical conformation may also play a role in soil biogeochemical processes and binding to pollutants in terrestrial environments. Abstract Organic matter (OM)–mineral interactions play an important role in OM preservation, global carbon cycling and contaminant transport. Studies have indicated that preferential sorption of OM is dependent on mineral type and solution conditions. In this study, 1H high resolution–magic angle spinning nuclear magnetic resonance (HR-MAS NMR) spectroscopy was employed to examine OM chemistry in organo-clay complexes. Dissolved OM from a forest soil, Leonardite humic acid and Peat humic acid were sorbed to Ca2+ enriched kaolinite and montmorillonite. As observed using 1H HR-MAS NMR spectroscopy, kaolinite sorbed mainly long-chain aliphatic compounds such as those from plant cuticles whereas montmorillonite sorbed a mixture of aliphatic components and proteins. These results show the preferential sorption of specific dissolved OM components on clay surfaces. This was tested further using solid-state 13C and 1H HR-MAS NMR analysis of whole soils containing kaolinite and montmorillonite as well as a Peat soil for contrast. The species present at the soil–water interface were mainly aliphatic components, carbohydrates and amino acids. Aromatic constituents were present in the soils (observed by solid-state 13C NMR and by 1H HR-MAS NMR spectroscopy when a more penetrating solvent was used) which signifies that these compounds likely exist in more hydrophobic domains that are buried and surface inaccessible. This study highlights the important role of OM interactions with clay minerals in the preservation of OM in soils and suggests that OM–OM associations may also play a role in the protection of specific OM components in soil.

A New Specific Succinate-Glutamate Metabolomic Hallmark in Sdhx-Related Paragangliomas

Paragangliomas (PGLs) are frequently associated with germline mutations in genes involved in energy metabolism. The purpose of the present study was to assess whether the tumor metabolomic profile of patients with hereditary and apparently sporadic PGLs enables the distinction of different subtypes of tumors. Twenty-eight unrelated patients with a histological diagnosis of PGLs were included in the present study. Twelve had germline mutations in SDHx genes (5 SDHB, 7 SDHD), 6 VHL, and 10 were apparently sporadic. Intact tumor samples from these patients (one per patient) were evaluated with 1H high-resolution magic angle spinning (HRMAS) NMR spectroscopy. SDHx-related tumors were characterized by an increase in succinate levels in comparison to other tumor subtypes (p = 0.0001 vs VHL and p = 0.000003 vs apparently sporadic). Furthermore, we found significantly lower values of glutamate in SDHx-related tumors compared to other subtypes (p = 0.0007 vs VHL and p = 0.003 vs apparently sporadic). Moreover, SDHx-tumors also exhibited lower values of ATP/ADP/AMP (p = 0.01) compared to VHL. VHL tumors were found to have the highest values of glutathione (GSH) compared to other tumors. Based on 4 metabolites (succinate, glutamate, GSH, and ATP/ADP/AMP), tumors were accurately distinguished from the other ones on both 3- and 2-class PLS-DA models. The present study shows that HRMAS NMR spectroscopy is a very promising method for investigating the metabolomic profile of various PGLs. The present data suggest the existence of a specific succinate-glutamate hallmark of SDHx PGLs. The relevance of such a metabolomic hallmark is expected to be very useful in designing novel treatment options as well as improving the diagnosis and follow-up of these tumors, including metastatic ones.

Long-term calorie restriction has minimal impact on brain metabolite and fatty acid profiles in aged rats on a Western-style diet

The effect of long-term calorie restriction (CR) on metabolites, fatty acid profiles and energy substrate transporter expression in the brain was assessed in aged rats. Three groups of male Sprague–Dawley rats were studied: (i) a 2month old ad libitum-fed (2AL group), (ii) a 19month old ad libitum-fed (19AL group), and (iii) a 19month old group subjected to 40% CR from the age of 7.5 to 19months (19CR group). The diet contained high sucrose and low n-3 polyunsaturated fatty acids (PUFA) so as to imitate a Western-style diet. High resolution magic angle spinning-1H NMR showed an effect of aging on brain cortex metabolites compared to 2AL rats, the largest differences being for myo-inositol (+251% and +181%), lactate (+203% and +188%), β-hydroxybutyrate (+176% and +618%) and choline (+148% and +120%), in 19AL and 19 CR rats, respectively. However, brain metabolites did not differ between the 19AL and 19CR groups. Cortex fatty acid profiles showed that n-3 PUFA were 35–47% lower but monounsaturated fatty acids were 40–52% higher in 19AL and 19CR rats compared to 2AL rats. Brain microvessel glucose transporter (GLUT1) was 68% higher in 19AL rats than in 2AL rats, while the monocarboxylate transporter, MCT1, was 61% lower in 19CR rats compared to 19AL rats. We conclude that on a high-sucrose, low n-3 PUFA diet, the brain of aged AL rats had higher metabolites and microvessel GLUT1 expression compared to 2AL rats. However, long-term CR in aged rats did not markedly change brain metabolite or fatty acid profile, but did reduce brain microvessel MCT1 expression.

Dissolution and Crystallization of Polyamides in Superheated Water and Concentrated Ionic Solutions

The dissolution and recrystallization of Polyamide 46 (PA46) from “superheated” (i) water and (ii) concentrated ionic solutions of strong solubilizing mono- and divalent Hofmeister ions are studied, utilizing in situ high-resolution magic-angle spinning (HR-MAS) nuclear magnetic resonance (NMR) supported by wide-angle X-ray diffraction (WAXD), attenuated total reflectance–Fourier transform infrared spectroscopy (ATR-FTIR), and gel permeation chromatography (GPC). The samples are sealed in glass capillaries, and employing variable-temperature 1H HR-MAS NMR spectroscopy, the dissolution process of PA46 as a function of temperature and pressure can be followed in situ. The purpose of such a study is to obtain molecular insight into the dissolution process of these hydrogen-bonded synthetic polymers in the different aqueous solutions. In pure water, at temperatures close to the dissolution of PA46, two distinct 1H resonances from water are observed. These resonances are associated with water molecules in the vicinity of PA46 and water in the bulk state. On further heating, the signal from water molecules in the vicinity of PA46 dominates. This sudden change in the environment suggests that water molecules, which have escaped the dense hydrogen-bonded network of bulk water, can diffuse into the structure of PA46, triggering dissolution of the polymer. This happens at a temperature that is more than 100 °C below the melting temperature of the polymer, notably without hydrolysis as verified by GPC performed prior to and after the dissolution experiments. On cooling, recrystallization of PA46 from aqueous solution is observed where water molecules are incorporated in the crystal structure. In the presence of salts, such as LiI and Cal2, weakening of the hydrogen-bonding network of the water molecules occurs. However, above room temperature, independent of the choice of salt, depopulation of the hydrogen bonding between the water molecules occurs, observed as a decrease in the 1H chemical shift value. The reduced hydrogen bonding in the presence of ions facilitates the dissolution of PA46 at much lower temperatures compared to pure water and ultimately results in the complete suppression of crystallization from solution even at room temperature. Depending on the valency of the cation a more mobile or frozen amorphous state of PA46 is obtained at low temperatures as verified by 13C HR-MAS NMR, ATR-FTIR, and WAXD.

Live-cell high resolution magic angle spinning magnetic resonance spectroscopy for in vivo analysis of Pseudomonas aeruginosa metabolomics

Pseudomonas aeruginosa (PA) is a pathogenic gram-negative bacterium that is widespread in nature, inhabiting soil, water, plants and animals. PA is a prevalent cause of deleterious human infections, particularly in patients whose host defense mechanisms have been compromised. Metabolomics is an important tool used to study host-pathogen interactions and to identify novel therapeutic targets and corresponding compounds. The aim of the present study was to report the metabolic profile of live PA bacteria using in vivo high-resolution magic angle spinning (HRMAS) nuclear magnetic resonance spectroscopy (NMR), in combination with 1- and 2-dimensional HRMAS NMR. This methodology provides a new and powerful technique to rapidly interrogate the metabolome of intact bacterial cells and has several advantages over traditional techniques that identify metabolome components from disrupted cells. Furthermore, application of multidimensional HRMAS NMR, in combination with the novel technique total through-Bond correlation Spectroscopy (TOBSY), is a promising approach that may be used to obtain in vivo metabolomics information from intact live bacterial cells and can mediate such analyses in a short period of time. Moreover, HRMAS 1H NMR enables the investigation of the associations between metabolites and cell processes. In the present study, we detected and quantified several informative metabolic molecules in live PA cells, including N-acetyl, betaine, citrulline, alanine and glycine, which are important in peptidoglycan synthesis. The results provided a complete metabolic profile of PA for future studies of PA clinical isolates and mutants. In addition, this in vivo NMR biomedical approach might have clinical utility and should prove useful in gene function validation, the study of pathogenetic mechanisms, the classification of microbial strains into functional/clinical groups, the testing of anti-bacterial agents and the determination of metabolic profiles of bacterial mutants.

Assessment of a 1H high-resolution magic angle spinning NMR spectroscopy procedure for free sugars quantification in intact plant tissue

In most plants, sucrose is the primary product of photosynthesis, the transport form of assimilated carbon, and also one of the main factors determining sweetness in fresh fruits. Traditional methods for sugar quantification (mainly sucrose, glucose and fructose) require obtaining crude plant extracts, which sometimes involve substantial sample manipulation, making the process time-consuming and increasing the risk of sample degradation. Here, we describe and validate a fast method to determine sugar content in intact plant tissue by using high-resolution magic angle spinning nuclear magnetic resonance spectroscopy (HR-MAS NMR). The HR-MAS NMR method was used for quantifying sucrose, glucose and fructose in mesocarp tissues from melon fruits (Cucumis melo var. reticulatus and Cucumis melo var. cantalupensis). The resulting sugar content varied among individual melons, ranging from 1.4 to 7.3g of sucrose, 0.4-2.5g of glucose; and 0.73-2.83g of fructose (values per 100g fw). These values were in agreement with those described in the literature for melon fruit tissue, and no significant differences were found when comparing them with those obtained using the traditional, enzymatic procedure, on melon tissue extracts. The HR-MAS NMR method offers a fast (usually <30min) and sensitive method for sugar quantification in intact plant tissues, it requires a small amount of tissue (typically 50mg fw) and avoids the interferences and risks associated with obtaining plant extracts. Furthermore, this method might also allow the quantification of additional metabolites detectable in the plant tissue NMR spectrum.

1 H-HRMAS NMR study of cold smoked Atlantic salmon (Salmo salar ) treated with E-beam

(1)H High resolution magic angle spinning (HRMAS) NMR spectroscopic data in combination with principal components analysis and analysis of variance were used to differentiate between irradiated and non-irradiated cold-smoked Atlantic salmon (Salmo salar). NMR profiling was obtained, with a spectral acquisition time of less than 8 min, from a small sample size of intact white salmon muscle, by non-destructive analysis that includes a very simple and rapid sample preparation step. Results obtained enable the use of creatine, trimethylamine oxide and the sum of phosphorylcholine and glycerophosphorylcholine as diagnostic compounds to detect irradiation treatment. This study shows the potential of (1)H-HRMAS to be a rapid method for investigating compositional changes due to food processing as well as to confirm the presence or absence of some bioactive compounds in irradiated samples.

Feasibility of 1H-High Resolution-Magic Angle Spinning NMR Spectroscopy in the Analysis of Viscous Cosmetic and Pharmaceutical Formulations

The feasibility of (1)H-High Resolution-Magic Angle Spinning (HR-MAS) nuclear magnetic resonance (NMR) spectroscopy for the direct analysis of viscous cosmetic and pharmaceutical formulations such as creams, gels, and pastes is presented. Three examples are described: (i) the detection of chitosan in toothpaste, (ii) the analysis of dexamethasone acetate (DMA) in a cream, and (iii) the analysis of the local anesthetics, lidocaine and prilocaine, in a gel and a cream. All active components could be directly detected in their original commercial formulations without the need for laborious sample preparation steps. In addition, the possibility for HR-MAS-based quantifications and the analysis of dynamic properties of active components in different formulations applying HR-MAS diffusion-ordered NMR spectroscopy are shown.

High‐resolution magic angle spinning 1H MRS in prostate cancer

Prostate cancer (PCa) is the most frequently diagnosed malignancy in men worldwide, largely as a result of the increased use of the annual serum prostate‐specific antigen (PSA) screening test for detection. PSA screening has saved lives, but it has also resulted in the overtreatment of many patients with PCa because of a limited ability to accurately localize and characterize PCa lesions through imaging. High‐resolution magic angle spinning (HRMAS) 1H MRS has proven to be a strong potential clinical tool for PCa diagnosis and prognosis. The HRMAS technique allows valuable metabolic information to be obtained from ex vivo intact tissue samples and also enables the performance of histopathology on the same tissue specimens. Studies have found that the quantification of individual metabolite levels and metabolite ratios, as well as metabolomic profiles, shows strong potential to improve accuracy in PCa detection, diagnosis and monitoring. Ex vivo HRMAS is also a valuable tool for the interpretation of in vivo results, including the localization of tumors, and thus has the potential to improve in vivo diagnostic tests used in the clinic. Here, we primarily review publications of HRMAS 1H MRS and its use for the study of intact human prostate tissue. Copyright © 2013 John Wiley &amp; Sons, Ltd.

Study of metabonomic profiles of human esophageal carcinoma by use of high-resolution magic-angle spinning 1H NMR spectroscopy and multivariate data analysis

Esophageal carcinoma (EC) is one of the most common malignant tumors. EC survival has remained disappointingly low because of the high malignancy of esophageal cancer and the lack of obvious clinical symptoms at an early stage. Early diagnosis is often difficult because the small tumor nodules are frequently missed. Metabonomics based on high-resolution magic-angle spinning (HRMAS) NMR has been popular for tumor detection because it is highly sensitive, provides rich biochemical information and requires no sample pretreatment. (1)H HRMAS spectra of non-involved adjacent esophageal tissues and of well differentiated and moderately differentiated esophageal carcinoma tumors were recorded and analyzed by use of multivariate and statistical analysis techniques. Moderately differentiated EC tumors were found to have increased total choline, alanine, and glutamate and reduced creatine, myo-inositol, and taurine compared with non-involved adjacent tissues. Moreover, clear differences between the metabonomic profiles of EC tissues enabled tumor differentiation. Furthermore, the integral Gly/MI ratio for samples of different tissue types were statistically significantly different; this was sufficient both for distinguishing non-involved tissues from esophageal carcinoma and for classification of well differentiated and moderately differentiated EC tumors.

Nano-cellulose 3D-networks as controlled-release drug carriers.

Bacterial cellulose (BC) membranes are used as the carrier for berberine hydrochloride and berberine sulphate to produce a new controlled release system. Release studies and transdermal experiments were carried out in vitro. Carrier BC can significantly extend the drug release time, in contrast to existing commercial carriers. Freeze-dried BC membranes 10 mm thick were optimized for drug delivery. The lowest release rate was in simulated gastric fluid (SGF) or in H2SO4 solution, the highest in simulated intestinal fluid (SIF) and an intermediate rate was found in alkaline conditions. The release curves closely followed the Ritger-Peppas model with free diffusion the most prominent mechanism. Scanning electron microscopy (SEM) analysis demonstrated that BC fibers were swollen in acid and base conditions. 1H high-resolution magic angle spinning nuclear magnetic resonance (1H HRMAS NMR) diffusion-ordered spectroscopy (DOSY) analysis showed that there was an interaction between the drugs and BC. The structure of BC, the media and the solubility of the drug all influenced the sustained-release behavior. The results from the release studies, the electron micrographs, and the transdermal experiments were in good agreement.

Exploring High-resolution Magic Angle Spinning (HR-MAS) NMR Spectroscopy for Metabonomic Analysis of Apples

Classical liquid-state high-resolution (HR) NMR spectroscopy has proved a powerful tool in the metabonomic analysis of liquid food samples like fruit juices. In this paper the application of (1)H high-resolution magic angle spinning (HR-MAS) NMR spectroscopy to apple tissue is presented probing its potential for metabonomic studies. The (1)H HR-MAS NMR spectra are discussed in terms of the chemical composition of apple tissue and compared to liquid-state NMR spectra of apple juice. Differences indicate that specific metabolic changes are induced by juice preparation. The feasibility of HR-MAS NMR-based multivariate analysis is demonstrated by a study distinguishing three different apple cultivars by principal component analysis (PCA). Preliminary results are shown from subsequent studies comparing three different cultivation methods by means of PCA and partial least squares discriminant analysis (PLS-DA) of the HR-MAS NMR data. The compounds responsible for discriminating organically grown apples are discussed. Finally, an outlook of our ongoing work is given including a longitudinal study on apples.

Garganica kid goat meat: Physico-chemical characterization and nutritional impacts

A detailed physico-chemical characterization of the Garganica kid goat meat was carried out on samples (quadriceps femoris muscles) coming from different farms located on the Promontory of Gargano, in Apulia (southern Italy) by determining conventional parameters (moisture, ashes, fat, and protein content), stable isotope ratios (15N/14N and 13C/12C), major (Ca, Mg, Na, and K), and trace metals (Zn, Mn, Cu, Fe, and Cr). Moreover, information on a large number of metabolites was obtained by means of 1H High-Resolution Magic-Angle Spinning Nuclear Magnetic Resonance (1H HR-MAS NMR) spectroscopy. The most interesting findings of this work, for their impact on human nutrition and health, were represented by a considerable protein content (18.9%) and a low fat content (2.98%), characterized by a remarkable average percentage of desirable fatty acids (62.76%) and a good (C18+C18:1)/C16 average ratio (2.30), that make the Garganica kid goat meat a highly appreciable food product.

Identification of Multiple Diffusion Rates in Mixed Solvent Anion Exchange Membranes Using High Resolution MAS NMR

High resolution magic angle spinning (HRMAS) 1H NMR in combination with 2D exchange NOESY and pulsed field gradient (PFG) NMR diffusion experiments have been used to characterize 1 N methanol swollen polymer anion exchange membranes (AEM) presently being developed for alkaline fuel cells. Standard static 1H NMR experiments on these materials have proven unsuccessful due to severe signal broadening. New experimental methods for increased resolution are needed to determine distinct solvent environments and transport properties. Using HRMAS NMR, resonances from water and methanol in both a free (bulk-like) environment and membrane-associated environment within the AEM were observed. 1H HRMAS PFG NMR experiments identified different molecular diffusion environments in the solvent, while 1H 2D NOESY exchange NMR experiments confirmed spatial contacts between membrane-associated species and the membrane. These results demonstrate that 1H HRMAS is an ideal technique for the characterization of individual environments and diffusion rates in polymer membranes with mixed solvent systems.

Cross-Link Density Estimation of PDMS Networks with Precise Consideration of Networks Defects

Two series of randomly cross-linked poly(dimethylsiloxane) (PDMS) networks with systematically varying and well-defined dangling chain and defect content were investigated by 1H double-quantum low-field NMR (DQ NMR), equilibrium swelling, and high-resolution magic-angle spinning NMR (MAS NMR) experiments in order to obtain information about the absolute molecular weight of the polymer chains between two cross-links, Mc, and its distribution. A comparison of the experimental results from swelling and DQ NMR experiments with the results of Miller–Macosko calculations, based upon the reaction turnover determined by MAS NMR, clarifies the importance of a precise estimation of the defect fraction of the polymer network for a reliable determination of Mc. In order to properly account for network defects in the evaluation of equilibrium swelling as well as DQ NMR experiments, we use the elastically effective polymer volume fraction ϕp,el (that can be obtained from DQ NMR or simple Hahn echo experiments on sample...

The assessment of the quality of the graft in an animal model for lung transplantation using the metabolomics 1H high‐resolution magic angle spinning NMR spectroscopy

Standards are needed to control the quality of the lungs from nonheart-beating donors as potential grafts. This was here assessed using the metabolomics 1H high-resolution magic angle spinning NMR spectroscopy. Selective perfusion of the porcine bilung block was set up 30 min after cardiac arrest with cold Perfadex®. Lung alterations were analyzed at 3, 6, and 8 h of cold ischemia as compared to baseline and to nonperfused lung. Metabolomics analysis of lung biopsies allowed identification of 35 metabolites. Levels of the majority of the metabolites increased over time at 4°C without perfusion, indicating cellular degradation, whereas levels of glutathione decreased. When lung was perfused at 4°C, levels of the majority of the metabolites remained stable, including levels of glutathione. Levels of uracil by contrast showed a reverse profile, as its signal increased over time in the absence of perfusion while being totally absent in perfused samples. Our results showed glutathione and uracil as potential biomarkers for the quality of the lung. The metabolomics 1H high-resolution magic angle spinning NMR spectroscopy can be efficiently applied for the assessment of the quality of the lung as an original technique characterized by a rapid assessment of intact biopsy samples without extraction and can be implemented in hospital environment.

Optimization of metabolite basis sets prior to quantitation in magnetic resonance spectroscopy: an approach based on quantum mechanics

High-resolution magic angle spinning (HRMAS) nuclear magnetic resonance (NMR) is playing an increasingly important role for diagnosis. This technique enables setting up metabolite profiles of ex vivo pathological and healthy tissue. The need to monitor diseases and pharmaceutical follow-up requires an automatic quantitation of HRMAS 1H signals. However, for several metabolites, the values of chemical shifts of proton groups may slightly differ according to the micro-environment in the tissue or cells, in particular to its pH. This hampers the accurate estimation of the metabolite concentrations mainly when using quantitation algorithms based on a metabolite basis set: the metabolite fingerprints are not correct anymore. In this work, we propose an accurate method coupling quantum mechanical simulations and quantitation algorithms to handle basis-set changes. The proposed algorithm automatically corrects mismatches between the signals of the simulated basis set and the signal under analysis by maximizing the normalized cross-correlation between the mentioned signals. Optimized chemical shift values of the metabolites are obtained. This method, QM-QUEST, provides more robust fitting while limiting user involvement and respects the correct fingerprints of metabolites. Its efficiency is demonstrated by accurately quantitating 33 signals from tissue samples of human brains with oligodendroglioma, obtained at 11.7 tesla. The corresponding chemical shift changes of several metabolites within the series are also analyzed.