Auswahl der wissenschaftlichen Literatur zum Thema „1H NMR quantification“

High-throughput metabolomics can be used to optimize cell growth for enhanced production or for monitoring cell health in bioreactors. It has applications in cell and gene therapies, vaccines, biologics, and bioprocessing. NMR metabolomics is a method that allows for fast and reliable experimentation, requires only minimal sample preparation, and can be set up to take online measurements of cell media for bioreactor monitoring. This type of application requires a fully automated metabolite quantification method that can be linked with high-throughput measurements. In this review, we discuss the quantifier requirements in this type of application, the existing methods for NMR metabolomics quantification, and the performance of three existing quantifiers in the context of NMR metabolomics for bioreactor monitoring.

A wheat field was sprayed with a dosage of 1.1 kg a.i./ha Roundup PowerMax 10 days before harvest. The 1H Nuclear Magnetic Resonance (NMR) spectroscopy was used for the detection and quantification of the glyphosate (GLYP) in dried wheat spikelets, leaves, and stems. The quantification was done by the integration of the CH2-P groups doublet at 3.00 ppm with good linearity. The GLYP content varied between different samples and parts of the plant. On average, the largest content of herbicide was found in leaves (20.0 mg/kg), followed by stems (6.4 mg/kg) and spikelets (6.3 mg/kg). Our study shows that the 1H-NMR spectroscopy can be a rapid and reliable tool for GLYP detection and quantification in the field studies.

Echo-detected 1H NMR spectra of water molecules hyperpolarized by dissolution-dynamic nuclear polarization alleviates deleterious dead time induced lineshape distortions and enables accurate nuclear spin polarization quantification.

The authors developed four variants of the qNMR technique (1H or 13C nucleus, DMSO-d6 or CDCl3 solvent) for identification and quantification by NMR of 22R and 22S epimers in budesonide active pharmaceutical ingredient and budesonide drugs (sprays, capsules, tablets). The choice of the qNMR technique version depends on the drug excipients. The correlation of 1H and 13C spectra signals to molecules of different budesonide epimers was carried out on the basis of a comprehensive analysis of experimental spectral NMR data (1H-1H gCOSY, 1H-13C gHSQC, 1H-13C gHMBC, 1H-1H ROESY). This technique makes it possible to identify budesonide epimers and determine their weight ratio directly, without constructing a calibration curve and using any standards. The results of measuring the 22S epimer content by qNMR are comparable with the results of measurements using the reference HPLC method.

Magnesium oxide (MgO) can convert to different magnesium-containing compounds depending on exposure and environmental conditions. Many MgO-based phases contain hydrated species allowing 1H-nuclear magnetic resonance (NMR) spectroscopy to be used in the characterization and quantification of proton-containing phases; however, surprisingly limited examples have been reported. Here, 1H-magic angle spinning (MAS) NMR spectra of select Mg-based minerals are presented and assigned. These experimental results are combined with computational NMR density functional theory (DFT) periodic calculations to calibrate the predicted chemical shielding results. This correlation is then used to predict the NMR shielding for a series of different MgO hydroxide, magnesium chloride hydrate, magnesium perchlorate, and magnesium cement compounds to aid in the future assignment of 1H-NMR spectra for complex Mg phases.

Environmental metabolomics is a promising approach to study pollutant impacts to target organisms in both terrestrial and aquatic environments. To this end, both nuclear magnetic resonance (NMR)- and mass spectrometry (MS)-based methods are used to profile amino acids in different environmental metabolomic studies. However, these two methods have not been compared directly which is an important consideration for broader comparisons in the environmental metabolomics field. We compared the quantification of 18 amino acids in the tissue extracts of Daphnia magna, a common model organism used in both ecotoxicology and ecology, using both 1H NMR spectroscopy and liquid chromatography with tandem MS (LC-MS/MS). 1H NMR quantification of amino acids agreed with the LC-MS/MS quantification for 17 of 18 amino acids measured. We also tested both quantitative methods in a D. magna sub-lethal exposure study to copper and lithium. Again, both NMR and LC-MS/MS measurements showed agreement. We extended our analyses with extracts from the earthworm Eisenia fetida and the plant model Nicotiana tabacum. The concentrations of amino acids by both 1H NMR and LC-MS/MS, agreed and demonstrated the robustness of both techniques for quantitative metabolomics. These findings demonstrate the compatibility of these two analytical platforms for amino acid profiling in environmentally relevant model organisms and emphasizes that data from either method is robust for comparisons across studies to further build the knowledge base related to pollutant exposure impacts and toxic responses of diverse environmental organisms.

Topical treatment containing undeclared corticosteroids and illegal topical treatment with corticosteroid content have been seen on the Swedish market. In creams and ointments corticosteroids in the category of glucocorticosteroids are used to reduce inflammatory reactions and itchiness in the skin. If the inflammation is due to bacterial infection or fungus, complementary treatment is necessary. Side effects of corticosteroids are skin reactions and if used in excess suppression of the adrenal gland function. Therefore the Swedish Medical Products Agency has published related warnings to make the public aware. There are many similar structures of corticosteroids where the anti-inflammatory effect is depending on substitutions on the corticosteroid molecular skeleton. In legal creams and ointments they can be found at concentrations of 0.025 ‑ 1.0 %, where corticosteroids with fluorine substitutions usually are found at concentrations up to 0.1 % due to increased potency. At the Medical Products Agency 19F-NMR and 1H-NMR have been used to detect and quantify corticosteroid content in creams and ointments. Nuclear Magnetic Resonance, NMR, is an analytical technique which is quite sensitive and can have a relative short experimental time. The low concentration of corticosteroids makes the signals detected in NMR small and in 1H‑NMR the signals are often overlapped by signals from the matrix. With 1H‑NMR characteristic signals could be detected in a less crowded spectral window between 5.96 ‑ 6.40 ppm where overlapping signals from the matrix often are absent. Since fluorine is less common in molecules, the option of using 19F‑NMR increases the possibility of finding fluorine-containing corticosteroids in creams and ointments. The corticosteroid signals in 19F‑NMR are detected at -165 ppm and -187 ppm, depending on where fluorine is located on the structure. Quantifying with 1H-NMR and 19F-NMR gave similar result with an accuracy of 98‑116 % and 89-106 % respectively, and RSD values between 2‑35 %, depending on the kind and amount of corticosteroid. Relations between the structure and some signals in 1H‑NMR were found, making it easier to determine the basic structure of unknown corticosteroids in creams and ointments. Screening experiments were performed on creams and ointments with known concentration corticosteroid in order to find minimum NS for analyzing products which might contain corticosteroids. In order to detect a corticosteroid concentration of 0.05 % 19F‑NMR needed 64 NS with an experimental time of 2 min and 1H-NMR needed 160 NS with an experimental time of 17 min. Concentrations of 0.025 % could for some corticosteroids be detected with these parameters. The possibility of spiking samples in order to discriminate between corticosteroids was also investigated. The corticosteroids available at the MPA could be discriminated from each other with at least one of the methods 1H‑NMR or 19F-NMR, and in most cases with both. A market research was done in order to search for counterfeits and salespersons in different health food stores were asked to recommend the best product to treat eczema or psoriasis. Nine recommended products were bought where one was found illegally containing a corticosteroid. In previous experiments at the MPA there had been occurrences of a split signal in 19F-NMR when analyzing creams. The split 19F‑NMR signal was shown to be related both to the presence of water and structural effects of the corticosteroid

Il cancro della prostata (PCa) è il tumore maligno non-cutaneo più diffuso tra gli uomini ed è il secondo tumore che miete più vittime nei paesi occidentali. La necessità di nuove tecniche non invasive per la diagnosi precoce del PCa è aumentata negli anni. 1H-MRS (proton magnetic resonance spectroscopy) e 1H-MRSI (proton magnetic resonance spectroscopy imaging) sono tecniche avanzate di spettroscopia in risonanza magnetica che permettono di individuare presenza di metaboliti come citrato, colina, creatina e in alcuni casi poliammine in uno o più voxel nel tessuto prostatico. L’abbondanza o l’assenza di uno di questi metaboliti rende possibile discriminare un tessuto sano da uno patologico. Le tecniche di spettroscopia RM sono correntemente utilizzate nella pratica clinica per cervello e fegato, con l’utilizzo di software dedicati per l’analisi degli spettri. La quantificazione di metaboliti nella prostata invece può risultare difficile a causa del basso rapporto segnale/rumore (SNR) degli spettri e del forte accoppiamento-j del citrato. Lo scopo principale di questo lavoro è di proporre un software prototipo per la quantificazione automatica di citrato, colina e creatina nella prostata. Lo sviluppo del programma e dei suoi algoritmi è stato portato avanti all’interno dell’IRST (Istituto Romagnolo per lo Studio e la cura dei Tumori) con l’aiuto dell’unità di fisica sanitaria. Il cuore del programma è un algoritmo iterativo per il fit degli spettri che fa uso di simulazioni MRS sviluppate con il pacchetto di librerie GAMMA in C++. L’accuratezza delle quantificazioni è stata testata con dei fantocci realizzati all’interno dei laboratori dell’istituto. Tutte le misure spettroscopiche sono state eseguite con il nuovo scanner Philips Ingenia 3T, una delle machine di risonanza magnetica più avanzate per applicazioni cliniche. Infine, dopo aver eseguito i test in vitro sui fantocci, sono stati acquisiti gli spettri delle prostate di alcuni volontari sani, per testare se il programma fosse in grado di lavorare in condizioni di basso SNR.

Neste trabalho prepararam-se ésteres de ácidos graxos por esterificação do ácido oléico e transesterificação do óleo de soja e do triéster oléico via catálise enzimática. Determinou-se a composição dos produtos obtidos por RMN 1H e CG-FID. Os padrões dos ésteres do ácido oléico via esterificação ácida foram preparados empregando ácido sulfúrico, os padrões dos ésteres do óleo de soja por transesterificação básica com hidróxido de sódio e o padrão do triéster oléico foi sintetizado utilizando ácido p-tolueno sulfônico como catalisador. A melhor enzima para essas reações foi determinada através de reações de esterificação do ácido oléico e transesterificação do óleo de soja com etanol na ausência de co-solventes, sendo que foi selecionada a lipase de Candida antarctica. A mesma enzima foi empregada nas esterificações enzimáticas do ácido oléico com diversos alcoóis (metanol, etanol, n-propanol e n-butanol), na transesterificação enzimática do óleo de soja e do triéster oléico com etanol. Em ambos os estudos, foram avaliados os fatores que influenciam as reações: quantidade de enzima; tempo de reação; água adicionada ao álcool e reuso do biocatalisador. Na esterificação enzimática do ácido oléico, o uso do etanol forneceu o melhor rendimento (96,5%) com 5,0% (m/m) de enzima em 24 horas de reação. Quando uma quantidade de 4,0% de água foi adicionada ao álcool, a reação utilizando metanol mostrou maior eficiência (98,5%) e os rendimentos com os outros alcoóis não foram alterados significativamente (acima de 90%). Também foi possível utilizar a enzima por até 10 ciclos sem perda de rendimento, com exceção do metanol, onde ocorreu um decréscimo acentuado de rendimentos nos ciclos seguintes. Na transesterificação enzimática do óleo de soja, os mesmos fatores foram avaliados e com 5,0% de enzima, após 24 horas, foi obtido um rendimento de 84,1% e com a adição de água o rendimento não foi significativamente alterado (83%). Na transesterificação, os métodos de quantificação por RMN 1H e CG-FID foram comparados sendo que uma maior diferença foi observada para as reações com baixos rendimentos por RMN 1H, porém em altos rendimentos a diferença entre os dois métodos não foi significativa. Monoglicerídeos e diglicerídeos foram quantificados por CG-FID e por RMN 1H onde foi possível calcular a razão dos produtos formados através de uma equação desenvolvida, sendo que a diferença entre esses tipos de análises foi pequena, de apenas 1,4%. A transesterificação enzimática do triéster oléico foi obtida em bom rendimento (90,4%) e uma pequena quantidade de mono- e diglicerídeos foi produzida. Em todas as reações de transesterificação, o glicerol não foi detectado após a lavagem dos produtos. A metodologia empregando a lipase de Candida antarctica mostrou-se eficiente para a produção de biodiesel a partir do óleo de soja e do ácido oléico com diferentes tipos de alcoóis.
In this work, it was prepared esters of the fatty acid by esterification of the oleic acid and transesterification of the soy oil through enzymatic catalysis. It was determined the composition of the products obtained by 1H NMR and GC-FID. The standards of esters of the oleic acid by acid esterification was prepared employing sulfuric acid, the standards of esters of the soy oil by alkaline transesterification with sodium hydroxide and the standard of the oleic triester was synthesized employing p-toluene sulfonic acid as catalyst. The best enzyme for those reactions was determined through reactions of esterification of the oleic acid and transesterification of the soy oil with ethanol and free co-solvents, and lipase from Candida antarctica was selected. The same enzyme was employed in the enzymatic esterifications of the oleic acid with various alcohols (methanol, ethanol, propanol and butanol), in the enzymatic transesterification of soy oil and the oleic triester with ethanol. In both studies, was assessed the factors that influence the reactions: amount of catalyst, reaction time, water added in the alcohol and the turnover of biocatalyst. In the enzymatic esterification of the oleic acid, the ethanol showed the better yield (96,5%) with 5,0% (m/m) of enzyme at 24 hours of reaction. When 4,0% of water was added to the alcohol, the methanol showed the high efficiency (98,5%) and the yield with another alcohols were not affected. It was also possible to use the enzyme for 10 cycles without lose yield, except for the methanol. In the enzymatic transesterification of the soy oil, the same factors were assessed using 5,0% of enzyme, after 24 hours, a yield of 84,1% was obtained and with the water addition the yield was not modified (83%). On the transesterification, 1H NMR and GC-FID were compared and a great difference was observed for low yields, but on high yields, the difference between methods was small. Monoglycerides and diglycerides were quantified by GC-FID and detected by 1H NMR, it was possible to calculate the ratio between them on the products formed through an equation developed and the difference for this type of analysis was small, only 1.4% . The enzymatic transesterification of the oleic triester was obtained with good yield (90,4%) and a small amount of the monoglycerides and diglycerides was produced. In all the transesterifications reactions, glycerol was not detected after washing mixture of products. The methodology employing Candida antarctica lipase was efficient for biodiesel production by soybean oil and oleic acid with different alcohols.

L’objectif de ce travail de thèse était de déterminer la composition chimique d’extraits de plantes de Corse encore peu ou pas étudiées et susceptibles d’avoir des activités biologiques valorisables. Cette étude a été menée en mettant en œuvre la méthodologie d’analyse des mélanges complexes par RMN 13C, développée depuis une trentaine d’années par l’équipe « Chimie et Biomasse » de l’UMR CNRS 6134 Sciences pour l’Environnement de l’université de Corse. Dans une première partie, nous avons étudié la composition chimique d’extraits de houx commun (Ilex aquifolium L.). Deux extraits de feuilles (obtenus à l’hexane et au dichlorométhane) ont chacun subi deux fractionnements successifs par chromatographie sur colonne ouverte de silice. L’analyse par RMN 13C (CPG(Ir) et CPG-SM également dans certains cas) des extraits bruts et des fractions de chromatographie a permis l’identification de onze triterpènes dont l’α-amyrine et la β-amyrine et certains de leurs esters. Parallèlement, deux acides triterpéniques d’intérêt (acide ursolique et acide oléanolique) ont été quantifiés par RMN 1H au sein de l’extrait brut au dichlorométhane via une méthode rapide mise au point et validée (justesse, linéarité, précision des mesures). Ces deux composés représentent respectivement 55,3% et 20,8% de l’extrait. Nous avons également réalisé, en collaboration avec l’équipe « Biochimie et Biologie Moléculaire du Végétal » de l’université de Corse, des tests antimicrobiens sur les extraits et certaines fractions de chromatographie. Les deux acides triterpéniques précédemment cités ont montré une activité antimicrobienne comparable à celle du chloramphénicol (antibiotique de référence) vis-à-vis de trois bactéries à Gram positif, Staphylococcus aureus, Staphylococcus epidermidis et Bacillus cereus (CMI = 4 et 8 mg.L-1 vs. 2 et 4 mg.L-1). Par ailleurs, nous avons préparé des extraits de baies de houx en utilisant différents solvants : hexane, dichlorométhane, dichlorométhane/acétate d’éthyle (50/50, v/v). Les deux derniers extraits (dichlorométhane et dichlorométhane/acétate d’éthyle) ont subi des fractionnements successifs et l’étude des spectres RMN 13C des extraits bruts et des fractions de chromatographie a permis d’identifier neuf triterpènes (précédemment identifiés), cinq dérivés phénoliques, six monosaccharides ainsi que quatre lactones (la ménisdaurilide, l’aquilégiolide, la 7-épi-griffonilide et la dasycarponilide) non encore répertoriées dans le houx commun. Dans une seconde partie, nous avons étudié la composition chimique d’extraits de fleurs et de racine de Calicotome villosa (Poir.) Link. Les extraits de fleurs obtenus au dichlorométhane et à l’acétate d’éthyle ont été soumis à des chromatographies successives. L’analyse par RMN 13C des extraits bruts et des fractions de chromatographie a permis l’identification de trois flavonoïdes, cinq dérivés glucosylés de flavonoïdes et quatre phénylpropanoïdes. Dans cette partie, nous nous sommes également intéressés à la composition chimique d’un extrait méthanolique de racine de C. villosa qui n’a fait l’objet d’aucune étude de ce type au préalable. Cet extrait a été soumis à plusieurs fractionnements successifs et dix-huit composés, notamment des stérols, des flavonoïdes, un polyphénol et des ptérocarpanes ont été identifiés par RMN 13C
The aim of this work was to determine chemical composition of wild growing corsican understudied plant species, with potential biological activities. This study was realized using the computerized NMR method developed over the past thirty years by the University of Corsica “Chimie et Biomasse” group, UMR CNRS “Sciences Pour l’environnement”. Identified secondary metabolites were then undertaken to evaluate their antimicrobial properties. As part of this study, we selected on the first place commun holly (Ilex aquifolium L.). After two successive column chromatography, hexane and dichloromethane leaves crude extracts and all chromatography fractions were analyzed by 13C NMR (GC(RI) and GC-MS sometimes) to allow the identification of eleven triterpens and α- and β-amyrin esters. Among identified compounds, ursolic acid and oleanolic acid were also quantified by 1H NMR in the dichloromethane crude extract using a reliable method developped and validated (accuracy, linearity precision of measurements). Ursolic acid accounted for 55.3% of the extract, followed by oleanolic acid, 20.8%. Evaluation of previous identified compounds antimicrobial activities has been performed in collaboration with « Biochimie et Biologie Moléculaire du Végétal » group (University of Corsica). Triterpen acids and chloramphenicol (reference antibiotic) displayed similar antibacterial activities against three Gram-positive bacteria, Staphylococcus aureus, Staphylococcus epidermidis and Bacillus cereus (MIC = 4 and 8 mg.L-1 vs. 2 and 4 mg.L-1). Moreover, dichloromethane and dichloromethane/ethyl acetate (50/50, v/v) berries extracts were submitted to successive column chromatography. Crude extracts and chromatography fractions 13C NMR spectra revealed presence of nine triterpens previously identidied, five phenolics derivatives, six monosaccharides and four lactones (menisdaurilide, aquilegiolide, dasycarponilide and 7-epi-griffonilide) were first time identified in berries holly extracts. On the second place, we determined chemical compositions of Calicotome villosa (Poir.) Link flowers and root extracts. 13C NMR analysis of dichloromethane and ethyl acetate flowers extracts and their chromatography fractions, allowed the identification of three flavonoids, five glycosyl flavonoids and four phenylpropanoids. We also studied chemical composition of a methanol root extract which has never been submitted to any chemical composition study. After successive column chromatography, eighteen compounds were identified by 13C NMR including sterols, flavonoids, a polyphenol and pterocarpans

Implementations of high-field nuclear magnetic resonance (NMR) facilities into metabolomics studies are unfortunately restricted by their large dimensions, high costings, and specialist technical staff requirements. Therefore, here the application and practical advantages offered by low-field (60 MHz), compact NMR spectrometers for probing the metabolic profiles of human saliva was explored, as was their value in salivary metabolomics studies. Saliva samples were collected from cigarette smoking (n = 11) and non-smoking (n = 31) human participants. 1H NMR spectra were acquired on both low-field (60 MHz) and medium-field (400 MHz) spectrometers. Metabolomics analyses were employed to evaluate the consistencies of salivary metabolite levels determined, and their abilities to distinguish between smokers and non-smokers. Low-field 1H NMR analysis detected up to 15, albeit permitted the reliable quantification of 5, potentially key diagnostic biomolecules simultaneously (LLOQ values 250–400 μmol/L), although these were limited to those with the most prominent resonances. Such low-field profiles were also found to be suitable for salivary metabolomics investigations, which confirmed the successful discrimination between smoking and non-smoking participant sample donors. Differences observed between these groups were largely ascribable to upregulated salivary levels of methanol, and its metabolite formate, in the smoking group, but higher smoking-mediated concentrations of acetate, propionate and glycine may arise from a diminished salivary flow-rate in these participants. In conclusion, determination of salivary biomolecules using low-field, benchtop 1H NMR analysis techniques were found to be valuable for bioanalytical and metabolomics investigations. Future perspectives for the applications of this non-stationary NMR technique, for example for the on-site ‘point-of-care’ testing of saliva samples for diagnostic oral disease screening purposes at dental surgeries and community pharmacies, are considered.

Natural organic matter (NOM) is a major intermediate in the global carbon, nitrogen, sulfur, and phosphorus cycles. NOM is also the environmental matrix that frequently controls binding, transport, degradation, and toxicity of many organic and inorganic contaminants. Despite its importance, NOM is poorly understood at the structural chemistry level because of its molecular complexity and heterogeniety. Nuclear magnetic resonance (NMR) spectroscopy is one of the most useful spectrometric methods used to investigate NOM structure because qualitative and quantitative organic structure information for certain organic elements can be generated by NMR for NOM in both the solution and solid states under nondegradative conditions. However, NMR spectroscopy is not as sensitive as infrared or ultraviolet-visible spectroscopy; it is not at present applicable to organic oxygen and sulfur, and quantification of NMR spectra is difficult under certain conditions. The purpose of this overview is to present briefly the “state of the art” of NMR characterization of NOM, and to suggest future directions for NMR research into NOM. More comprehensive texts concerning the practice of NMR spectroscopy and its application to NOM in various environments have been produced by Wilson and by Wershaw and Mikita. Carbon, hydrogen, and oxygen are the major elements of NOM; together they comprise about 90% of the mass. The minor elements that constitute the remainder are nitrogen, sulfur, phosphorus, and trace amounts of the various halogen elements. With the exception of coal, in which carbon is the most abundant element, the order of relative abundance in NOM on an atomic basis is H > C > O > N > S > P = halogens. The optimum NMR-active nuclei for these elements are 1H, 13C, 17O, 15N, 33S, 31P, and 19F. The natural abundances and receptivities of these nuclei relative to 1H are given in Table 12.1. Quadrupolar effects for 17O, 33S, and halogen elements other than 19F lead to line broadening that greatly limits resolution in NMR studies of these elements in NOM.

Lipid oxidation is a chemical process that is detrimental for the quality of oil-based foods. Historically, hydroperoxides and aldehydes are considered the most important molecules in the assessment of lipid oxidation. While they do provide insights in kinetics of primary and secondary oxidation, it has been proposed that a third class of oxidation products, the epoxides, could help resolve mechanistical anomalies that are hitherto unexplained.Here, we developed a 2D 1H-13C HSQC NMR method that allows for a quantitative assessment of epoxides in real food products. The method was applicable in vegetable oils and mayonnaises in a reproducible (RSD ‰¤ 11.6 %) and repeatable (RSD ‰¤ 8.2 %) manner with an LoD and LoQ of respectively 0.18 and 0.62 mmol/kg.This novel method was applied in combination with existing NMR methods to simultaneously quantify lipid hydroperoxides, aldehydes and epoxides, providing a more comprehensive view on lipid oxidation pathways under different conditions. Epoxides showed to be a major product during intermediate and late stage of oxidation. However, its potential as an early marker for lipid oxidation appeared to be limited. A combination of hydroperoxides, epoxides, and aldehydes, on the other hand, could be a powerful combination to capture the main radical mechanisms during oxidation. Moreover, the characterization and quantification of individual epoxide sub-classes by 2D 1H-13C HSQC NMR enables mechanistical oxidation studies.