A comparative NMR-based metabolomics study of lung parenchyma of severe COVID-19 patients.

COVID-19 was the most significant infectious-agent-related cause of death in the 2020-2021 period. On average, over 60% of those admitted to ICU facilities with this disease died across the globe. In severe cases, COVID-19 leads to respiratory and systemic compromise, including pneumonia-like symptoms, acute respiratory distress syndrome, and multiorgan failure. While the upper respiratory tract and lungs are the principal sites of infection and injury, most studies on the metabolic signatures in COVID-19 patients have been carried out on serum and plasma samples. In this report we attempt to characterize the metabolome of lung parenchyma extracts from fatal COVID-19 cases and compare them with that from other respiratory diseases. Our findings indicate that the metabolomic profiles from fatal COVID-19 and non-COVID-19 cases are markedly different, with the former being the result of increased lactate and amino acid metabolism, altered energy pathways, oxidative stress, and inflammatory response. Overall, these findings provide additional insights into the pathophysiology of COVID-19 that could lead to the development of targeted therapies for the treatment of severe cases of the disease, and further highlight the potential of metabolomic approaches in COVID-19 research.

A nitroalkene derivative of salicylate alleviates diet-induced obesity by activating creatine metabolism and non-shivering thermogenesis.

Obesity-related type II diabetes (diabesity) has increased global morbidity and mortality dramatically. Previously, the ancient drug salicylate demonstrated promise for the treatment of type II diabetes, but its clinical use was precluded due to high dose requirements. In this study, we present a nitroalkene derivative of salicylate, 5-(2-nitroethenyl)salicylic acid (SANA), a molecule with unprecedented beneficial effects in diet-induced obesity (DIO). SANA reduces DIO, liver steatosis and insulin resistance at doses up to 40 times lower than salicylate. Mechanistically, SANA stimulated mitochondrial respiration and increased creatine-dependent energy expenditure in adipose tissue. Indeed, depletion of creatine resulted in the loss of SANA action. Moreover, we found that SANA binds to creatine kinases CKMT1/2, and downregulation CKMT1 interferes with the effect of SANA in vivo. Together, these data demonstrate that SANA is a first-in-class activator of creatine-dependent energy expenditure and thermogenesis in adipose tissue and emerges as a candidate for the treatment of diabesity.

Beyond cannabinoids: Application of NMR-based metabolomics for the assessment of Cannabis sativa L. crop health.

While Cannabis sativa L. varieties have been traditionally characterized by their major cannabinoid profile, it is now well established that other plant metabolites can also have physiological effects, including minor cannabinoids, terpenes, and flavonoids. Given the multiple applications of cannabis in the medical field, it is therefore critical to characterize it according to its chemical composition (i.e., its metabolome) and not only its botanical traits. With this in mind, the cannabinoid and metabolomic profiles from inflorescences of two C. sativa varieties with either high Δ9-tetrahydrocannabinolic acid (THCA) or high cannabidiolic acid (CBDA) contents harvested at different times were studied. According to results from HPLC and NMR-based untargeted metabolomic analyses of organic and aqueous plant material extracts, we show that in addition to expected variations according to cannabinoid profiles, it is possible to distinguish between harvests of the same variety. In particular, it was possible to correlate variations in the metabolome with presence of powdery mildew, leading to the identification of molecular markers associated with this fungal infection in C. sativa.

Synthesis and antitumoral evaluation of natural product-like compounds based on tropolone and benzotropolone derivatives.

We present the preparation of a series of novel natural product-like homobarrelenones, norcaranes, and dihydrofluorenones through a diversity-oriented synthetic (DOS) strategy that combines Diels-Alder reactions and phototransformations, as well as their biological evaluation against MCF-7, HT-29, and NCI-H460 human tumor cells. Six of these demonstrated activities in the micromolar range against the three cell lines, and none were predicted as cytotoxic against human nontumor cells according to in silico studies. In addition, within the set of active derivatives, three exhibited low unspecific cytotoxicity in a sperm motility assay. The rich functionality of the new compounds makes them ideal candidates for exhaustive structure-activity relationship studies.

Predicting Mandarin Fruit Acceptability: From High-Field to Benchtop NMR Spectroscopy.

Recent advances in nuclear magnetic resonance (NMR) have led to the development of low-field benchtop NMR systems with improved sensitivity and resolution suitable for use in research and quality-control laboratories. Compared to their high-resolution counterparts, their lower purchase and running costs make them a good alternative for routine use. In this article, we show the adaptation of a method for predicting the consumer acceptability of mandarins, originally reported using a high-field 400 MHz NMR spectrometer, to benchtop 60 MHz NMR systems. Our findings reveal that both instruments yield comparable results regarding sugar and citric acid levels, leading to the development of virtually identical predictive linear models. However, the lower cost of benchtop NMR systems would allow cultivators to implement this chemometric-based method as an additional tool for the selection of new cultivars.

Identification of chia, flax and sesame seeds authenticity markers by NMR-based untargeted metabolomics and their validation in bakery products containing them.

Chia, flax, and sesame seeds are considered superseeds due to their beneficial nutritional properties, and they are frequently included as functional ingredients in foods. Authenticity markers of these seeds, including bakery products containing them, have been identified by both liquid and gas chromatography coupled to mass spectrometry (LC-MS/MS and GC-MS/MS, respectively) targeted metabolomics. However, there are no reports describing the use of nuclear magnetic resonance (NMR) spectroscopy based metabolomics to identify authenticity markers in either the raw seeds or foods containing them. We herein report the application of an untargeted NMR-based metabolomics workflow to the identification of authenticity markers for the three seeds. Seven markers, belonging to the families of polyphenols and cyanogenic glycosides, allowed good differentiation of the raw materials. Validation in cookies containing different seed percentages showed that two markers resisted the processing stage, making them feasible authenticity markers for the food trade.

New Insights into the Chemical Composition of Ayahuasca.

Ayahuasca is a psychedelic beverage originally from the Amazon rainforest used in different shamanic settings for medicinal, spiritual, and cultural purposes. It is prepared by boiling in water an admixture of the Amazonian vine Banisteriopsis caapi, which is a source of ß-carboline alkaloids, with plants containing N,N-dimethyltryptamine, usually Psychotria viridis. While previous studies have focused on the detection and quantification of the alkaloids present in the drink, less attention has been given to other nonalkaloid components or the composition of the solids suspended in the beverage, which may also affect its psychoactive properties. In this study, we used nuclear magnetic resonance (NMR) and liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) to study the composition of ayahuasca samples, to determine their alkaloid qualitative and quantitative profiles, as well as other major soluble and nonsoluble components. For the first time, fructose was detected as a major component of the samples, while harmine (a ß-carboline previously described as an abundant alkaloid in ayahuasca) was found to be present in the solids suspended in the beverage. In addition, N,N-dimethyltryptamine (DMT), harmine, tetrahydroharmine, harmaline, and harmol were identified as the major alkaloids present in extracts of all samples. Finally, a novel, easy, and fast method using quantitative NMR was developed and validated to simultaneously quantify the content of these alkaloids found in each ayahuasca sample.

Characterization of Melanoidins and Color Development in Dulce de Leche, a Confectionary Dairy Product With High Sucrose Content: Evaluation of pH Effect, an Essential Manufacturing Process Parameter.

The effect on color of the initial pH employed in dulce de leche (DL) production was evaluated through physicochemical and spectroscopical characterization of the melanoidins formed in the process. Melanoidins originated at pH values of 6.5, 7.0, and 7.5, and they were released by the enzymatic hydrolysis of the protein backbone and purified by gel filtration. They showed a significant degree of polydispersity, in general, with molecular weights (MWs) below 1,800 Da. DL produced at a higher pH released melanoidins with higher average MW after the enzymatic hydrolysis. They also presented darker colors (dE*ab, C*), more closely resembling those typical of the commercial product. Analysis of the fractions isolated by gel filtration using HPLC-DAD and multinuclear NMR showed an heterogeneous and complex composition. Even though structurally related, the 1H NMR spectra of melanoidins showed a higher degree of aromaticity at higher pH values. In conclusion, the pH employed in DL production affects the amount and structure of the colored products originated by MR reactions, and thus the color of the final product.

An Integrative Approach for the Characterization of Plant-Pathogenic Streptomyces spp. Strains Based on Metabolomic, Bioactivity, and Phylogenetic Analysis.

Actinomycetes are generally recognized as a diverse group of gram-positive, mycelium-forming, soil bacteria that play an important role in mineralization processes and plant health, being Streptomyces the most well-known genus from this group. Although plant pathogenicity is a rare attribute in this genus, some species have significant impact worldwide due to their ability to cause important crop diseases such as potato common scab (CS). In this work, an integrative approach was applied to investigate the pathogenic potential of Streptomyces spp. isolates obtained from a local collection of actinomycetes isolated from potato fields. Secretion of phytotoxic compounds was verified in most pathogenic strains from our collection (27 out of 29), and we followed metabolomic analysis to investigate those phytotoxins. We first evaluated the production of the known phytotoxins thaxtomin A (TXT) and desmethylmensacarcin (DMSN) in phytotoxic Streptomyces spp. by HPLC analysis, resulting in 17 TXT and 6 DMSN producers. In addition, NMR-based metabolomic models were able to classify strains according to their phytotoxicity, and metabolomic data was also used to infer chemotaxonomy within pathogenic species. A correlation between phylogeny and the production of distinct phytotoxins was found, supporting the idea that there are "species specific" metabolites produced by this genus. The recently discovered polyketide DMSN was associated unequivocally with S. niveiscabiei strains and was not produced by other species in the growth conditions employed. Two S. niveiscabiei and two S. puniciscabiei phytotoxic strains that did not produce TXT nor DMSN suggest the production of other kind of metabolites involved in phytotoxicity, and allowed the prioritization of these strains for further chemical studies. Indeed, we found two S. niveiscabiei strains whose supernatants were not phytotoxic in the radish assay, suggesting other pathogenic mechanisms involved. We believe our work will be useful to help understand relations between metabolites and phylogenetic clades within actinomycetes.

Isolation and Characterization of Melanoidins from Dulce de Leche, A Confectionary Dairy Product.

Melanoidins, the brown-colored compounds formed through the Maillard reaction, are responsible for color development in dulce de leche (DL), a popular confectionary dairy product in the Río de la Plata region, particularly in Uruguay and Argentina. Color is a critical quality parameter that strongly influences consumer preference. This work aimed to develop a method to perform preliminary structural characterization of the chromophores produced by the Maillard reaction. Melanoidins are present in a water-insoluble fraction, linked to a protein backbone, conforming melanoproteins of high molecular weight. The insoluble melanoprotein fraction (10% total solids) was isolated, and the chromophores released by proteolysis and isolated by gel-permeation chromatography. The analysis of the products revealed that they present a high degree of molecular weight (MW) polydispersity, in a range of 300 to 2000 Da, where the compounds of higher molecular weight contributed the most to the color of the product. The isolated fractions were further analyzed by RP-HPLC using a diode array detector (DAD) detector. These results, together with H-NMR data, suggested that the chromophores isolated belonged to a relatively simple mixture of aromatic products with higher hydrophobic character relative to other products of the melanoprotein digestion.

Isolation and structural characterization of a non-diketopiperazine phytotoxin from a potato pathogenic Streptomyces strain.

Two Streptomyces spp. strains responsible for potato common scab infections in Uruguay which do not produce diketopiperazines were identified through whole-genome sequencing, and the virulence factor produced by one of them was isolated and characterized. Phylogenetic analysis showed that both pathogenic strains can be identified as S. niveiscabiei, and the structure of the phytotoxin was elucidated as that of the polyketide desmethylmensacarcin using MS and NMR methods. The metabolite is produced in yields of ∼200 mg/L of culture media, induces deep necrotic lesions on potato tubers, stuns root and shoot growth in radish seedlings, and is comparatively more aggressive than thaxtomin A. This is the first time that desmethylmensacarcin, a member of a class of compounds known for their antitumor and antibiotic activity, is associated with phytotoxicity. More importantly, it represents the discovery of a new virulence factor related to potato common scab, an economically-important disease affecting potato production worldwide.

Determining the relative strengths of aromatic and aliphatic C-H⋠⋠⋠X hydrogen bonds in imidazolium ionic liquids through measurement of H/D isotope effects on 19 F nuclear shielding.

The relative strengths of aromatic and aliphatic C-H⋅⋅⋅X hydrogen bonds in imidazolium ionic liquids were investigated through measurement of H/D isotope effects on the 19 F nuclear shielding of deuterated isotopologues of 1-n-butyl-3-methylimidazolium hexafluorophosphate and tetrafluoroborate ([C4 mim]PF6 and [C4 mim]BF4 ). Δ19 F(H,D) values ranging from 9.7 to 49.7 ppb were observed for [C4 mim]PF6 isotopologues, while for the [C4 mim]BF4 series these went from 26.2 to 83.8 ppb. Our findings indicate that the interactions between the fluorinated anions and protons on the C-1' and C-1″ position of the N-alkyl sidechains are comparable to, and in some cases stronger than, those involving protons on the aromatic ring, underscoring the role that these weak interionic forces have on the local ordering of imidazolium salts in the liquid state. Copyright © 2017 John Wiley & Sons, Ltd.

Xanthane sesquiterpenoids from the roots and flowers of Xanthium cavanillesii.

The sesquiterpene lactones xanthanodiene, 4-epi-xanthanol, 4-epi-isoxanthanol, and 4-epi-xanthinin, as well as the xanthanolide derivative 4-oxo-bedfordia acid were isolated from the chloroform extracts of roots and flowers of Xanthium cavanillesii Schouw. The identities of these compounds were corroborated through comparison of their spectroscopic data, including IR, MS, and (1)H and (13)C NMR assignments, with literature reports. In addition, the structural characterization of 4-oxo-bedfordia acid was revisited and a comprehensive spectroscopic study of the compound is presented. This is to our knowledge the first phytochemical investigation of the roots of X. cavanillesii, and of flowers in the whole Xanthium genus.

Negligible elongation of mucin glycans with Gal ß1-3 units distinguishes the laminated layer of Echinococcus multilocularis from that of Echinococcus granulosus.

The larval stages of the cestodes Echinococcus multilocularis and Echinococcus granulosus cause the important zoonoses known as larval echinococcoses. These larvae are protected by a unique, massive, mucin-based structure known as the laminated layer. The mucin glycans of the E. granulosus laminated layer are core 1- or core 2-based O-glycans in which the core Galpß1-3 residue can initiate a chain comprising one to three additional Galpß1-3 residues, a motif not known in mammalian carbohydrates. This chain can be capped with a Galpα1-4 residue, and can be ramified with GlcNAcpß1-6 residues. These, as well as the GlcNAcpß1-6 residue in core 2, can be decorated with the Galpα1-4Galpß1-4 disaccharide. Here we extend our analysis to the laminated layer of E. multilocularis, showing that the non-decorated cores, together with Galpß1-3(Galpα1-4Galpß1-4GlcNAcpß1-6)GalNAc, comprise over 96% of the glycans in molar terms. This simple laminated layer glycome is exhibited by E. multilocularis grown either in vitro or in vivo. Interestingly, all the differences with the complex laminated layer glycome found in E. granulosus may be explained in terms of strongly reduced activity in E. multilocularis of a putative glycosyltransferase catalysing the elongation with Galpß1-3. Comparative inter-species analysis of available genomic and transcriptomic data suggested a candidate for this enzyme, amongst more than 20 putative (non-core 1) Gal/GlcNAc ß1-3 transferases present in each species as a result of a taeniid-specific gene expansion. The candidate gene was experimentally verified to be transcribed at much higher levels in the larva of E. granulosus than that of E. multilocularis.

Aggregation Behavior of 6-Isocassine and N-Methyl-6-Isocassine: Insights into the Biological Mode of Action of Lipid Alkaloids.

The aggregation behavior of 6-isocassine and N-methyl-6-isocassine, two piperidin-3-ol alkaloids isolated respectively from the barks of Prosopis nigra and P. affinis, was investigated using a combination of NOE experiments and diffusion measurements in solvents of varying polarity and hydrogen bonding capacity. While the NOE enhancements for N-methyl-6-isocassine are positive, regardless of the solvent, those for 6-isocassine shift from negative to positive when going from chloroform-d to methanol-d(4) solution. In addition, despite the self-diffusion coefficients of both compounds being virtually identical in methanol-d4, N-methyl-6-isocassine diffuses nearly twice as fast as the non-methylated alkaloid in chloroform-d. The changes in rotational and translational dynamics observed between solvents for 6-isocassine suggest that the molecule forms dimeric head-to-head aggregates in non-polar aprotic environments, a behavior that could help explain the biological mode of action that has been proposed for this type of alkaloids.

Observation of aliphatic C-H···X hydrogen bonds in imidazolium ionic liquids.

Measurement of H/D isotope effects on the (19)F resonances of sequentially deuterated 1-n-butyl-3-methylimidazolium hexafluorophosphate and tetrafluoroborate isotopologues reveals the formation of aliphatic C-H···X hydrogen bonds between the fluorinated anions and protons along the N-alkyl sidechains of the cations.

Synthesis, characterization, microbiological evaluation, genotoxicity and synergism tests of new nano silver complexes with sulfamoxole: X-ray diffraction of [Ag2(SMX)2]·DMSO.

The synthesis and microbiological evaluation of two new Ag(I) complexes with sulfamoxole (SMX), [Ag2(SMX)2]·H2O and [Ag4(SCN)3(SMX)]·H2O are described. Both were characterized by elemental analysis, thermogravimetry, powder and single crystal X-ray diffraction, NMR, Raman and experimental and theoretical IR spectroscopies. Their antibacterial and antifungal properties were evaluated by agar and broth dilution assays, respectively. In addition, synergism tests for Pseudomonas aeruginosa were performed, and genotoxicity studies were carried out employing the Allium cepa test. Both complexes displayed good activity against Escherichia coli, Staphylococcus aureus, P. aeruginosa, and 10 fungi strains, with lower minimum inhibitory concentrations (MICs) than that of free SMX in all cases. The nanometrical crystallite particle size determined from XRPD, DLS and TEM might explain the good microbiological activity in spite of the low solubility of both complexes. The fractional inhibitory concentration (FIC) calculated from the P. aeruginosa test data indicated that the activity of the complexes is not due to synergism of the free components in the concentration ratios studied. Moreover, none of the complexes displayed cytotoxic effects on onions in the concentration range tested, and chromosome aberrations were not observed.

Conformational preferences of furan- and thiophene-based arylamides: a combined computational and experimental study.

We examine the conformational preferences of the furan- and thiophene-based arylamides, N-methylfuran-2-carboxamide (3) and N-methylthiophene-2-carboxamide (4), using a combination of computational methods and NMR experiments. The compound choice stems from their use as foldamer building blocks. We quantify the differences in the conformational rigidity of the two compounds, which governs corresponding foldamer conformations. Specifically, we demonstrate the effects of intramolecular hydrogen bonding (H-bonding), geometrical patterns and solvent polarity on arylamide conformations by comparing 3, 4 and previously studied ortho-methoxy N-methylbenzamide (1) and ortho-methylthio N-methylbenzamide (2). The study reveals that compound 3, despite its non-optimal S(5)-type H-bond geometry, retains a large portion of the H-bonded (eclipsed) conformation even in polar protic solvents. This behaviour is consistent with the quantum mechanical (QM) torsional energy profile. The percentages of H-bonded conformers that 3 retains are just slightly smaller than those of 1, which has a stronger S(6)-type H-bond. As for 2 and 4, the replacement of the O atom in 1 by an S atom in 2 results in a 70­90% loss of the H-bonded conformer in solution. However, the equivalent O to S replacement in 3 (leading to 4) causes only 15­30% loss of the eclipsed conformers in 4. Therefore, conformational preferences of 4 are very different from 2, in contrast to the similarity between 3 and 1. This study shows how the interplay of several forces modulates the conformational flexibility of arylamides. It also attests the strategy we are developing, which leads to accurate prediction of foldamer structure. The vital component of this strategy is the re-parameterization of critical force field parameters based on QM potential energy profiles, as well as validation of these parameters using experimental data in solution.

Synthesis and evaluation as potential anticancer agents of novel tetracyclic indenoquinoline derivatives.

We report the synthesis and evaluation as potential anticancer agents of a series of tetracyclic indenoquinolines. The compounds, which are obtained through the photoisomerization of Diels-Alder adducts formed between purpurogallin derivatives and nitrosobenzene, have in vitro antiproliferative activities in the µM to nM range against breast (MCF-7), lung epithelial (A-549), and cervical (HeLa) adenocarcinoma cells. The cytotoxicities of several of the novel tetracycles are comparable to or better than that of camptothecin. A strong correlation between the activity of the compounds and their aromaticity and planarity was observed, suggesting a mode of action similar to that of topoisomerase poisons.

Sequence-controlled oligomers fold into nanosolenoids and impart unusual optical properties.

Controlled syntheses give unique block oligomers with alternating flexible ethylene glycol and rigid perylenetetracarboxylic diimide (PDI) units. The number of rigid units vary from n=1 to 10. PDI units were stitched together by using efficient phosphoramidite chemistry. The resulting oligomers undergo folding in most solvents, including chloroform. In their ground state, these folded oligomers were characterized by using Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS), NMR spectroscopy, and electronic absorption spectroscopy. FTICR-MS revealed the exact masses of these sequence-controlled oligomers, which confirmed the chemical composition and validated the synthetic strategy. The NMR neighboring ring-current effect (NRE) indicates the formation of cofacial π stacks; the stacked aromatic rings have nearly coaxial alignment akin to a nanosoleniod. Nanosolenoidal shielding in π stacks causes all aromatic protons to shift upfield, whereas NOE in a cyclic hetero-chromophoric dimer supports a rotated, cofacial π-stacking orientation separated by about 3.5 Å. Electron-phonon coupling is much stronger than excitonic coupling in these self-folded PDI oligomers; thus, Franck-Condon factors dictate the observed spectral features in visible spectra. The absorbance spectrum exhibits weak hypochromism due to π stacking with increasing stacking units n. Finally, ab initio calculations support the experimental observations, indicating 3.5 Å cofacial spacing in which one molecule is rotated 30° from the eclipsed orientation and higher oligomers can adopt, without a compensating energy penalty, either the right/left-handed helices or the 1,3-eclipsed structures. Both theory and experiments validate the nano-π-solenoids and their novel photophysical properties.