Rising temperatures due to climate change may affect the quality of open-field cultivated processing tomatoes by altering the nutrient content. Bioinoculants are growing in popularity as a nature-based strategy to mitigate these environmental stresses. Untargeted quantitative NMR spectroscopy was leveraged to characterize the metabolome of tomato fruits exposed to abiotic stress during the year 2022, which was marked by unexpected high temperatures and low rainfall compared to the year 2021 with average conditions. This study was conducted at growing sites in Tarquinia and Viterbo, comparing untreated plants to ones treated with a Trichoderma-based bioinoculant. The hotter year affected the water-soluble fraction (28 compounds), causing an increase in amino acids, citrate, and formate contents while decreasing carbohydrates together with a significant drop in β-sitosterol + campesterol in the organic fraction (11 compounds). The site mainly affected the linolenic acid levels, which were more abundant in Tarquinia than Viterbo in the hotter year, whereas ascorbate and myo-inositol were higher in Tarquinia in both years. The year × site interaction significantly affected the content of several amino acids, glucose, sucrose, and trigonelline. The bioinoculant effect was significant only for sucrose, while its interactions with the other factors showed little to no significance across all the measured metabolites.

In the context of organic farming, the introduction of a local product to wider markets and an evaluation of storage effects, metabolic and transcriptomic variations in two broccoli rabe genotypes from production cycles of two different years were studied by comparing florets of stored fresh (SF) and packaged (P) for 4 days with those harvested fresh from the field (H). Twenty-five hydrosoluble compounds, including amino acids, carbohydrates, and organic acids, were quantified by untargeted nuclear magnetic resonance (NMR). Principal component analysis produced a neat separation among the three commodity statuses with P being the most divergent and SF closer to H. In the packaged florets, carbohydrate levels dropped significantly (over -52%), while the levels of amino acids and organic acids varied. There was an increase in stress-responsive phenylalanine and valine (over 30%) and succinic and alpha-ketoglutaric acids (over 75%). Compound correlation analyses indicated a carbohydrate sink towards gamma-aminobutyric acid (GABA) and lactic acid (LA) metabolism under hypoxic conditions in packaged florets. RNA-seq analysis revealed that over 4000 genes were differentially expressed in SF vs H and 8000 in P vs H. Several CAR and AA pathways were significantly enriched in S and even more significantly in P, when compared to H. A map of gene expression (175 genes) and metabolite contents (14 compounds) was constructed to elucidate the gene routes that lead to accumulation of GABA and LA, known for healthy properties, in P. WGCNA and promoter binding site analyses enabled the identification of transcription factors (bZIP, WRKY, ERF types), interactions, and targeted genes encoding key enzymes in GABA and LA accumulation.

The introduction of benchtop FT-NMR spectrometers in recent years represents a remarkable innovation in various fields, including the food sector. Modern benchtop FT-NMR spectrometers are low-field instruments, with a magnetic field ranging from 1 T to 2.35 T (1H resonance frequency from 43 MHz to 100 MHz), characterized by compact design, ease of use, and low maintenance costs. As in the case of high-field NMR instruments, benchtop NMR spectra (obtained by Fourier transformation) contain important information useful for compound identification and quantification.In this review, a description of the fundamental steps useful both to acquire benchtop NMR spectra and to treat the obtained data is reported together with a wide range of applications in the food field. In particular, peculiar aspects of commercial benchtop instruments as well as NMR data acquisition, processing and treatment are reviewed reporting also a practical pipeline and a list of good practices for benchtop NMR applications. Benchtop FT-NMR applications, mainly focused on food adulteration detection and quality control, are discussed here using targeted, metabolomic, and fingerprinting approaches. Finally, the industrial applicability of benchtop NMR methods in either static or continuous mode is reported.

Polyethylene nanoplastics (NPs) are widely diffused in terrestrial environments, including soil ecosystems, but the stress mechanisms in plants are not well understood. This study aimed to investigate the effects of two increasing concentrations of NPs (20 and 200 mg kg−1 of soil) in lettuce. To this aim, high-throughput hyperspectral imaging was combined with metabolomics, covering both primary (using NMR) and secondary metabolism (using LC-HRMS), along with lipidomics profiling (using ion-mobility-LC-HRMS) and plant performance.Hyperspectral imaging highlighted a reduced plant growth pattern. Several vegetative indexes indicated plant toxicity, with 20 mg kg−1 NPs significantly decreasing lettuce density and vegetation health (as indicated by NDVI and plant senescence reflectance indexes). Consistently, photosynthetic activity also decreased.At the biochemical level, metabolomics and lipidomics pointed out a multi-layered broad biochemical reprogramming of primary and secondary metabolism involving a decrease in sterols, sphingolipids, glycolipids, and glycerophospholipids in response to NPs. The reduction in phosphatidylinositol coincided with an accumulation of diacylglycerols (DAG), suggesting the activation of the phospholipase C lipid signaling pathway. Moreover, nanoplastic treatments down-modulated different biosynthetic pathways, particularly those involved in N-containing compounds and phenylpropanoids. Our mechanistic basis of NPs stress in plants will contribute to a better understanding of their environmental impact.

BACKGROUND:Cauliflower (Brassica oleracea L.) and globe artichoke (Cynara scolymus L.) are vegetables with a high waste index mainly related to stems and leaves. In this study, enzymatic hydrolysates obtained from these wastes were proposed to be used as plant biostimulants. Life cycle assessment methodology was also applied to evaluate environmental performances related to cauliflower and artichoke byproducts. RESULTS:Hydrolysates (HYs) were chemically and biologically characterized. Amino acids, organic acids, amines, polyols, mineral elements, phenols, tannins, flavonoids and sulfur compounds were identified and quantified by means of NMR, inductively coupled plasma mass spectrometry and UV-visible analyses. Cauliflower leaf and flower HYs showed the highest concentration of free amino acids, whereas stems showed the highest concentration of Ca. Regarding artichoke, asparagine, glutamine and aspartic acid were exclusively detected in stems, whereas artichoke leaves showed the highest Mg and Mn levels together with the highest antioxidant activity. The HYs diluted in water were tested as biostimulants. The impacts of five concentrations of HYs (0.00, 0.28, 0.84, 2.52 and 7.56 g L-1) on seed germination and early seedling growth of crimson clover, alfalfa, durum wheat and corn were investigated. CONCLUSIONS:The application of artichoke biostimulant (0.28 g L-1) positively influenced the coefficient of velocity of germination in alfalfa, crimson clover and durum wheat, whereas cauliflower biostimulant significantly improved corn germination speed. © 2024 The Author(s). Journal of the Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Considering that maize (Zea mays L.) is a staple food for a large segment of the population worldwide, many attempts have been made to improve the nutritional value of its grain and at the same time to achieve sustainable cropping systems. The present study aimed to characterize the composition and nutritional value of maize grain as influenced by cropping system, genetic background (variety), and growing year using untargeted NMR metabolomics. The composition of both water- (sugars and polyols, organic acids, and amino acids) and liposoluble metabolites (free and esterified fatty acids, sterols, and lipids) extracted from the maize grain was determined. Multivariate statistical analyses (PCA and ANOVA) pointed to the growing year and the variety as the most important random and fixed factors, respectively, influencing the metabolite profile. The samples were separated along PC1 and PC3 according to the growing year and the variety, respectively. A higher content of citric acid and diunsaturated fatty acids and a lower content of tyrosine, trigonelline, and monounsaturated fatty acids was observed in the organic with respect to the conventional variety. The effect of the cropping system was overwhelmed by the random effect of the growing year. The results provide novel knowledge on the influence of agronomic practices on maize micronutrient contents.

The health-promoting properties of blueberries are widely recognized and are mainly attributed to anthocyanins. However, fruit's chemical composition includes also other components and strongly depends on varieties and climatic conditions. Here, H-1 NMR metabolite profiling and biological activity of four blueberry cultivars (Spartan, Jewels, Misty, Camelia) grown in Central Italy over two years were reported. Untargeted and targeted NMR analyses allowed the quantification of sugars, organic acids, amino acids, anthocyanins, lipids, and other compounds. Spectrophotometric assays evaluated total phenolic and flavonoid content, antioxidant activity, and enzyme inhibitory activity toward cholinesterase, alpha-amylase, alpha-glucosidase, and tyrosinase. Statistical analysis showed a correlation between chemical composition and biological activity, revealing markers specific to blueberry cultivars (quinic acid, quercitrin, myo-inositol, myrtillin, and petunidin-3-O-glucoside). Almost all antioxidant assays were correlated with the chlorogenic acid levels. A strong effect of harvesting on chemical composition and biological activities was observed, with Misty cultivar having the highest antioxidant activity.

The chloroplast (cp) genome diversity has been used in phylogeny studies, breeding, and variety protection, and its expression has been shown to play a role in stress response. Smooth- and curly-leafed endives (Cichorium endivia var. latifolium and var. crispum) are of nutritional and economic importance and are the target of ever-changing breeding programmes. A reference cp genome sequence was assembled and annotated (cultivar 'Confiance'), which was 152,809 base pairs long, organized into the angiosperm-typical quadripartite structure, harboring two inverted repeats separated by the large- and short- single copy regions. The annotation included 136 genes, 90 protein-coding genes, 38 transfer, and 8 ribosomal RNAs and the sequence generated a distinct phyletic group within Asteraceae with the well-separated C. endivia and intybus species. SSR variants within the reference genome were mostly of tri-nucleotide type, and the cytosine to uracil (C/U) RNA editing recurred. The cp genome was nearly fully transcribed, hence sequence polymorphism was investigated by RNA-Seq of seven cultivars, and the SNP number was higher in smooth- than curly-leafed ones. All cultivars maintained C/U changes in identical positions, suggesting that RNA editing patterns were conserved; most cultivars shared SNPs of moderate impact on protein changes in the ndhD, ndhA, and psbF genes, suggesting that their variability may have a potential role in adaptive response. The cp transcriptome expression was investigated in leaves of plants affected by pre-harvest rainfall and rainfall excess plus waterlogging events characterized by production loss, compared to those of a cycle not affected by extreme rainfall. Overall, the analyses evidenced stress- and cultivar-specific responses, and further revealed that genes of the Cytochrome b6/f, and PSI-PSII systems were commonly affected and likely to be among major targets of extreme rain-related stress.

Acidolysis lignins from the species Quercus cerris L. and Eucalyptus camaldulensis Dehnh. were isolated and characterized using high pressure size exclusion chromatography (HP-SEC), Fourier-transform (FTIR) infrared spectroscopy, analytical pyrolysis–gas chromatography–mass spectrometry (Py-GCMS), and two-dimensional heteronuclear single quantum coherence (2D HSQC) NMR spectroscopy. The acidolysis lignins from the two different species varied in chemical composition and structural characteristics, with Q. cerris L. lignin having a higher S/G ratio and higher molar mass averages with a bimodal molar mass distribution. The different analytical techniques FTIR spectroscopy, Py-GCMS, and 2D NMR spectroscopy provided consistent results regarding the S/G ratio of the lignins from the two wood species. Based on the determined high S/G ratio of both oak and eucalypt lignin, the two wood sources could be promoted as substrates for efficient lignin isolation in modern forest biorefineries in order to develop innovative lignin-based value-added biorefinery products.

The recovery of bioactive compounds from crop byproducts leads to a new perspective way of waste reutilization as a part of the circular economy. The present study aimed at an exhaustive metabolite profile characterization of globe artichoke and cauliflower byproducts (leaves, stalks, and florets for cauliflower only) as a prerequisite for their valorization and future implementations. The metabolite profile of aqueous and organic extracts of byproducts was analyzed using the NMR-based metabolomics approach. Free amino acids, organic acids, sugars, polyols, polyphenols, amines, glucosinolates, fatty acids, phospho- and galactolipids, sterols, and sesquiterpene lactones were identified and quantified. In particular, globe artichoke byproducts are a source of health-beneficial compounds including chiro-inositol (up to 10.1 mg/g), scyllo-inositol (up to 1.8 mg/g), sesquiterpene lactones (cynaropicrin, grosheimin, dehydrocynaropicrin, up to 45.5 mg/g in total), inulins, and chlorogenic acid (up to 7.5 mg/g), whereas cauliflower byproducts enclose bioactive sulfur-containing compounds S-methyl-L-cysteine S-oxide (methiin, up to 20.7 mg/g) and glucosinolates. A variable content of all metabolites was observed depending on the crop type (globe artichoke vs. cauliflower) and the plant part (leaves vs. stalks). The results here reported can be potentially used in different ways, including the formulation of new plant biostimulants and food supplements.

In this review, the three different NMR-based approaches usually used to study foodstuffs are described, reporting specific examples. The first approach starts with the food of interest that can be investigated using different complementary NMR methodologies to obtain a comprehensive picture of food composition and structure; another approach starts with the specific problem related to a given food (frauds, safety, traceability, geographical and botanical origin, farming methods, food processing, maturation and ageing, etc.) that can be addressed by choosing the most suitable NMR methodology; finally, it is possible to start from a single NMR methodology, developing a broad range of applications to tackle common food-related challenges and different aspects related to foods.

Chondroitin sulfate methacrylate (CS-MA) is a semisynthetic biopolymer increasingly used for the fabrication of chemical hydrogels. In this study, the methacrylation reaction of native CS was carried out with glycidyl methacrylate in dimethyl sulfoxide and optimized to obtain tunable and reproducible methacrylation degrees in a short reaction time. The methacrylation reaction was deeply characterized by mono- and bi-dimensional (1D, 2D) NMR spectroscopy of CS-MA derivatives with different methacrylation degrees. In contrast to what previously reported in the literature, HSQC, HMBC and TOCSY analyses revealed that the methacrylation reaction proceeds via both epoxy ring-opening and transesterification, involving predominantly the primary hydroxyl groups of CS, while preserving sulfate and carboxyl groups of the biopolymer. These findings are of fundamental importance for appropriate and rational design of CS-MA-based biomaterials.

In this study, the effect of several agronomical practices on the chemical composition of hemp inflorescences, a potential novel food that needs to be further studied, was observed. Here, the case study of inflorescences from Ferimon cultivars is discussed and submitted to different agronomical practices (irrigation and fertilizers) in different years, and the inflorescences harvested in different periods were analyzed by a multimethodological approach. Targeted and untargeted methodologies allowed cannabinoids, total phenolic content, metabolite profile and antioxidant activity to be determined. The biomass and inflorescence yields were also reported. The whole data set was submitted to ANOVA-simultaneous component analysis. The statistic results allowed us to observe that irrigation was responsible for the (–)-Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD) increment. THC, cannabichromene (CBC), cannabigerol (CBG), succinate, and fructose resulted as higher in full female flowering than in the period of seed maturity. On the other hand, nitrogen supplementation led to an increase of iso-leucine, valine, and threonine. The obtained results underlined both the potential food application of hemp inflorescences, due to the rich chemical profile, and the strong effect of agronomical practices, mainly irrigation and harvesting, on the qualitative and quantitative characteristics of its metabolite profile.

Down syndrome (DS) is characterized by muscle hypotonia and low muscle strength associated with motor dysfunction. Elucidation of the determinants of muscle weakness in DS would be relevant for therapeutic approaches aimed at treating/mitigating a physical disability with a strong impact on the quality of life in persons with DS. The Ts65Dn mice is a recognized mouse model of DS, with trisomic mice presenting gross motor and muscle phenotypes. The aim of this work was to assess the effect of physical exercise, a well-known tool to improve skeletal muscle condition, in the hindlimbs of trisomic and euploid male mice using quantitative magnetic resonance imaging (MRI). Magnetic resonance spectroscopy (MRS) metabolomics and histological fiber typing were used to further characterize the post-exercise muscle. Quantitative MRI showed not significantly different amounts of skeletal muscle in proximal hindlimbs in trisomic and euploid mice both at baseline and after physical exercise (P>0.05). Similar results were obtained for hindlimbs subfascia adipose tissue, and subcutaneous adipose tissue (P>0.05). MRS showed lower amounts of exercise-related metabolites (valine, isoleucine, leucine) in euploid vs. trisomic mice after exercise (P≤0.05). The percentage of slow-twitch fibers was similar in the two genotypes (P>0.05). We conclude that in DS adapted physical exercise (one month of training) does not induce quantitative changes in skeletal muscle or fiber type composition therein; however, the metabolic response of skeletal muscle to exercise may be affected by trisomy. These findings prompt further research investigating the role of physical exercise as a cue to clarify the mechanisms of the muscular deficit found in DS.

Semen cryopreservation represents the main tool for preservation of biodiversity; however, in avian species, the freezing–thawing process results in a sharp reduction in sperm quality and consequently fertility. Thus, to gain a first insight into the molecular basis of the cryopreservation of turkey sperm, the NMR-assessed metabolite profiles of fresh and frozen–thawed samples were herein investigated and compared with sperm qualitative parameters. Cryopreservation decreased the sperm viability, mobility, and osmotic tolerance of frozen–thawed samples. This decrease in sperm quality was associated with the variation in the levels of some metabolites in both aqueous and lipid sperm extracts, as investigated by NMR analysis. Higher amounts of the amino acids Ala, Ile, Leu, Phe, Tyr, and Val were found in fresh than in frozen–thawed sperm; on the contrary, Gly content increased after cryopreservation. A positive correlation (p < 0.01) between the amino acid levels and all qualitative parameters was found, except in the case of Gly, the levels of which were negatively correlated (p < 0.01) with sperm quality. Other water-soluble compounds, namely formate, lactate, AMP, creatine, and carnitine, turned out to be present at higher concentrations in fresh sperm, whereas cryopreserved samples showed increased levels of citrate and acetyl-carnitine. Frozen–thawed sperm also showed decreases in cholesterol and polyunsaturated fatty acids, whereas saturated fatty acids were found to be higher in cryopreserved than in fresh sperm. Interestingly, lactate, carnitine (p < 0.01), AMP, creatine, cholesterol, and phosphatidylcholine (p < 0.05) levels were positively correlated with all sperm quality parameters, whereas citrate (p < 0.01), fumarate, acetyl-carnitine, and saturated fatty acids (p < 0.05) showed negative correlations. A detailed discussion aimed at explaining these correlations in the sperm cell context is provided, returning a clearer scenario of metabolic changes occurring in turkey sperm cryopreservation.

The application of Nuclear Magnetic Resonance in food science has grown exponentially over the past few decades [...]

Following a similar approach on carvacrol-based derivatives, we investigated the synthesis and the microbiological screening against eight strains of H. pylori, and the cytotoxic activity against human gastric adenocarcinoma (AGS) cells of a new series of ether compounds based on the structure of thymol. Structural analysis comprehended elemental analysis and 1H/13C/19F NMR spectra. The analysis of structure–activity relationships within this molecular library of 38 structurally-related compounds reported that some chemical modifications of the OH group of thymol led to broad-spectrum growth inhibition on all isolates. Preferred substitutions were benzyl groups compared to alkyl chains, and the specific presence of functional groups at para position of the benzyl moiety such as 4-CN and 4-Ph endowed the most anti-H. pylori activity toward all the strains with minimum inhibitory concentration (MIC) values up to 4 µg/mL. Poly-substitution on the benzyl ring was not essential. Moreover, several compounds characterized by the lowest minimum inhibitory concentration/minimum bactericidal concentration (MIC/MBC) values against H. pylori were also tested in order to verify a cytotoxic effect against AGS cells with respect to 5-fluorouracil and carvacrol. Three derivatives can be considered as new lead compounds alternative to current therapy to manage H. pylori infection, preventing the occurrence of severe gastric diseases. The present work confirms the possibility to use natural compounds as templates for the medicinal semi-synthesis.

Beer antioxidants originate mainly from malts, classified as colored, caramel, and roasted, according to the malting process. This study aimed to characterize, in terms of phenolic antioxidants, three types of Pale Ale craft beers brewed using increasing percentage of dark malt (0, 5, and 15% Caraamber malt, called PA100, PA95, PA85, respectively) and to evaluate the impact of dealcoholization by osmotic distillation (OD) on the same antioxidants. All the alcoholic (PA, 6.2-6.8 vol %) and low alcoholic (LA-PA, 1 vol %) beers were analyzed by HPLC-ESI-MS/MS, total phenolic content (TPC), and antioxidant activity (AA): similar phenolic profiles were evidenced and 43 compounds identified or tentatively identified. Some differences were found among PA100, PA95, and PA85: PA85 was richer in free phenolic compounds (10.55 mg/L) and had a higher TPC (463.7 GAE mg/L) and AA (852.1 TE mg/L). LA-PA beers showed the same phenolic profile and similar TPC and AA compared to PA beers; however, there were some differences regarding LA-PA85 (5.91 mg/L). Dealcoholization by OD seemed to weakly affect the phenolic fraction. ESI-MS/MS infusion experiments evidenced oligosaccharides, small organic acids, and amino acids, whose presence was confirmed and quantitated by NMR: besides ethanol and other alcohols, weak to strong loss of low-molecular-weight metabolites was evidenced in LA-PA beers.

Untargeted (NMR) and targeted (RP-HPLC-PDA-ESI-MSn, RP-HPLC-FD) analytical methodologies were used to determine the bioactive components of 19 tea samples, characterized by different production processes (common tea and GABA tea), degrees of fermentation (green and oolong teas), and harvesting season (autumn and spring). The combination of NMR data and a multivariate statistical approach led to a statistical model able to discriminate between GABA and non-GABA teas and green and oolong teas. Targeted analyses showed that green and GABA green teas had similar polyphenol and caffeine contents, but the GABA level was higher in GABA green teas than in regular green tea samples. GABA oolong teas showed lower contents of polyphenols, caffeine, and amino acids, and a higher content of GABA, in comparison with non-GABA oolong teas. In conclusion, the results of this study suggest that the healthy properties of teas, especially GABA teas, have to be evaluated via comprehensive metabolic profiling rather than only the GABA content.

Endive ( Cichorium endivia L.), a vegetable consumed as fresh or packaged salads, is mostly cultivated outdoors and known to be sensitive to waterlogging in terms of yield and quality. Phenotypic, metabolic and transcriptomic analyses were used to study variations in curly- (‘Domari’, ‘Myrna’) and smooth-leafed (‘Flester’, ‘Confiance’) cultivars grown in short-term waterlog due to rainfall excess before harvest. After recording loss of head weights in all cultivars (6-35%), which was minimal in ‘Flester’, NMR untargeted profiling revealed variations as influenced by genotype, environment and interactions, and included drop of total carbohydrates (6–50%) and polyols (3–37%), gain of organic acids (2–30%) and phenylpropanoids (98–560%), and cultivar-specific fluctuations of amino acids (−37 to +15%). The analysis of differentially expressed genes showed GO term enrichment consistent with waterlog stress and included the carbohydrate metabolic process. The loss of sucrose, kestose and inulin recurred in all cultivars and the sucrose-inulin route was investigated by covering over 50 genes of sucrose branch and key inulin synthesis ( fructosyltransferases ) and catabolism ( fructan exohydrolases ) genes. The lowered expression of a sucrose gene subset together with that of SUCROSE:SUCROSE-1-FRUCTOSYLTRANSFERASE ( 1-SST ) may have accounted for sucrose and kestose contents drop in the leaves of waterlogged plants. Two anti-correlated modules harbouring candidate hub-genes, including 1-SST , were identified by weighted gene correlation network analysis, and proposed to control positively and negatively kestose levels. In silico analysis further pointed at transcription factors of GATA, DOF, WRKY types as putative regulators of 1-SST .