Showing papers by "Marco Trevisan published in 2020"

Untargeted metabolomics with multivariate analysis to discriminate hazelnut (Corylus avellana L.) cultivars and their geographical origin

Abstract: Background In the present study a metabolomics-based approach was used to discriminate among different hazelnut cultivars and to trace their geographical origins. Ultra-high-pressure liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry (UHPLC-ESI/QTOF-MS) was used to profile phenolic and sterolic compounds. Results Compounds were identified against an in-house database using accurate monoisotopic mass and isotopic patterns. The screening approach was designed to discern 15 hazelnut cultivars and to discriminate among the geographical origins of six cultivars from the four main growing regions (Chile, Georgia, Italy, and Turkey). This approach allowed more than 1000 polyphenols and sterols to be annotated. The metabolomics data were elaborated with both unsupervised (hierarchical clustering) and supervised (orthogonal projections to latent structures discriminant analysis, OPLS-DA) statistics. These multivariate statistical tools allowed hazelnut samples to be discriminated, considering both 'cultivar type' and 'geographical origin'. Flavonoids (anthocyanins, flavanols and flavonols - VIP scores 1.34-1.49), phenolic acids (mainly hydroxycinnamics - VIP scores 1.35-1.55) together with cholesterol, ergosterol, and stigmasterol derivatives (VIP scores 1.34-1.49) were the best markers to discriminate samples according to geographical origin. Conclusions This work illustrates the potential of untargeted profiling of phenolics and sterols based on UHPLC-ESI/QTOF mass spectrometry to discriminate hazelnut and support authenticity and origin. © 2019 Society of Chemical Industry.

Extending the concept of terroir from grapes to other agricultural commodities: an overview

Abstract: The term terroir is widely used in oenology to link geographical origin and environmental conditions used for grapes training with the sensorial and hedonistic value of the wine. Overall, this definition includes the pedo-climatic (such as soil type and climate) as well as agronomic practices able to affect the composition of a plant-derived food. This concept is rather underestimated in matrices other than wine, even though recent advance in analytics is providing clear evidence that the organoleptic and nutritional value of foods is strongly affected by the area of cultivation. This mini review reports the molecular basis of the effect of terroir on phytochemical profile and highlights the need to extend this concept from grape to other crops and food products, thus linking geographical origin and quality of agricultural commodities.

Impact of a Pitanga Leaf Extract to Prevent Lipid Oxidation Processes during Shelf Life of Packaged Pork Burgers: An Untargeted Metabolomic Approach

Abstract: In this work, the comprehensive metabolomic changes in pork burgers treated with different antioxidants, namely, (a) a control without antioxidants, (b) 200 mg/kg butylated hydroxytoluene (BHT), and (c) 250 mg/kg pitanga leaf extract (PLE, from Eugenia uniflora L.), each one packaged under modified atmosphere (80% O2 and 20% CO2) for 18 days storage at 2 ± 1 °C, were deeply studied. In particular, untargeted metabolomics was used to evaluate the impact of the antioxidant extracts on meat quality. The PLE phytochemical profile revealed a wide variety of antioxidant compounds, such as polyphenols, alkaloids, and terpenoids. Multivariate statistics (both unsupervised and supervised) allowed to observe marked differences in BHT and PLE burgers metabolomic profiles during storage. Most of the differences could be attributed to hexanoylcarnitine, 4-hydroxy-2-nonenal, 6-hydroxypentadecanedioic acid, 9S,11S,15S,20-tetrahydroxy-5Z,13E-prostadienoic acid (20-hydroxy-PGF2a), sativic acid, followed by glycerophospholipids. In addition, significant correlations (p < 0.01) were observed between thiobarbituric acid reactive substances and metabolites related to lipid oxidation processes. Therefore, the approach used showed a clear modulation of lipid oxidation, likely promoted by the plant leaf extract, thus confirming the ability of PLE to delay lipid oxidative phenomena during storage.

Isolation and Screening of Extracellular PGPR from the Rhizosphere of Tomato Plants after Long-Term Reduced Tillage and Cover Crops.

Abstract: Plant growth promoting rhizobacteria provide an innovative solution to address challenges in sustainable agro-ecosystems, improving plant growth as well as acting as agents of biocontrol. In this study autochthonous bacteria were isolated from the rhizosphere of processing tomato plants (Solanum lycopersicum L.) cultivated with conservation agriculture practices (i.e., reduced tillage and cover crops), and evaluated for both growth-promoting activities (PGPAs), and antagonistic potential against the phytopathogenic pest Sclerotinia sclerotiorum. Considering the several activities of PGPR, we decided to structure the screening with a hierarchic approach, starting from testing the capability of fixing nitrogen. The obtained bacteria were processed through the molecular typing technique rep-PCR (Repetitive Extragenic Palindromic) in order to discriminate microbial strains with the same profiles, and identified via 16S rDNA sequencing. Thirty-eight selected isolates were screened in vitro for different activities related to plant nutrition and plant growth regulation as well as for antifungal traits. Isolated bacteria were found to exhibit different efficiencies in indoleacetic acid production and siderophore production, phosphate solubilization and biocontrol activity against the widespread soil-borne plant pathogen S. sclerotiorum. All the 38 bacterial isolates showed at least one property tested. With a view to detect the suitable candidates to be developed as biofertilizers, the selected isolates were ranked by their potential ability to function as PGPR. Thus, consortium of native PGPR bacteria inoculants may represent a suitable solution to address the challenges in sustainable agriculture, to ensure crop yield and quality, lowering the application of chemicals input.

Metabolomic Study to Evaluate the Transformations of Extra-Virgin Olive Oil's Antioxidant Phytochemicals During In Vitro Gastrointestinal Digestion.

Abstract: In this work, different commercial extra-virgin olive oils (EVOO) were subjected to in vitro gastrointestinal digestion and the changes in bioactive compounds were evaluated by ultra-high-pressure liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry, using untargeted metabolomics. As expected, raw EVOO samples were abundant in total sterols (on average: 3007.4 mg equivalents/kg) and tyrosol equivalents (on average: 334.1 mg equivalents/kg). However, the UHPLC-QTOF screening allowed us to annotate 310 compounds, with a large abundance of sterols (219 compounds), followed by polyphenols (67 compounds) and terpenoids. The in vitro gastrointestinal digestion was found to affect the phytochemical composition of the different EVOO samples. In particular, both unsupervised and supervised statistics depicted the modifications of the bioactive profile following gastric and pancreatic phases. Overall, the compounds which resulted as the most affected by the in vitro digestion were flavonoids (cyanidin and luteolin equivalents), whilst relatively high % bioaccessibility values were recorded for tyrosol equivalents during the pancreatic phase (on average, 66%). In this regard, oleuropein-aglycone (i.e., the major phenolic compound in EVOO) was converted to hydroxytyrosol, moving from an average value of 1.3 (prior to the in vitro digestion) up to 9.7 mg equivalents/kg during the pancreatic step. As proposed in the literature, the increase in hydroxytyrosol might be the result of the combined effect of lipase(s) activity and acidic conditions. Taken together, the present findings corroborate the suitability of untargeted metabolomics coupled to in vitro digestion methods to investigate the bioaccessibility of phenolic compounds. In this regard, a significant impact of in vitro gastrointestinal digestion on polyphenolic profiles has been detected, thus suggesting the need to account for actual bioaccessibility values rather than just considering the amounts in the raw commodity.

Identification of markers of sensory quality in ground coffee: an untargeted metabolomics approach.

Abstract: In the last years, consumers increased the demand for high-quality and healthy beverages, including coffee. To date, among the techniques potentially available to determine the overall quality of coffee beverages, metabolomics is emerging as a valuable tool. In this study, 47 ground coffee samples were selected during the 2018 Edition of the “International coffee tasting” (ICT) in order to provide discrimination based on both chemical and sensory profiles. In particular, 20 samples received a gold medal (“high quality” group), while lower sensory scores characterized 27 samples (without medal). Untargeted metabolomics based on ultra-high pressure liquid chromatography coupled with quadrupole-time-of-flight (UHPLC-QTOF) and head space-gas chromatography coupled with mass spectrometry platforms followed by multivariate statistical approaches (i.e., both supervised and unsupervised) were used to provide new insight into the searching of potential markers of sensory quality. Several compounds were identified, including polyphenols, alkaloids, diazines, and Maillard reaction products. Also, the headspace/GC-MS highlighted the most important volatile compounds. Polyphenols were scarcely correlated to the sensory parameters, whilst the OPLS-DA models built using typical coffee metabolites and volatile/Maillard compounds possessed prediction values > 0.7. The “high quality” group showed specific metabolomic signatures, thus corroborating the results from the sensory analysis. Overall, methyl pentanoate (ROC value = 0.78), 2-furfurylthiol (ROC value = 0.75), and L-Homoserine (ROC value = 0.74) established the higher number of significant (p < 0.05) correlations with the sensory parameters. Although ad-hoc studies are advisable to further confirm the proposed markers, this study demonstrates the suitability of untargeted metabolomics for evaluating coffee quality and the potential correlations with the sensory attributes.

New insights in the allelopathic traits of different barley genotypes: Middle Eastern and Tibetan wild-relative accessions vs. cultivated modern barley.

Abstract: The two alkaloids gramine and hordenine have been known for playing a role in the allelopathic ability in barley (Hordeum vulgare L.). These allelochemicals can be both found in leaves and roots in some barley cultivars whereas in others one seems to exclude the other. In this study eighteen accessions of barley from the Middle-East area, one accession from Tibet and the modern spring cultivar Barke, already used as parental donor in a nested associated mapping (NAM) population, were screened for their gramine, hordenine and N-methyltyramine (the direct precursor of hordenine) content in leaves, roots and exudates. Moreover, the toxicity of the three allelochemicals on root growth inhibition on lettuce (Lactuca sativa L.) was evaluated. Results of this study showed the preferential production of gramine and hordenine in leaves and roots, respectively, in the nineteen barley accessions. On the other hand, in the modern barley cultivar Barke, the highest content of hordenine in roots and the general lack of gramine suggests a favored biosynthesis of the former. Gramine was not detected in the root exudates. In additions, different metabolomic profiles were observed in wild relatives compared to modern barley genotypes. The results also showed the phytotoxic effects of the three compounds on root growth of lettuce seedlings, with a reduction in root length and an increase of root surface area and diameter. In conclusion, this study highlighted the impact of the domestication effects on the production and distribution of the two allelopathic alkaloids gramine and hordenine in barley.

Nutraceutical Profiles of Two Hydroponically Grown Sweet Basil Cultivars as Affected by the Composition of the Nutrient Solution and the Inoculation With Azospirillum brasilense.

Abstract: Sweet basil (Ocimum basilicum L.) is one of the most produced aromatic herbs in the world, exploiting hydroponic systems. It has been widely assessed that macronutrients, like nitrogen (N) and sulfur (S), can strongly affect the organoleptic qualities of agricultural products, thus influencing their nutraceutical value. In addition, plant-growth-promoting rhizobacteria (PGPR) have been shown to affect plant growth and quality. Azospirillum brasilense is a PGPR able to colonize the root system of different crops, promoting their growth and development and influencing the acquisition of mineral nutrients. On the bases of these observations, we aimed at investigating the impact of both mineral nutrients supply and rhizobacteria inoculation on the nutraceutical value on two different sweet basil varieties, i.e., Genovese and Red Rubin. To these objectives, basil plants have been grown in hydroponics, with nutrient solutions fortified for the concentration of either S or N, supplied as SO42- or NO3-, respectively. In addition, plants were either non-inoculated or inoculated with A. brasilense. At harvest, basil plants were assessed for the yield and the nutraceutical properties of the edible parts. The cultivation of basil plants in the fortified nutrient solutions showed a general increasing trend in the accumulation of the fresh biomass, albeit the inoculation with A. brasilense did not further promote the growth. The metabolomic analyses disclosed a strong effect of treatments on the differential accumulation of metabolites in basil leaves, producing the modulation of more than 400 compounds belonging to the secondary metabolism, as phenylpropanoids, isoprenoids, alkaloids, several flavonoids, and terpenoids. The primary metabolism that resulted was also influenced by the treatments showing changes in the fatty acid, carbohydrates, and amino acids metabolism. The amino acid analysis revealed that the treatments induced an increase in arginine (Arg) content in the leaves, which has been shown to have beneficial effects on human health. In conclusion, between the two cultivars studied, Red Rubin displayed the most positive effect in terms of nutritional value, which was further enhanced following A. brasilense inoculation.

Changes in physiological activities and root exudation profile of two grapevine rootstocks reveal common and specific strategies for Fe acquisition.

Abstract: In several cultivation areas, grapevine can suffer from Fe chlorosis due to the calcareous and alkaline nature of soils. This plant species has been described to cope with Fe deficiency by activating Strategy I mechanisms, hence increasing root H+ extrusion and ferric-chelate reductase activity. The degree of tolerance exhibited by the rootstocks has been reported to depend on both reactions, but to date, little emphasis has been given to the role played by root exudate extrusion. We studied the behaviour of two hydroponically-grown, tolerant grapevine rootstocks (Ramsey and 140R) in response to Fe deficiency. Under these experimental conditions, the two varieties displayed differences in their ability to modulate morpho-physiological parameters, root acidification and ferric chelate reductase activity. The metabolic profiling of root exudates revealed common strategies for Fe acquisition, including ones targeted at reducing microbial competition for this micronutrient by limiting the exudation of amino acids and sugars and increasing instead that of Fe(III)-reducing compounds. Other modifications in exudate composition hint that the two rootstocks cope with Fe shortage via specific adjustments of their exudation patterns. Furthermore, the presence of 3-hydroxymugenic acid in these compounds suggests that the responses of grapevine to Fe availability are rather diverse and much more complex than those usually described for Strategy I plants.

Nutritional inter-dependencies and a carbazole-dioxygenase are key elements of a bacterial consortium relying on a Sphingomonas for the degradation of the fungicide thiabendazol

Abstract: Background: Thiabendazole (TBZ), is a benzimidazole fungicide and anthelminthic whose high persistence and toxicity pose a serious environmental threat. In our quest for environmental mitigation we previously isolated the first TBZ-degrading bacterial consortium and provided preliminary evidence for its composition and the degrading role of a Sphingomonas. Here, we employed a multi-omic approach combined with DNA-stable isotope probing (SIP) to determine the genetic make-up of the key consortium members, to disentangle nutritional and metabolic interdependencies, to identify the transformation pathway of TBZ and to understand the genetic network driving its transformation. Results: Time-series SIP in combination with amplicon sequencing analysis verified the key role of Sphingomonas in TBZ degradation by assimilating over 80% of the 13C-labelled phenyl moiety of TBZ. Non-target mass spectroscopy (MS) analysis showed the accumulation of thiazole-4-carboxamidine as a single dead-end transformation product and no phenyl-containing derivative, in line with the phenyl moiety assimilation in the SIP analysis. Time series metagenomic analysis of the consortium supplemented with TBZ or succinate led to the assembly of 18 metagenome-assembled genomes (MAGs) with >80% completeness, six (Sphingomonas 3X21F, γ-Proteobacterium 34A, Bradyrhizobiaceae 9B and Hydrogenophaga 19A, 13A, and 23F) being dominant. Meta-transcriptomic and -proteomic analysis suggested that Sphingomonas mobilize a carbazole dioxygenase (car) operon during the initial cleavage of TBZ to thiazole-4-carboxamidine and catechol, the latter is further transformed by enzymes encoded in a catechol ortho-cleavage (cat) operon; both operons being up-regulated during TBZ degradation. Computational docking analysis of the terminal oxygenase component of car, CarAa, showed high affinity to TBZ, comparable to carbazole, reinforcing its high potency for TBZ transformation. These results suggest no interactions between consortium members in TBZ transformation, performed solely by Sphingomonas. In contrast, gene expression network analysis revealed strong interactions between Sphingomonas MAG 3X12F and Hydrogenophaga MAG 23F, with Hydrogenophaga activating its cobalamin biosynthetic pathway and Sphingomonas its cobalamin salvage pathway along TBZ degradation. Conclusions: Our findings suggest interactions between consortium members which align with the "black queen hypothesis": Sphingomonas detoxifies TBZ, releasing consortium members by a toxicant; in return for this, Hydrogenophaga 23F provides cobalamin to the auxotrophic Sphingomonas.