Genetic study of Camelina sativa oilseed crop and selection of a new variety by the bulk method.

Camelina sativa, commonly referred to as camelina or false flax, has emerged as a promising cover crop with the potential to mitigate climate change-a pressing global challenge that demands urgent and sustainable solutions. Belonging to the Brassicaceae family and native to Europe and Central Asia, camelina is an oilseed crop known for its resilience in diverse climates, including arid and semi-arid regions, making it adaptable to various environments. A breeding program started from a study of six winter varieties and five spring varieties of camelina is described: these genetic materials were characterized by SSRs molecular markers and by GBS technique. Molecular data clearly showed all spring varieties were genetically similar and distinguishable from the winter varieties, which, in turn, clustered together. Using molecular data, parental varieties belonging to the two different clusters were selected to generate new genetic variability. The new variety obtained, selected through the bulk method based on three parameters: yield, earliness, and weight of 1000 seeds, has allowed the generation of the new genetic material provisionally named C1244. Chemical characterization was performed (bromatological and glucosinolates analysis) to better describe C1244 in comparison with benchmark varieties. The new variety exhibited early maturity, similar to spring varieties, making this genetic material promising for use in intercropping systems, a high weight of 1000 seeds (1.46 g) which improves and facilitates seeding/harvesting operations and a high oil content (33.62%) akin to winter varieties making it valuable for human and animal food purposes.

Biochemical Characterization of the Seed Quality of a Collection of White Lupin Landraces from Southern Italy.

Lupin species provide essential nutrients and bioactive compounds. Within pulses, they have one of the highest contents of proteins and fibers and are among the poorest in carbohydrates. The Mediterranean region is an important cradle area of the origin and domestication of cultivated white lupin (Lupinus albus L.). In this work, we present the characterization of 19 white lupin landraces collected from several sites in southern Italy, characterized by different pedoclimatic conditions. The protein contents and electrophoretic patterns, total polyphenols, phytic acid, lipids and phosphorous content, and reducing and anti-tryptic activities have been determined for each landrace. The relationships of the compositional characteristics, the area of origin of landraces and between compositional characteristics and thermo-pluviometric trends that occurred in the genotype comparison field during the two-year period between 2019 and 2020 are compared and discussed. From a nutritional point of view, some of the analyzed landraces differ from the commercial reference. The panel of molecular analyses performed can help in building an identity card for the grain to rapidly identify those varieties with the desired characteristics.

Protein Concentration Affects the Food Allergen γ-Conglutin Uptake and Bacteria-Induced Cytokine Production in Dendritic Cells.

γ-Conglutin (γ-C) from lupin seeds has been identified as a potent allergen with cross reactivity to peanuts. Here, we investigated how γ-C affected the response in bone marrow-derived dendritic cells (DCs) to bacterial stimuli. γ-C enhanced L. acidophilus NCFM (LaNCFM)-induced IL-12, IL-10, and IL-23 dose-dependently. In contrast, together with E. coli Nissle or LPS, γ-C reduced the production of IL-12 but not of IL-23 and IL-10. Enzyme-hydrolyzed γ-C also enhanced LaNCFM-induced IL-12 and IL-23 production. All preparations induced ROS production in the DCs. The mannose receptor ligands mannan and dextran and the clathrin inhibitor monodansylcadaverine partly inhibited the endocytosis of γ-C. Kunitz trypsin inhibitor and the scavenger receptor ligand polyG also enhanced LaNCFM-induced IL-12, indicating the involvement of receptors other than C-type lectin receptors. The endocytosis of labeled γ-C increased dose-dependently by addition of unlabeled γ-C, which coincided with γ-C's tendency to aggregate. Taken together, γ-C aggregation affects endocytosis and affects the cytokine production induced by gram-positive and gram-negative bacteria differently. We suggest that γ-C is taken up by the same mechanism as other food proteins but due to aggregation is present in higher concentration in the DCs. This could influence the resulting T-cell response in a microbial stimuli-dependent way.

A not-glycosylated isoform of γ-conglutin, a hexameric glycoprotein of Lupinus albus seed, participates in the oligomerization equilibrium.

γ-conglutin (γ-C) is a hexameric glycoprotein accumulated in lupin seeds and has long been considered as a storage protein. Recently, it has been investigated for its possible postprandial glycaemic regulating action in human nutrition and for its physiological role in plant defence. The quaternary structure of γ-C results from the assembly of six monomers in reversible pH-dependent association/dissociation equilibrium. Our working hypothesis was that the γ-C hexamer is made up of glycosylated subunits in association with not-glycosylated isoforms, that seem to have 'escaped' the correct glycosylation process in the Golgi. Here we describe the isolation of not-glycosylated γ-C monomers in native condition by two in tandem lectin-based affinity chromatography and the characterization of their oligomerization capacity. We report, for the first time, the observation that a plant multimeric protein may be formed by identical polypeptide chains that have undergone different post-translational modifications. All obtained considered, the results strongly suggest that the not-glycosylated isoform can also take part in the oligomerization equilibrium of the protein.

Binary Alginate-Whey Protein Hydrogels for Antioxidant Encapsulation.

Encapsulation of antioxidants in hydrogels, i.e., three-dimensional networks that retain a significant fraction of water, is a strategy to increase their stability and bioaccessibility. In fact, low oxygen diffusivity in the viscous gelled phase decreases the rate of oxidation. Moreover, some hydrocolloids such as alginate and whey proteins provide a pH-dependent dissolution mechanism, allowing the retention of encapsulated compounds in the gastric environment and their release in the intestine, where they can be absorbed. This paper reviews the information on alginate-whey protein interactions and on the strategies to use binary mixtures of these polymers for antioxidant encapsulation. Results showed that alginate and whey proteins strongly interact, forming hydrogels that can be modulated by alginate molecular mass, mannuronic acid: guluronic acid ratio, pH, Ca2+ or transglutaminase addition. Hydrogels of alginate and whey proteins, in the forms of beads, microparticles, microcapsules, and nanocapsules, generally provide better encapsulation efficiency and release properties for antioxidants with respect to the hydrogel of alginate alone. The main challenges for future studies are to extend knowledge on the interactions among three components, namely alginate, whey proteins, and the encapsulated bioactive compounds, and to investigate the stability of these structures under food processing conditions. This knowledge will represent the rationale basis for the development of structures that can be tailored to specific food applications.

Lupinus albus γ-Conglutin: New Findings about Its Action at the Intestinal Barrier and a Critical Analysis of the State of the Art on Its Postprandial Glycaemic Regulating Activity.

γ-Conglutin (γ-C) is the glycoprotein from the edible seed L. albus, studied for long time for its postprandial glycaemic regulating action. It still lacks clear information on what could happen at the meeting point between the protein and the organism: the intestinal barrier. We compared an in vitro system involving Caco-2 and IPEC-J2 cells with an ex vivo system using pig ileum and jejunum segments to study γ-C transport from the apical to the basolateral compartment, and its effects on the D-glucose uptake and glucose transporters protein expression. Finally, we studied its potential in modulating glucose metabolism by assessing the possible inhibition of α-amylase and α-glucosidase. RP-HPLC analyses showed that γ-C may be transported to the basolateral side in the in vitro system but not in the pig intestines. γ-C was also able to promote a decrease in glucose uptake in both cells and jejunum independently from the expression of the SGLT1 and GLUT2 transporters.

Antifungal Activity and Biochemical Profiling of Exudates from Germinating Maize Nostrano di Storo Local Variety.

Plant pathogens are responsible for important damages to valuable crops causing important economic losses. Agrobiodiversity protection is crucial for the valorization of local varieties that could possess higher resistance to biotic and abiotic stress. At the beginning of germination, seeds are susceptible to pathogens attacks, thus they can release endogenous antimicrobial compounds of different natures in the spermosphere, to contrast proliferation of microorganisms. The work aimed at characterizing the maize of local variety Nostrano di Storo seed exudates secreted during the first phases of germination, to identify compounds active in the defense towards pathogens. Storo seed exudates were proven to inhibit F. verticilloides germination. In order to investigate the cause of the described effect, compositional profiling of the exudates was performed through NMR, lipidomic, and proteomic analyses. This study suggests an important role of microbial endophytic communities in the protection of the seed during the early phases of the germination process and their interplay with fatty acids released by the seeds, rather than a specific antifungal compound. The valorization of agronomically acceptable maize lines with pre-harvest enhanced resistances to pathogens contamination could lead, in the near future, to commercially available varieties potentially requiring more limited chemical protective treatments.

Digestive and gastroprotective effects of Achillea erba-rotta subsp. moschata (Wulfen) I.Richardson (syn. A. moschata Wulfen) (Asteraceae): From traditional uses to preclinical studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Achillea erba-rotta subsp. moschata (Wulfen) I.Richardson (syn. A. moschata Wulfen) (Asteraceae) is an alpine endemic plant whose aerial parts are harvested by the locals mainly for the digestive properties. Despite its widespread use, few studies have been conducted to date to verify its bioactivity. AIM OF THE STUDY: The purpose of the work was to meet the tradition confirming with experimental data the popular belief that the consumption of this species offers beneficial effects to the gastrointestinal system. MATERIALS AND METHODS: Using Soxhlet apparatus, the dried aerial parts of A. erba-rotta subsp. moschata were successively extracted with petroleum ether (PET), dichloromethane (DCM) and methanol (MeOH). The essential oil (EO) was obtained by hydrodistillation using a Clevenger apparatus while infusion (AE) was prepared following the traditional local recipe. Their chemical characterization was performed by various techniques including SPME-GC/MS, GC/MS and HPLC/MS-MS. An in vitro biological screening was carried out. The influence of AE on lipid digestion was monitored by titration of free fatty acids (FFA) during pancreatic lipase activity with the pH-stat method. For all extracts and EO, the anti-Helicobacter pylori activity was assessed by the broth microdilution method, the influence on cell viability was evaluated against NCI-N87, OE21 and Caco-2 cell lines and a preliminary toxicity evaluation was done using Brine Shrimp lethality (BSL) assay. The anti-inflammatory potential was evidenced by interleukin IL-1- induced IL8 expression on Caco-2 cells. RESULTS: AE increased by 15% the FFA releasing compared to the pancreatic lipase alone. PET, DCM and MeOH extracts as well as AE and EO were considered active against the growth of both antimicrobial susceptible and resistant strains of H. pylori with MIC values starting from 16 µg/mL. PET and DCM (IC50 = 89 µg/mL and 96 µg/mL, respectively, against Caco-2 cell line) extracts showed the high effect on cell viability while the EO reduced in 50% of cell viability at 1.48 µL/mL (NCI-N87 cells), 1.42 µL/mL (OE21 cells), and 3.44 µL/mL (Caco-2 cells) corroborating the BSL results. In different degrees, all extracts and EO inhibited the IL-1ß-stimulated IL-8 production in Caco-2 cells. CONCLUSIONS: The obtained data are encouraging and provide a scientific basis for the traditional use of A. erba-rotta subsp. moschata as a digestive agent although they need to be further corroborated by studies involving the investigation of both the in vivo activities and the role of the compounds detected in the extracts.

Thermal Shift Assay as a Tool to Evaluate the Release of Breakdown Peptides from Cowpea ß-Vignin during Seed Germination.

The present work aimed to characterize the molecular relationships between structure and function of the seed storage protein ß-vignin, the vicilin storage protein of cowpea (Vigna unguiculata, l. Walp) seeds. The molecular characterization of ß-vignin was carried out firstly by assessing its thermal stability, under different conditions of pH and ionic strength, by thermal shift assay (TSA) using SYPRO Orange fluorescent dye. Secondly, its aggregation propensity was evaluated using a combination of chromatographic and electrophoretic techniques. Two forms of ß-vignin were considered: the native form purified from mature quiescent seeds, and a stable breakdown intermediate of 27 kDa produced while seeds germinate. TSA is a useful tool for determining and following over time the structural changes that occur to the protein during germination. The main result was the molecular characterization of the 27 kDa intermediate breakdown polypeptide, which, to the best of our knowledge, has never been described before. ß-vignin seems to retain its trimeric conformation despite the evident degradation of its polypeptides.

Polyphenol bioactivity evolution during the spontaneous fermentation of vegetal by-products.

Food industry by-products such as grape pomace (GP), tomato pomace (TP), and spent coffee grounds (SCG) are rich in polyphenols (PP) but are easily biodegradable. The aim of this study is to test Spontaneous Fermentation (SF) as treatment to modify PP profile and bioactivity. The results highlighted that the by-products' organic matter and the microbial populations drove the SF evolution; heterolactic, alcoholic, and their mixtures were the predominant metabolisms of TP, GP, and SCG + GP co-fermentation. Increases in the extractable amounts and antiradical activity occurred for all the biomasses. Regarding the aglycate-PPs (APP), i.e. the most bioreactive PPs, significant changes occurred for TP and GP but did not influence the anti-inflammatory bioactivity. The co-fermentation increased significantly chlorogenic acid and consumed most of the APPs, acting as a purification system to obtain a highly concentrated APP fraction, so that the extract might be employed for a specific purpose.

Valorization of Okara by Enzymatic Production of Anti-Fungal Compounds for Plant Protection.

Okara is a soybean transformation agri-food by-product, the massive production of which currently poses severe disposal issues. However, its composition is rich in seed storage proteins, which, once extracted, can represent an interesting source of bioactive peptides. Antimicrobial and antifungal proteins and peptides have been described in plant seeds; thus, okara is a valuable source of compounds, exploitable for integrated pest management. The aim of this work is to describe a rapid and economic procedure to isolate proteins from okara, and to produce an enzymatic proteolyzed product, active against fungal plant pathogens. The procedure allowed the isolation and recovery of about 30% of okara total proteins. Several proteolytic enzymes were screened to identify the proper procedure to produce antifungal compounds. Antifungal activity of the protein digested for 24 h with pancreatin against Fusarium and R. solani mycelial growth and Pseudomonas spp was assessed. A dose-response inhibitory activity was established against fungi belonging to the Fusarium genus. The exploitation of okara to produce antifungal bioactive peptides has the potential to turn this by-product into a paradigmatic example of circular economy, since a field-derived food waste is transformed into a source of valuable compounds to be used in field crops protection.

Bioactivities of Pseudocereal Fractionated Seed Proteins and Derived Peptides Relevant for Maintaining Human Well-Being.

Food proteins and peptides are able to exert a variety of well-known bioactivities, some of which are related to well-being and disease prevention in humans and animals. Currently, an active trend in research focuses on chronic inflammation and oxidative stress, delineating their major pathogenetic role in age-related diseases and in some forms of cancer. The present study aims to investigate the potential effects of pseudocereal proteins and their derived peptides on chronic inflammation and oxidative stress. After purification and attribution to protein classes according to classic Osborne's classification, the immune-modulating, antioxidant, and trypsin inhibitor activities of proteins from quinoa (Chenopodium quinoa Willd.), amaranth (Amaranthus retroflexus L.), and buckwheat (Fagopyrum esculentum Moench) seeds have been assessed in vitro. The peptides generated by simulated gastro-intestinal digestion of each fraction have been also investigated for the selected bioactivities. None of the proteins or peptides elicited inflammation in Caco-2 cells; furthermore, all protein fractions showed different degrees of protection of cells from IL-1ß-induced inflammation. Immune-modulating and antioxidant activities were, in general, higher for the albumin fraction. Overall, seed proteins can express these bioactivities mainly after hydrolysis. On the contrary, higher trypsin inhibitor activity was expressed by globulins in their intact form. These findings lay the foundations for the exploitation of these pseudocereal seeds as source of anti-inflammatory molecules.

Chromatography-Independent Fractionation and Newly Identified Molecular Features of the Adzuki Bean (Vigna angularis Willd.) ß-vignin Protein.

Adzuki seed ß-vignin, a vicilin-like globulin, has proven to exert various health-promoting biological activities, notably in cardiovascular health. A simple scalable enrichment procedure of this protein for further nutritional and functional studies is crucial. In this study, a simplified chromatography-independent protein fractionation procedure has been optimized and described. The electrophoretic analysis showed a high degree of homogeneity of ß-vignin isolate. Furthermore, the molecular features of the purified protein were investigated. The adzuki bean ß-vignin was found to have a native size of 146 kDa, and the molecular weight determined was consistent with a trimeric structure. These were identified in two main polypeptide chains (masses of 56-54 kDa) that are glycosylated polypeptides with metal binding capacity, and one minor polypeptide chain with a mass 37 kDa, wherein these features are absent. The in vitro analysis showed a high degree of digestibility of the protein (92%) and potential anti-inflammatory capacity. The results lay the basis not only for further investigation of the health-promoting properties of the adzuki bean ß-vignin protein, but also for a possible application as nutraceutical molecule.

Chemical Composition, Tocopherol and Carotenoid Content of Seeds from Different Andean Lupin (Lupinus mutabilis) Ecotypes.

Andean lupin (Lupinus mutabilis) seeds are appreciated for their high protein and lipid contents and have potential applications as ingredients in food, cosmetic, and pharmaceutical industries. Nevertheless, the information about the seed composition (especially in lipophilic antioxidants) of ecotypes from distinct cropping areas is currently limited. Thus, the aim of the present research was to assess the morphological characteristics, chemical composition, tocopherol and carotenoid contents of the seeds of 33 Andean lupin ecotypes from different Peruvian regions, along with three L. albus, one L. angustifolius and one L. luteus controls. Significant differences were noted among the Andean ecotypes for all analyzed features. The protein, lipid and ash contents were 32.0-46.9, 13.6-18.6 and 2.7-4.4 g/100 g dry matter (DM), respectively. The seeds were rich in tocopherols (172.1-249.8 mg/kg DM; γ-tocopherol was 98% of total tocols) and low in carotenoids (0.69-2.89 mg/kg DM). Debittering increased the tocopherol content (227.0-378.2 mg/kg DM), probably because of the soluble components loss, although the carotenoid concentration remained unchanged. The Andean lupins had higher protein, lipid and tocopherol contents than L. albus and L. angustifolius; the L. luteus values were within the L. mutabilis range. These results suggest that L. mutabilis harbors nutritional characteristics that are well suited to modern food trends.

Lupinus albus γ-Conglutin, a Protein Structurally Related to GH12 Xyloglucan-Specific Endo-Glucanase Inhibitor Proteins (XEGIPs), Shows Inhibitory Activity against GH2 ß-Mannosidase.

γ-conglutin (γC) is a major protein of Lupinus albus seeds, but its function is still unknown. It shares high structural similarity with xyloglucan-specific endo-glucanase inhibitor proteins (XEGIPs) and, to a lesser extent, with Triticum aestivum endoxylanase inhibitors (TAXI-I), active against fungal glycoside hydrolases GH12 and GH11, respectively. However, γC lacks both these inhibitory activities. Since ß-galactomannans are major components of the cell walls of endosperm in several legume plants, we tested the inhibitory activity of γC against a GH2 ß-mannosidase (EC 3.2.1.25). γC was actually able to inhibit the enzyme, and this effect was enhanced by the presence of zinc ions. The stoichiometry of the γC/enzyme interaction was 1:1, and the calculated Ki was 1.55 µM. To obtain further insights into the interaction between γC and ß-mannosidase, an in silico structural bioinformatic approach was followed, including some docking analyses. By and large, this work describes experimental findings that highlight new scenarios for understanding the natural role of γC. Although structural predictions can leave space for speculative interpretations, the full complexity of the data reported in this work allows one to hypothesize mechanisms of action for the basis of inhibition. At least two mechanisms seem plausible, both involving lupin-γC-peculiar structures.

Characterization of Chenopodin Isoforms from Quinoa Seeds and Assessment of Their Potential Anti-Inflammatory Activity in Caco-2 Cells.

Several food-derived molecules, including proteins and peptides, can show bioactivities toward the promotion of well-being and disease prevention in humans. There is still a lack of information about the potential effects on immune and inflammatory responses in mammalian cells following the ingestion of seed storage proteins. This study, for the first time, describes the potential immunomodulation capacity of chenopodin, the major protein component of quinoa seeds. After characterizing the molecular features of the purified protein, we were able to separate two different forms of chenopodin, indicated as LcC (Low charge Chenopodin, 30% of total chenopodin) and HcC (High charge Chenopodin, 70% of total chenopodin). The biological effects of LcC and HcC were investigated by measuring NF-κB activation and IL-8 expression studies in undifferentiated Caco-2 cells. Inflammation was elicited using IL-1ß. The results indicate that LcC and HcC show potential anti-inflammatory activities in an intestinal cell model, and that the proteins can act differently, depending on their structural features. Furthermore, the molecular mechanisms of action and the structural/functional relationships of the protein at the basis of the observed bioactivity were investigated using in silico analyses and structural predictions.

Antioxidant and Anti-Inflammatory Activities of the Crude Extracts of Raw and Fermented Tomato Pomace and Their Correlations with Aglycate-Polyphenols.

Two tomato pomace (TP) were studied as feedstocks to obtain extracts that are rich in polyphenols. TPs prompt degradation impairs biomass safety, thus naturally present microflora were tested to perform conservation, and own lactic bacteria became predominant after 60 days of treatment. The extracts of TPs and TPs fermented (TPF) were chemically characterized and tested for antioxidant and anti-inflammatory activities. Flavonoids and phenolic acids were classed as aglycone-polyphenols (A-PP), the most bioactive polyphenol fraction. Fermentation led to a reduction of the A-PP amount, but no significant change in composition. Antioxidant power increased, despite the A-PP reduction, for the presence of fermentation metabolites having aromatic-substituent. TP and TPF both have anti-inflammatory properties that were strictly dependent upon the A-PP content. Fermentation preserved the anti-inflammatory activity and the Partial Least Square (PLS) identified as the most active molecules naringenin chalcone, kaempferol, gallic acid, and cinnamic acid, together with the definition of the active dose.

Bio-Functional and Structural Properties of Pasta Enriched with a Debranning Fraction from Purple Wheat.

A colored and fiber-rich fraction from the debranning of purple wheat was incorporated at 25% into semolina- and flour-based pasta produced on a pilot-plant scale, with the aim of increasing anthocyanin and total phenolic content with respect to pasta obtained from whole pigmented grains. The debranning fraction impaired the formation of disulfide-stabilized protein networks in semolina-based systems. Recovery of phenolics was impaired by the pasta making process, and cooking decreased the phenolic content in both enriched samples. Cooking-related losses in anthocyanins and total phenolics were similar, but anthocyanins in the cooked semolina-based pasta were around 20% of what was expected from the formulation. HPLC (High Performance Liquid Chromatography) profiling of phenolics was carried out on extracts from either type of enriched pasta both before and after cooking and indicate possible preferential retention of specific compounds in each type of enriched pasta. Extracts from cooked samples of either enriched pasta were tested as inhibitors of enzymes involved in glucose metabolism and uptake, as well as for their capacity of suppressing the response to inflammatory stimuli. Results of both biological tests indicate that the phenolics in extracts from both cooked pasta samples had inhibitory capacities higher than extracts of the original debranning fraction at identical concentrations of total bioactives.

Effects on the Caco-2 Cells of a Hypoglycemic Protein from Lupin Seeds in a Solution and Adsorbed on Polystyrene Nanoparticles to Mimic a Complex Food Matrix.

The search for bioactivities influencing the human wellbeing of food proteins and peptides is a topic of broad and current interest. γ-Conglutin (γC) is a lupin seed protein drawing remarkable pharmacological and/or nutraceutical interest, as it is able to reduce hyperglycemia in humans and animal models. The present work deepens our investigations to understand the molecular basis of the in vitro effects of γC by testing the possible metabolic effects on cultivated Caco-2 cells. γC and its derived peptides (obtained via simulated gastrointestinal digestion) did not influence the cell viability at incubation times up to 24 h. The incubation of cells with native or digested γC caused no detectable inflammation processes mediated by Nuclear Factor kappa B (NFκB). We checked if treatment with γC or its derived peptides can elicit the expression of two peptide transporters (Pept-1 and Htp-1) by using an RT-qPCR approach. Native γC caused the halving of Pept-1 expression compared to untreated cells, but this effect disappeared when γC was digested. Either native γC or γC peptides reduced the expression levels of Hpt-1. Finally, this work also sheds light on the possible structural modifications of γC that may occur in the gastrointestinal tract, using an in vitro simulated dispersed system with polystyrene nanoparticles (NPs).

Soil Application of Effective Microorganisms (EM) Maintains Leaf Photosynthetic Efficiency, Increases Seed Yield and Quality Traits of Bean (Phaseolus vulgaris L.) Plants Grown on Different Substrates.

EM (effective microorganisms) is a biofertilizer consisting of a mixed culture of potentially beneficial microorganisms. In this study, we investigated the effects of EM treatment on leaf in vivo chlorophyll a fluorescence of photosystem II (PSII), yield, and macronutrient content of bean plants grown on different substrates (nutrient rich substrate vs. nutrient poor sandy soil) in controlled environmental conditions (pot experiment in greenhouse). EM-treated plants maintained optimum leaf photosynthetic efficiency two weeks longer than the control plants, and increased yield independent of substrate. The levels of seed nutritionally-relevant molecules (proteins, lipids, and starch) were only slightly modified, apart from the protein content, which increased in plants grown in sandy soil. Although EM can be considered a promising and environmentally friendly technology for sustainable agriculture, more studies are needed to elucidate the mechanism(s) of action of EM, as well as its efficacy under open field conditions.