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The Camellia genus comprises a vast array of underexplored medicinal plants that merit a systematic valorization to exploit their potential as natural sources of phytochemicals with associated health-promoting properties. In this work, flower extracts from eight poorly characterized Camellia japonica L. cultivars were subjected to polyphenol profiling through untargeted metabolomics combined with in vitro functional analysis. Anthocyanins, mostly represented by cyanidin 3-O-glycosides, flavones, and flavonols, were found as the major constituents of C. japonica flowers, together with hydroxycinnamic acids, tyrosols, alkylphenols, and stilbenes, which were detected for the first time in this species. The application of multivariate statistics revealed a flower color-dependent fingerprint of C. japonica cultivars, featuring anthocyanins and other flavonoids as metabolite markers associated with color-flowered cultivars with respect to white-flowered ones. The accumulation of anthocyanins, especially reported in ‘Eugenia de Montijo’ flowers, was highly correlated with the cytotoxic and anti-inflammatory properties of the derived extracts, including AGS, Caco-2, and MCF7 cancer cell lines. Moreover, the flavones accumulation reported in ‘Carolyn Tuttle’ extracts was also associated with high rates of free-radical scavenging activity, as well as a potent cytotoxicity against the Caco-2 cell line. In general, C. japonica anthocyanin-enriched flower extracts were revealed as promising candidates for the industrial production of polyphenols with associated biological activities of high interest for critical sectors in the food, pharmaceutical, and cosmetic industries.

The modulation of plant secondary metabolism by the manipulation of nutrient solution (NS) electrical conductivity and biostimulant application has the potential to increase crop growth and the content of bioactive compounds, thus ameliorating the qualitative attributes of vegetables. A Graminaceae-derived protein hydrolysate (pH) and its molecular fractions PH1 (>10 kDa), PH2 (1 < x <10 kDa) and PH3 (<1 kDa) were applied on lettuce grown in a soilless system and irrigated with two levels of NaCl in the NS (0 and 30 mM). The different pH fractions provided distinct responses, with PH2 in saline conditions increasing lettuce fresh weight by 10.48%, and decreased Na, Cl, malate and citrate concentrations in the leaves by 37.24%, 15.45%,31.4% and 33.73% compared to control. Similar results were achieved by the fraction PH3. However, pH and its fractions PH2 and PH3 increased total flavonoids in salinity conditions, while pH and its fraction PH3 increased total phenolic acids in both saline and non-saline conditions. At the same time, all the 3 fractions of pH increased lutein in both conditions. Therefore, pH fractions should be casted, under tailored conditions, to achieve the desired growth and modulation of qualitative attributes. Nonetheless, omic approaches should be further recommended to support and clarify the mechanisms of these fractions.

Root exudates play an essential role in helping plants to cope with abiotic stress. However, the modulation of root exudation profiles under multiple stress conditions is still poorly understood. Using targeted and untargeted metabolomics, here we tested the effect of drought, heat stress, and their combination on maize root exudates, also considering the differences that might exist between root types (seminal and primary) and root zones (apical and sub-apical). In addition, we built an analytical framework that relates the root exudation profile with the rhizosphere bacterial community, enabling us to dissect the interactions between specific root exudates and bacterial taxa. The composition of root exudates undergoes distinct modulation according to the single or combined stress and to the root zone, but not according to the root type. In addition, we found that stress-specific exudates can influence the relative abundance of specific bacterial taxa, some of which are known to be beneficial microorganisms. Our results contribute to the understanding of plant-soil interactions under the influence of abiotic stressors, which is key in paving the way towards an increased resilience to abiotic stresses, representing a powerful tool to craft the next generation of agricultural practices.

The use of plant biostimulants such as seaweed extracts (SWE) and protein hydrolysates (PH) has grown in the recent years due to their beneficial effects on yield under both optimal and sub-optimal conditions such as salt stress. The comprehension of the mode of action of these two important categories of biostimulants on plant performance will allow to use them more efficiently under different growth conditions. This study aimed to examine the efficacy of a seaweed and plant-based biostimulants on greenhouse lettuce (Lactuca sativa L.) grown under non-saline (0 mM NaCl) and saline conditions (40 mM NaCl) in terms of growth, yield, SPAD index, leaf mineral composition and metabolomic profiling. Shoot fresh weight of lettuce was reduced by 15.3% under 40 mM of NaCl. Nonetheless, it was boosted by both used biostimulants by 9 and 18%, on average, under 0 and 40 mM NaCl salinity, respectively. Na content under saline conditions was reduced in the presence of the biostimulants treatment, where PH reduced it significantly by 15.6% and SWE by 9.4%. On the other hand, Cl content was significantly reduced only under PH treatment. Both biostimulants elicited a broad metabolic reprogramming, involving the accumulation of stress-related compounds such as glucosinolates, terpenoid phytoalexins, and jasmonates. Interestingly, distinctive metabolomic signatures could be observed following the application of the different biostimulants under salinity conditions. In more detail, PH promoted the accumulation of glucosinolates and phytoalexins precursors, while SWE induced a down accumulation of secondary metabolites. Our findings indicate different processes being modulated by PH and SWE, with possible synergistic effects, thus paving the way towards integrated strategies to alleviate the detrimental effects of salinity in lettuce.

Microplastics (MP) are ubiquitous contaminants of great concern due to their high persistence and potential hazardous impact on the environment. Depending on their size and shape, as well as the chemical additives they can have in their polymeric structure, MP can be taken up by organisms, ultimately leading to direct and indirect toxic effects. In this review, we discuss the primary sources, fate, and impact of MP on the rhizosphere ecology, focusing in particular on how soil physical-chemical properties, plant physiology, and soil biodiversity are modulated by the interaction of MP in the plant-soil system. Current knowledge on soil ecotoxicology shows that MP directly affects soil quality and fertility via alteration of soil nutrient cycling and microbial communities and indirectly by changing soil bulk density, pH, porosity, electric conductivity, and nutrient bioavailability. MP is also known to affect soil animals by altering their feeding, mobility, and reproductive behavior. Toxic effects of MP on the multiple and co-occurring interactions among the soil, soil organisms and plants, particularly in the long-term, remain unstudied. Indeed, a better understanding of rhizosphere-MP interactions at a functional level (e.g., nutrient availability, pollutant immobilization, root exudates, etc.) is urgently needed to develop risk assessment frameworks of soil pollution by MP.

An optimized ultrasound-assisted extractive method was developed to obtain a polyphenol-enriched extract from the aerial parts of Thymus comosus Heuff. ex Griseb. et Schenk. Optimization process was conducted based on Design of Experiment (DoE) principles, determining the influence of three independent variables (time, ultrasound amplitude, ethanol concentration) on the total phenolic content of the extract (dependent variable). Additionally, the phenolic composition of the extract was characterized through UHPLC-HRMS, revealing beside the most abundant flavonoid-type compounds the presence of salvianolic acids C, D and L in high amounts. Phytochemical profile of the extract was correlated with its antioxidant activity (tested through five complementary assays) and enzyme-inhibitory potential, showing important antiglucosidase and anticholinesterase effects. Overall, it was concluded that the developed method is suitable for obtaining a good recovery of both phenolic and non-phenolic compounds from Thymus comosus aerial parts, and their presence in the optimized extract is responsible for its pharmacological potential.

Salinization of agricultural lands is of increasing concern at global scale due to its negative impacts on several physiological and developmental processes of most crops, that in turn can lead to threatening food security. Salt-tolerant plant growth-promoting bacteria (PGPB) represent an emerging strategy to protect plants and enhance the productivity of agroecosystems suffering from salinization. This study aimed at investigating the individual effect of two PGPB in alleviating salt-stressed tomato plants, as well as the rhizosphere functioning and metabolomic profiles under soil salinity and PGPB inoculation treatments. The Biolog Eco-Plate functional analysis and untargeted metabolomics approaches were used to this aim. Under saline conditions, plants inoculated with Pseudomonas 16S showed higher biomass (p < 0.05) than both uninoculated and Enterobacter 15S-inoculated plants, indicating that the former strain was efficient in alleviate the saline stress. Moreover, independently from PGPB inoculation, the microbial metabolism in tomato's rhizosphere was significantly increased by salinity, especially related to the catabolism of amines, carbohydrates, and polymers. The UHPLC/QTOF-MS approach allowed the identification of signature metabolomic patterns for the rhizosphere of tomato, with inflected accumulation of compounds imposed by either Pseudomonas 16S, Enterobacter 15S or saline stress. Crucial compounds from primary and secondary metabolism related to salinity stress or PGPB inoculation were detected, including low molecular weight osmolytes (e.g., amino acids), as well as anthocyanins, hydroxycinnamic acids, coumarins and saponins. An increase in the content of ROS-scavenging and antioxidant compounds in addition to the facilitation of Fe acquisition induced by Pseudomonas 16S, are suggested as mechanisms responsible for the higher biomass accumulation by tomato plants grown under saline stress.

Nanomaterials-mediated contamination (including the highly reactive metal oxides ZnO nanoparticles) is becoming one of the most concerning issues worldwide. In this study, the toxic effects of two chemical species of Zn (ZnO nanoparticles and bulk ZnSO4) were investigated in bean plants, following either foliar or soil application, at concentrations from 250 to 2000 mg L−1 using biochemical assays, proteomics and metabolomics. The accumulation of Zn in plant tissues depended on the application type, zinc chemical form and concentration, in turn triggering distinctive morphological, physiological, and redox responses. Bean plants were more sensitive to the foliar than to the soil application, and high concentrations of ZnO NP and bulk ZnSO4 determined the highest plant growth inhibition and stress symptoms. However, low dosages of ZnSO4 induced a slight plant growth promotion and better physiological and antioxidative response. Low concentration of Zn leaded to increased activity of stress-related proteins and secondary metabolites with antioxidant capacity, while increasing concentration reached the exhausted phase of the plant stress response, reducing the antioxidant defense system. Such high concentrations increased lipids peroxidation, protein degradation and membranes integrity. Oxidative damage occurred at high concentrations of both chemical species of Zn. Foliar spraying impaired photosynthetic efficiency, while soil applications (especially ZnSO4) elicited antioxidant metabolites and proteins, and impaired chloroplast-related proteins involved in the electron transport chain and ATP production. Taken together, the results highlighted distinctive and nanoparticles-related toxic effects of ZnO in bean, compared to ionic forms of Zn.

In this work, the combination of elicited plant suspension cultured cells (PSCCs) with untargeted metabolomics establishes a powerful, cutting-edge strategy to unravel the effects of elicitors on the biosynthetic potential of medicinal plants, as Bryophyllum sp. The PSCC technology constitutes a successful biotechnological system for the study and production of bioactive compounds throughout the inclusion of elicitors with the ability to modulate secondary metabolism. The use of methyl jasmonate (MJ) and salicylic acid (SA) as abiotic elicitors on bryophyllum PSCCs, resulted in differential effects on cell growth and secondary metabolism, depending on the species, including synergistic and antagonistic effects. This fact suggests that both elicitors play a pleiotropic effect on plant secondary metabolism, showing complex interactions, according to the UHPLC-QTOF mass spectrometry profiling. Thus, the combination of both elicitors induced a strong synergistic on B. daigremontianum PSCCs, with 2272 putatively annotated compounds, whereas it caused a negative effect on the secondary metabolism of B. × houghtonii PSCCs, being MJ the only elicitor driving a positive effect, presenting 2972 annotated compounds. Meanwhile, B. tubiflorum PSCCs did not show a significant modulation of secondary metabolism, with 1521 annotated compounds. The metabolite annotation indicated that three families of secondary metabolites were mainly affected by elicitation: phenolic compounds constituted the most affected family by elicitation, mainly represented by flavonoids and lignans; N-containing compounds included glucosinolates, amines, and alkaloids, reported to Bryophyllum sp. for the first time; and terpenoids included mainly phytoalexins and saponins. The results depict a deep genotype-dependent metabolomic reprogramming of secondary metabolism in response to elicitors, thanks to the application of untargeted metabolomics. This knowledge will allow the consideration of Bryophyllum sp.

Inoculation with plant growth-promoting bacteria (PGPB) represents an efficient method in sustainable agriculture to improve nutrients availability and crop production in diverse environmental conditions. In the present work, untargeted metabolomics and community-level physiological profiles (CLPP) approaches were employed to investigate the shaping of tomato rhizosphere functioning and potential metabolic activity imposed by rhizosphere-associated microbiome, following inoculation with two different PGPB (Enterobacter sp. 15S and Pseudomonas sp. 16S). Significant increases in root and shoot dry biomass were observed in both inoculated treatments, when compared to uninoculated plants (p < 0.05). The untargeted metabolomics allowed discriminating the metabolic profiles of tomato rhizosphere, with distinct modulations imposed by either Enterobacter 15S or Pseudomonas 16S. Flavonoids and other phenolics were among the most frequently identified differential metabolites in the rhizosphere from both Enterobacter 15S and Pseudomonas 16S inoculated plants. Nevertheless, other metabolites like phytohormones and amino acids were also decisive to this specific modulation. The metabolic activity profile rhizosphere-associated microbiome of tomato plants unveiled by the CLPP analysis was congruent with the metabolomic data and reinforced the influence exerted by the bacterial inoculation on modulating the rhizosphere microbiome functioning. In particular, the microbiome associated with control and 16S-treated plants showed a higher functional diversity than those treated with Enterobacter 15S. Carbohydrates, carboxylic acids, amino acids, and polymers were the main classes of substrates which contributed to such differences.

Muscari turcicum, endemic to south Anatolia, Turkey, represents an unexplored crop plant, with potential therapeutic uses related to its phytochemical composition. In this work, the in vitro antioxidant and enzyme inhibitory activity of flower, leaf and bulb extracts, obtained using different extraction methods were evaluated. A comprehensive polyphenolic and sesquiterpene lactones profiling of the different extracts was also undertaken. For this purpose, UHPLC-QTOF mass spectrometry allowed us to putatively annotate 280 phytochemical compounds of which 162 were polyphenols and 118 were sesquiterpene lactones. The most abundant polyphenols were flavonoids (77 compounds), phenolic acids (34 compounds), and low molecular weight phenols (38 compounds). Muscari turcicum leaf methanol extract possessed the highest concentrations of low-molecular-weight phenolics, phenolic acids, and sesquiterpene lactones (20.61, 7.00, and 3.44 mg standard equivalent/g, respectively). The water extract of M. turcicum flower obtained by infusion showed prominent reducing (120.52 mg Trolox equivalent [TE]/g mg TE/g for both CUPRAC and FRAP) and radical scavenging potential (91.39 mg TE/g, for DPPH assay). Besides, M. turcicum flower methanol extract (13.44 mg EDTA equivalent/g) showed the highest metal chelating activity. Interestingly, methanol extracts obtained by Soxhlet extraction and maceration actively inhibited tyrosinase (129.36 mg kojic acid equivalent/g) and cholinesterases (5.15 mg galantamine equivalent [GALAE]/g and 6.16 mg GALAE/g, for acetyl and butyryl cholinesterase) respectively. Strong correlations (p < 0.01) were observed between polyphenols/sesquiterpenoids and observed biological activities. Scientific evidences presented in this study has provided baseline data for bioprospection of novel pharmaceutical/cosmetic candidates from Muscari turcicum, thus supporting its therapeutic exploitation.

Synthetic pesticides may have non-target effects on soil microorganisms which have been identified as a specific protection goal in pesticide environmental risk assessment. Most studies to date have focused on the effects of synthetic pesticides on soil microorganisms, whereas little is known about the response of the soil microbial community to the so-called low-risk pesticide classes. The main objective of this study was to assess the impact of a botanical (azadirachtin) and a low-dose pesticide (trifloxystrobin), applied as commercial formulations, on soil microbial functions. In a microcosm study, pesticides were applied in soil at increasing dose rates (up to ×100 the recommended dose rate). Their dissipation and the formation of the major transformation product (TP) of trifloxystrobin were determined. Enzymatic activities and the abundance of key functional microbial groups were measured via fluorometric assays, potential nitrification (PNT) and q-PCR. Trifloxystrobin and azadirachtin did not persist in soil at all dose rates with DT 50s of 1.1–1.4 and 1.3 days respectively. No pesticide dose-dependent inhibitory effects on the activity of soil enzymes, the abundance of ammonia-oxidizing bacteria (AOB), ammonia-oxidizing archaea (AOA), sulfur-oxidizing bacteria (SOB) and on PNT were seen. Instead the application of both pesticides at all dose rates stimulated PNT. Trifloxystrobin was transformed to trifloxystrobin acid (TFSA), which persisted in soil. However its formation did not correlate with adverse effects on soil microbial functions. We provide first evidence that trifloxystrobin and azadirachtin do not impose unacceptable effects on soil microbial functions even at high dose rates.