Chemical Profile and Biological Activity of Cherimoya (Annona cherimola Mill.) and Atemoya (Annona atemoya) Leaves.

Annona cherimola (Cherimoya) and Annona atemoya (Atemoya) are tropical plants known for their edible fruit. Scientific data suggest that their leaves, used in traditional medicine in the form of teas or infusions without evidence of toxicity, contain several bioactive compounds. However, only Annona muricata among all the Annona species is currently used in the nutraceutical field, and its dried leaves are marketed for tea preparation. In this work, we explored the nutraceutical potential of Atemoya and Cherimoya leaves, by evaluating their chemical profile and functional properties. Phytochemical analyses showed large amounts of phenolic compounds, in particular proanthocyanidins, and identified 18 compounds, either flavonoids or alkaloids. Concerning biological activity, we found antioxidative properties correlated with polyphenols, and antiproliferative activity against HeLa and HepG2 cell lines correlated with alkaloids. The obtained results demonstrate the potential use of Annona cherimola leaves for the preparation of dietary supplements aimed to promote the physiological redox balance. Moreover, the varietal comparison suggests that two commercial cultivars (Campas and White) and the local Torre 1, better suit this purpose. On the other hand, among the studied cultivars, Campas and Torre 1 are also the richest in alkaloids and, in consideration of the anti-proliferative properties of their extracts, dietary supplements based on these cultivars might also have chemo-preventive effects.

Phytochemical profile and antioxidant properties of the edible and non-edible portions of black sapote (Diospyros digyna Jacq.).

This study evaluated the phytochemical profile and antioxidative properties of the edible and non-edible portions of black sapote. The phytochemical analysis highlighted the presence of several bioactive compounds, differently distributed among peel, pulp and seeds. In particular, the peel resulted rich of flavan-3-ols and proanthocyanidins, whereas seeds contained high amount of organic acids, including ferulic, citric and sinapic acids. Concerning functional properties, both edible and non-edible portions showed a significant prevention of lipid peroxidation in a cell-based model. Moreover, the results suggested that the antioxidant protection involved both redox active properties and gene expression modulation. Concerning redox active properties, peel extracts showed an antioxidant activity 7/12-fold higher than the edible portion, while seed extracts were more active in increasing catalase gene expression. The obtained results confirmed that black sapote is a good source of antioxidant phytochemicals and its non-edible portions have a great potential in the production of functional foods and supplements.

Combining metabolite doping and metabolic engineering to improve 2-phenylethanol production by engineered cyanobacteria.

2-Phenylethanol (2-PE) is a rose-scented aromatic compound, with broad application in cosmetic, pharmaceutical, food and beverage industries. Many plants naturally synthesize 2-PE via Shikimate Pathway, but its extraction is expensive and low-yielding. Consequently, most 2-PE derives from chemical synthesis, which employs petroleum as feedstock and generates unwanted by products and health issues. The need for "green" processes and the increasing public demand for natural products are pushing biotechnological production systems as promising alternatives. So far, several microorganisms have been investigated and engineered for 2-PE biosynthesis, but a few studies have focused on autotrophic microorganisms. Among them, the prokaryotic cyanobacteria can represent ideal microbial factories thanks to their ability to photosynthetically convert CO2 into valuable compounds, their minimal nutritional requirements, high photosynthetic rate and the availability of genetic and bioinformatics tools. An engineered strain of Synechococcus elongatus PCC 7942 for 2-PE production, i.e., p120, was previously published elsewhere. The strain p120 expresses four heterologous genes for the complete 2-PE synthesis pathway. Here, we developed a combined approach of metabolite doping and metabolic engineering to improve the 2-PE production kinetics of the Synechococcus elongatus PCC 7942 p120 strain. Firstly, the growth and 2-PE productivity performances of the p120 recombinant strain were analyzed to highlight potential metabolic constraints. By implementing a BG11 medium doped with L-phenylalanine, we covered the metabolic burden to which the p120 strain is strongly subjected, when the 2-PE pathway expression is induced. Additionally, we further boosted the carbon flow into the Shikimate Pathway by overexpressing the native Shikimate Kinase in the Synechococcus elongatus PCC 7942 p120 strain (i.e., 2PE_aroK). The combination of these different approaches led to a 2-PE yield of 300 mg/gDW and a maximum 2-PE titer of 285 mg/L, 2.4-fold higher than that reported in literature for the p120 recombinant strain and, to our knowledge, the highest recorded for photosynthetic microorganisms, in photoautotrophic growth condition. Finally, this work provides the basis for further optimization of the process aimed at increasing 2-PE productivity and concentration, and could offer new insights about the use of cyanobacteria as appealing microbial cell factories for the synthesis of aromatic compounds.

Modulation of Antioxidant Defense in Farmed Rainbow Trout (Oncorhynchus mykiss) Fed with a Diet Supplemented by the Waste Derived from the Supercritical Fluid Extraction of Basil (Ocimum basilicum).

Phytotherapy is based on the use of plants to prevent or treat human and animal diseases. Recently, the use of essential oils and polyphenol-enriched extracts is also rapidly increasing in the aquaculture sector as a means of greater industrial and environmental sustainability. Previous studies assessed the antibacterial and antiparasitic effects of these bioactive compounds on fish. However, studies on the modulation of oxidative stress biomarkers are still scant to date. Thus, in this study, the modulation of antioxidant defense against oxidative stress exerted by fish diets supplemented with a basil supercritical extract (F1-BEO) was assessed in rainbow trout Oncorhynchus mykiss. The F1-BEO extracted with supercritical fluid extraction was added to the commercial feed flour (0.5, 1, 2, 3% w/w) and mixed with fish oil to obtain a suitable compound for pellet preparation. Fish were fed for 30 days. The levels of stress biomarkers such as superoxide dismutase, catalase, glutathione peroxidase, glutathione S-transferase, glutathione reductase, glyoxalase I, glyoxalase II, lactate dehydrogenase, glutathione and malondialdehyde showed a boost in the antioxidant pathway in fish fed with a 0.5% F1-BEO-supplemented diet. Higher F1-BEO supplementation led to a failure of activity of several enzymes and the depletion of glutathione levels. Malondialdehyde concentration suggests a sufficient oxidative stress defense against lipid peroxidation in all experimental groups, except for a 3% F1-BEO-supplemented diet (liver 168.87 ± 38.79 nmol/mg prot; kidney 146.86 ± 23.28 nmol/mg prot), compared to control (liver 127.76 ± 18.15 nmol/mg prot; kidney 98.68 ± 15.65 nmol/mg prot). Our results suggest supplementing F1-BEO in fish diets up to 0.5% to avoid potential oxidative pressure in farmed trout.

Combination of Poly(ε-Caprolactone) Biomaterials and Essential Oils to Achieve Anti-Bacterial and Osteo-Proliferative Properties for 3D-Scaffolds in Regenerative Medicine.

Biomedical implants, an essential part of the medical treatments, still suffer from bacterial infections that hamper patients' recovery and lives. Antibiotics are widely used to cure those infections but brought antibiotic resistance. Essential oils (EOs) demonstrate excellent antimicrobial activity and low resistance development risk. However, EO application in medicine is still quite scarce and almost no research work considers its use in combination with bioresorbable biomaterials, such as the poly(ε-caprolactone) (PCL) polymer. This work aimed to combine the antibacterial properties of EOs and their components, particularly eugenol and cinnamon oil, against Staphylococcus aureus, S. epidermidis and Escherichia coli, with those of PCL for medical applications in which good tissue regeneration and antimicrobial effects are required. The PCL porous scaffolds, added with increasing (from 30% to 50%) concentrations of eugenol and cinnamon oil, were characterized by square-shaped macropores. Saos-2 cells' cell viability/proliferation was hampered by 40 and 50% EO-enriched PCL, whereas no cytotoxic effect was recorded for both 30% EO-added PCL and pure-PCL. The antibacterial tests revealed the presence of a small inhibition halo around the 30% eugenol and cinnamon oil-functionalized PCL scaffolds only for staphylococci, whereas a significant decrease on both adherent and planktonic bacteria was recorded for all the three microorganisms, thus proving that, even if the EOs are only in part released by the EO-added PCL scaffolds, an anti-adhesive feature is anyway achieved. The scaffold will have the ability to support new tissue formation and simultaneously will be able to prevent post-surgical infection. This research shows the great potential in the use of EOs or their single components, at low concentrations, for biomaterial functionalization with enhanced anti-bacterial and biointegration properties.

Proanthocyanidins and Where to Find Them: A Meta-Analytic Approach to Investigate Their Chemistry, Biosynthesis, Distribution, and Effect on Human Health.

Proanthocyanidins (PACs) are a class of polyphenolic compounds that are attracting considerable interest in the nutraceutical field due to their potential health benefits. However, knowledge about the chemistry, biosynthesis, and distribution of PACs is limited. This review summarizes the main chemical characteristics and biosynthetic pathways and the main analytical methods aimed at their identification and quantification in raw plant matrices. Furthermore, meta-analytic approaches were used to identify the main plant sources in which PACs were contained and to investigate their potential effect on human health. In particular, a cluster analysis identified PACs in 35 different plant families and 60 different plant parts normally consumed in the human diet. On the other hand, a literature search, coupled with forest plot analyses, highlighted how PACs can be actively involved in both local and systemic effects. Finally, the potential mechanisms of action through which PACs may impact human health were investigated, focusing on their systemic hypoglycemic and lipid-lowering effects and their local anti-inflammatory actions on the intestinal epithelium. Overall, this review may be considered a complete report in which chemical, biosynthetic, ecological, and pharmacological aspects of PACs are discussed.

A new protein hydrolysate-based biostimulant applied by fertigation promotes relief from drought stress in Capsicum annuum L.

Recently, biostimulants have been used in sustainable agriculture as priming agents able to increase crop tolerance to abiotic stressors. Here, a soil application of GHI_16_VHL, a plant protein hydrolysate-based biostimulant, was tested for its capability to mitigate severe water stress effects on Capsicum annuum at flowering time. The biostimulant influence on plant physiological status was monitored upon stress and its relief, by measuring chlorophyll levels, stomatal density, stem water potential, leaf gas exchanges and plant growth. Moreover, leaf osmoregulation and oxidative stress levels were also evaluated by quantifying free proline, total non-structural carbohydrates (NSC), ROS-scavenging activity and H2O2 level. Although biostimulant-primed plants showed a quicker decrease of stem water potential with respect to untreated plants upon drought imposition, they recovered faster probably due to the higher leaf osmolyte accumulation, namely NSC during drought. Moreover, leaf gas exchange recovery was prompted in biostimulant-treated plants, which showed an incremented stomatal density and the same chlorophyll level of well-watered plants at the end of the recovery phase. Hydrogen peroxide level was significantly lower during stress and early recovery in biostimulant primed plants, probably due to the higher catalase activity in treated plants before drought or to the higher level of non-enzymatic antioxidant scavengers in primed stressed plants. Finally, the biostimulant priming increased aboveground relative growth rate and final fruit yield of stressed plants. Taken together, our data suggest that the biostimulant priming treatment promotes a faster and more efficient plant recovery after drought.

Clostridium cellulovorans Proteomic Responses to Butanol Stress.

Combination of butanol-hyperproducing and hypertolerant phenotypes is essential for developing microbial strains suitable for industrial production of bio-butanol, one of the most promising liquid biofuels. Clostridium cellulovorans is among the microbial strains with the highest potential for direct production of n-butanol from lignocellulosic wastes, a process that would significantly reduce the cost of bio-butanol. However, butanol exhibits higher toxicity compared to ethanol and C. cellulovorans tolerance to this solvent is low. In the present investigation, comparative gel-free proteomics was used to study the response of C. cellulovorans to butanol challenge and understand the tolerance mechanisms activated in this condition. Sequential Window Acquisition of all Theoretical fragment ion spectra Mass Spectrometry (SWATH-MS) analysis allowed identification and quantification of differentially expressed soluble proteins. The study data are available via ProteomeXchange with the identifier PXD024183. The most important response concerned modulation of protein biosynthesis, folding and degradation. Coherent with previous studies on other bacteria, several heat shock proteins (HSPs), involved in protein quality control, were up-regulated such as the chaperones GroES (Cpn10), Hsp90, and DnaJ. Globally, our data indicate that protein biosynthesis is reduced, likely not to overload HSPs. Several additional metabolic adaptations were triggered by butanol exposure such as the up-regulation of V- and F-type ATPases (involved in ATP synthesis/generation of proton motive force), enzymes involved in amino acid (e.g., arginine, lysine, methionine, and branched chain amino acids) biosynthesis and proteins involved in cell envelope re-arrangement (e.g., the products of Clocel_4136, Clocel_4137, Clocel_4144, Clocel_4162 and Clocel_4352, involved in the biosynthesis of saturated fatty acids) and a redistribution of carbon flux through fermentative pathways (acetate and formate yields were increased and decreased, respectively). Based on these experimental findings, several potential gene targets for metabolic engineering strategies aimed at improving butanol tolerance in C. cellulovorans are suggested. This includes overexpression of HSPs (e.g., GroES, Hsp90, DnaJ, ClpC), RNA chaperone Hfq, V- and F-type ATPases and a number of genes whose function in C. cellulovorans is currently unknown.

Towards a new application of amaranth seed oil as an agent against Candida albicans.

Amaranthus spp. (Amaranthaceae family), known as amaranth, are plants native of Central America, today produced in many parts of the world. due to their popularity popular as a health food. Because of its composition, amaranth can be considered to be attractive not only as a food but also for pharmaceutical and cosmetics uses. To date, antifungal activity of amaranth extracts has not been totally investigated, therefore the scope of this study was to evaluate the antifungal effect of the apolar fraction from Amaranthus cruentus L. seeds extract, alone and in association with antifungal drugs terbinafine, a common antifungal agent, which itself has only fungistatic effect on Candida albicans strains without exerting fungicidal activity. Our results demonstrate that this amaranth oil in combination with terbinafine has synergic fungistatic and fungicidal activity, with FICI of 0.466 and 0.496, respectively. No fungistatic and fungicidal activity of terbinafine alone at concentrations up to 64 µg/mL and amaranth oil alone at concentrations up to 2000 µg/mL, against all tested C. albicans strains, were observed. does not show activity towards Candida albicans strains but it can effectively potentiate the antifungal activity of terbinafine, a common antifungal agent which itself This result suggests the possible application of amaranth oil in the preparation of formulations with terbinafine for topical use.

Transcriptome Analyses and Antioxidant Activity Profiling Reveal the Role of a Lignin-Derived Biostimulant Seed Treatment in Enhancing Heat Stress Tolerance in Soybean.

Soybean (Glycine max Merr.) is a worldwide important legume crop, whose growth and yield are negatively affected by heat stress at germination time. Here, we tested the role of a biostimulant based on lignin derivatives, plant-derived amino acids, and molybdenum in enhancing soybean heat stress tolerance when applied on seeds. After treatment with the biostimulant at 35 °C, the seed biometric parameters were positively influenced after 24 h, meanwhile, germination percentage was increased after 72 h (+10%). RNA-Seq analyses revealed a modulation of 879 genes (51 upregulated and 828 downregulated) in biostimulant-treated seeds as compared with the control, at 24 h after incubation at 35 °C. Surprisingly, more than 33% of upregulated genes encoded for ribosomal RNA (rRNA) methyltransferases and proteins involved in the ribosome assembly, acting in a specific protein network. Conversely, the downregulated genes were involved in stress response, hormone signaling, and primary metabolism. Finally, from a biochemical point of view, the dramatic H2O2 reduction 40%) correlated to a strong increase in non-protein thiols (+150%), suggested a lower oxidative stress level in biostimulant-treated seeds, at 24 h after incubation at 35 °C. Our results provide insights on the biostimulant mechanism of action and on its application for seed treatments to improve heat stress tolerance during germination.

Squalene: More than a Step toward Sterols.

Squalene (SQ) is a natural triterpene widely distributed in nature. It is a metabolic intermediate of the sterol biosynthetic pathway and represents a possible target in different metabolic and oxidative stress-related disorders. Growing interest has been focused on SQ's antioxidant properties, derived from its chemical structure. Strong evidence provided by ex vivo models underline its scavenging activity towards free radicals, whereas only a few studies have highlighted its effect in cellular models of oxidative stress. Given the role of unbalanced free radicals in both the onset and progression of several cardiovascular diseases, an in depth evaluation of SQ's contribution to antioxidant defense mechanisms could represent a strategic approach in dealing with these pathological conditions. At present experimental results overall show a double-edged sword role of squalene in cardiovascular diseases and its function has to be better elucidated in order to establish intervention lines focused on its features. This review aims to summarize current knowledge about endogenous and exogenous sources of SQ and to point out the controversial role of SQ in cardiovascular physiology.

Chemical, Physiological and Molecular Responses of Host Plants to Lepidopteran Egg-Laying.

Plant-lepidopteran interactions involve complex processes encompassing molecules and regulators to counteract defense responses they develop against each other. Lepidoptera identify plants for oviposition and exploit them as larval food sources to complete their development. In turn, plants adopt different strategies to overcome and limit herbivorous damages. The insect egg deposition on leaves can already induce a number of defense responses in several plant species. This minireview deals with the main features involved in the interaction between plants and lepidopteran egg-laying, focusing on responses from both insect and plant side. We discuss different aspects of direct and indirect plant responses triggered by lepidopteran oviposition. In particular, we focus our attention on the mechanisms underlying egg-induced plant defenses that can i) directly damage the eggs such as localized hypersensitive response (HR)-like necrosis, neoplasm formation, production of ovicidal compounds and ii) indirect defenses, such as production of oviposition-induced plant volatiles (OIPVs) used to attract natural enemies (parasitoids) able to kill the eggs or hatching larvae. We provide an overview of chemical, physiological, and molecular egg-mediated plant responses induced by both specialist and generalist lepidopteran species, also dealing with effectors, elicitors, and chemical signals involved in the process. Egg-associated microorganisms are also discussed, although little is known about this third partner participating in plant-lepidopteran interactions.

Anti-Proliferative Effects of an Extra-Virgin Olive Oil Extract Enriched in Ligstroside Aglycone and Oleocanthal on Human Liver Cancer Cell Lines.

Oleocanthal and ligstroside aglycone are olive oil-derived polyphenols. The former interferes with tumor growth with minor or no cytotoxicity on non-tumorigenic primary cell lines. The information about the bioactivity of ligstroside aglycone are scanty, with the exception of a known antioxidant power. Hepatocellular carcinoma is a malignant tumor with high mortality rates. Systemic chemotherapy is only marginally effective and is frequently complicated by toxicity. Previous observations have shown that hepatocellular carcinoma cell lines become more sensitive to taxol when it is combined with Tumor Necrosis Factor α (TNFα). The present work aimed to assess the effects of a polyphenolic extract containing both oleocanthal and ligstroside aglycone on proliferation and/or death in three liver cancer cell lines (HepG2, Huh7 and Hep3B). The possibility to enhance such effect by the addition of TNFα was also investigated. Both cell proliferation and death were enhanced by the exposure to the polyphenolic extract. Such effect was associated with induction of autophagy and could be potentiated by TNFα. The presence of ligstroside aglycone in the extract lowered the oleocanthal concentration required for cytotoxicity. These results show for the first time that the effects of a polyphenol extract can be potentiated by TNFα and that modulation of autophagy likely account for these effects.

The effect of isabelin, a sesquiterpene lactone from Ambrosia artemisiifolia on soil microorganisms and human pathogens.

Ambrosia artemisiifolia L. (common ragweed) is an invasive weed, which is well known for the strong allergenic effect of its pollen as well as for its invasiveness and impact in crop fields (e.g. causing yield losses). This species produces a broad range of sesquiterpenoids. In recent years, new bioactive molecules have been discovered in this plant, e.g. isabelin, a sesquiterpene dilactone. The bioactivity of isabelin has been already demonstrated on allergy-related receptors and its inhibitory effect on seeds of various plant species. Isabelin was tested for potential antimicrobial effects by using a selection of soil-borne bacteria and fungi and three human pathogens as model organisms. For the majority of microorganisms tested, no antimicrobial activity of isabelin was observed. However, isabelin revealed strong antimicrobial activity against the Gram-positive soil bacterium Paenibacillus sp. and against the Gram-positive, multidrug-resistant Staphylococcus aureus. The observed inhibitory activity of isabelin can enlighten the importance to study similar compounds for their effect on human pathogens and on soil and rhizosphere microorganisms.

Restriction fragment length polymorphism of the 5S-rRNA-NTS region: a rapid and precise method for plant identification.

Molecular genetic methods have several advantages over classical morphological and chemical analyses. The genetic method requires genotype instead than phenotype, therefore PCR-based techniques have been widely used for a rapid identification of plant species, varieties and chemotypes. Recently, the molecular discrimination of some higher plant species has been evaluated using sequences of a 5S-rRNA gene spacer region. The variation in the nontranscribed sequence (NTS) region has been used in a number of plant species for studying intraspecific variation, genome evolution, and phylogenetic reconstruction. Here, we describe a rapid method based on the use of the 5S-rRNA-NTS region as a tool for plant DNA fingerprinting, which combines PCR, sequencing and restriction fragment length polymorphism analyses.