A New Wound-Healing Tool Based on Glycyrrhiza glabra Extract-Loaded Ufasomes on Spanish Broom Dressings.

The development of innovative products for restoring skin integrity and promoting wound healing is still a challenge. The aim of this work was to evaluate an innovative Spanish broom wound dressing impregnated with Glycyrrhiza glabra extract-loaded ufasomes to improve wound healing. Ufasomes were characterized in terms of size, polydispersity index, entrapment efficiency, zeta potential, and stability. In addition, in vitro release studies and biocompatibility, biosafety, and scratch tests on WS1 fibroblasts were performed. The loaded ufasomes showed a nanometric size (<250 nm), good size distribution (lower than 0.3), and appropriate encapsulation efficiency (~67%). Moreover, the lipid vesicles showed good stability during the storage period and allowed for a slow release of glycyrrhizin, the main bioactive compound of the extract. Biological studies revealed that loaded vesicles are not cytotoxic, are hemocompatible, and lead to the complete closure of the scratch after about 33 h. To conclude, the results suggest that the developed dressings can be efficiently used to promote the healing process.

Metabolomic analysis and bioactivities of Arbutus unedo leaves harvested across the seasons in different natural habitats of Sardinia (Italy).

BACKGROUND: Arbutus unedo L. is a wild tree of Mediterranean regions used as food and in traditional medicine and important for afforestation programs. There is no detailed information available on the variation of A. unedo leaves metabolome across the seasons. The leaves were analyzed by Proton nuclear magnetic resonance (1 H NMR)-based metabolomics, comparing samples harvested across the seasons and in ten different natural habitats of Sardinia (Italy). RESULTS: Multivariate analysis showed the impact of seasonal variation on the metabolome: glucose and quinic acid increased in summer, while in spring sucrose was accumulated. ß-Arbutin, the main known active principle of A. unedo, generally reached the highest concentration in autumn. In winter, O-ß-methylglucose, γ-aminobutyric acid (GABA), flavonols (quercetin-3-O-α-rhamnoside, myricetin-3-O-α-rhamnoside, kaempferol-3-O-α-rhamnoside), catechin, and gallocatechin increased. Characteristic metabolomic features were found also for samples collected in different locations. For instance, trees growing at the highest altitude and exposed to lower temperatures produced less flavonols and catechins. The only sample collected on trees growing on limestones, dolomites, and dolomitic limestones type of soil showed generally the highest content of arbutin. The highest phenolics content was found during spring, while samples collected on flowering branches in winter were the ones with the highest flavonoid content. The antioxidant activity was also variated, ranging from 1.3 to 10.1 mg of Trolox equivalents (TE)/mL of extract, and it was positively correlated to both total phenolics and flavonoid content. Winter samples showed the lowest antibacterial activity, while summer and autumn ones exhibited the highest activity (IC50 values ranging from 17.3 to 42.3 µg/mL against Staphylococcal species). CONCLUSION: This work provides 1 H-NMR fingerprinting of A. unedo leaves, elucidating the main metabolites and their variations during seasons. On the basis of arbutin content, autumn could be considered the balsamic period of this taxon. Samples collected in this season were also the most active ones as antibacterial. Moreover, an interesting metabolomic profile enriched in catechins and flavonols was observed in leaves collected in winter on flowering branches which were endowed with high antioxidant potential.

Biotic and Abiotic Stressors in Plant Metabolism.

Plants are subject to a variety of biotic and abiotic stress that affect their metabolism, physiology, morphology, and growth [...].

Extraction, Encapsulation into Lipid Vesicular Systems, and Biological Activity of Rosa canina L. Bioactive Compounds for Dermocosmetic Use.

Valorization of wild plants to obtain botanical ingredients could be a strategy for sustainable production of cosmetics. This study aimed to select the rosehip extract containing the greatest amounts of bioactive compounds and to encapsulate it in vesicular systems capable of protecting their own antioxidant activity. Chemical analysis of Rosa canina L. extracts was performed by LC-DAD-MS/MS and 1H-NMR and vitamins, phenolic compounds, sugars, and organic acids were detected as the main compounds of the extracts. Liposomes, prepared by the film hydration method, together with hyalurosomes and ethosomes, obtained by the ethanol injection method, were characterized in terms of vesicle size, polydispersity index, entrapment efficiency, zeta potential, in vitro release and biocompatibility on WS1 fibroblasts. Among all types of vesicular systems, ethosomes proved to be the most promising nanocarriers showing nanometric size (196 ± 1 nm), narrow polydispersity (0.20 ± 0.02), good entrapment efficiency (92.30 ± 0.02%), and negative zeta potential (-37.36 ± 0.55 mV). Moreover, ethosomes showed good stability over time, a slow release of polyphenols compared with free extract, and they were not cytotoxic. In conclusion, ethosomes could be innovative carriers for the encapsulation of rosehip extract.

Plant Secondary Metabolites: An Opportunity for Circular Economy.

Moving toward a more sustainable development, a pivotal role is played by circular economy and a smarter waste management. Industrial wastes from plants offer a wide spectrum of possibilities for their valorization, still being enriched in high added-value molecules, such as secondary metabolites (SMs). The current review provides an overview of the most common SM classes (chemical structures, classification, biological activities) present in different plant waste/by-products and their potential use in various fields. A bibliographic survey was carried out, taking into account 99 research articles (from 2006 to 2020), summarizing all the information about waste type, its plant source, industrial sector of provenience, contained SMs, reported bioactivities, and proposals for its valorization. This survey highlighted that a great deal of the current publications are focused on the exploitation of plant wastes in human healthcare and food (including cosmetic, pharmaceutical, nutraceutical and food additives). However, as summarized in this review, plant SMs also possess an enormous potential for further uses. Accordingly, an increasing number of investigations on neglected plant matrices and their use in areas such as veterinary science or agriculture are expected, considering also the need to implement "greener" practices in the latter sector.

Screening of ninety herbal products of commercial interest as potential ingredients for phytocosmetics.

Elastase and tyrosinase are important targets both for cosmetics and for dermatological disorders. In this work, ninety herbal products were tested as inhibitors of these two enzymes. Eleven extracts resulted strongly active. Four out of them (Camellia sinensis, Ginkgo biloba, Rhodiola rosea, Vitis vinifera) inhibited both enzymes, five (Glycyrrhiza glabra, Ribes nigrum, Rheum officinale, Salvia officinalis, Tilia platyphyllos) were active against tyrosinase only, and two (Ceterach officinarum and Cinnamomum zeylanicum) proved selectively active against elastase. The IC50 ranged from 3.1 to 104.9 µg/mL and 19.3 to 164.3 µg/mL, against elastase and tyrosinase, respectively. The most active extracts resulted enriched in flavonoids (from 1.47 to 56.47 mg RE/g of extract) and phenolics (from 37.43 to 123.56 mg GAE/g of extract), indicating also an antioxidant potential. Finally, a positive correlation between enzymatic bioactivities and phenolic content was also established.

Grapevine Green Pruning Residues as a Promising and Sustainable Source of Bioactive Phenolic Compounds.

Green pruning residues (GPRs) and leaves from 16 red and white Vitis vinifera L. cultivars from Piedmont (Italy) were studied. The investigated samples were extracted by ultrasound-assisted extraction optimized by an experimental design, and quali- and quantitatively analyzed by HPLC-PDA-MS/MS. GPRs and leaves show a similar polyphenolic pattern, with quercetin 3-O-glucuronide, caftaric acid, and quercetin 3-O-glucoside as the main components, although in variable proportions. The HPLC results were related to the antioxidant activity, measured as total phenolic content and through DPPH and ABTS assays with similar results. Colorimetric in vitro assays, offline combined with HPLC-PDA analysis, determine which compounds contribute to the antioxidant activity in terms of radical scavenging abilities. Valorization of GPRs is a potential source of natural compounds that could be of interest in the health field, increasing their economic value together with a positive effect on the environment.

Identification of a Collagenase-Inhibiting Flavonoid from Alchemilla vulgaris Using NMR-Based Metabolomics.

This paper describes the use of 1H NMR profiling and chemometrics in order to facilitate the selection of medicinal plants as potential sources of collagenase inhibitors. A total of 49 plants with reported ethnobotanical uses, such as the healing of wounds and burns, treatment of skin-related diseases, rheumatism, arthritis, and bone diseases, were initially chosen as potential candidates. The in vitro collagenase inhibitory activity of hydroalcoholic extracts of these plants was tested. Moreover, their phytochemical profiles were analyzed by 1H NMR and combined with the inhibitory activity data by an orthogonal partial least squares model. The results showed a correlation between the bioactivity and the concentration of phenolics, including flavonoids, phenylpropanoids, and tannins, in the extracts. Considering the eventual false-positive effect on the bioactivity given by tannins, a tannin removal procedure was performed on the most active extracts. After this procedure, Alchemilla vulgaris was the most persistently active, proving to owe its activity to compounds other than tannins. Thus, this plant was selected as the most promising and further investigated through bioassay-guided fractionation, which resulted in the isolation of a flavonoid, quercetin-3-O-ß-glucuronide, as confirmed by NMR and HRMS spectra. This compound showed not only a higher activity than other flavonoids with the same aglycone moiety, but was also higher than doxycycline (positive control), the only Federal Drug Administration-approved collagenase inhibitor. The approach employed in this study, namely the integration of metabolomics and bioactivity-guided fractionation, showed great potential as a tool for plant selection and identification of bioactive compounds in natural product research.

Hemidesmus indicus (L.) R. Br. extract inhibits the early step of herpes simplex type 1 and type 2 replication.

Herpes simplex virus types 1 (HSV-1) and 2 (HSV-2) cause several clinically relevant syndromes in both adults and neonates. Despite the availability of efficient anti-HSV agents, the search for new therapeutic approaches is highly encouraged due to the increasing drug resistance of virus strains. Medicinal plants represent a source of potential bioactive compounds. In this context we evaluated the anti-herpetic activity of Hemidesmus indicus (L.) R. Br., a plant widely used in traditional Indian medicine. The hydroalcoholic extract prepared from roots was characterized by NMR and HPLC analysis and assayed in vitro by CPE reduction and virus infectivity assays to define its anti-viral effect. The extract's mechanism of action was investigated by virucidal and time-of-addition assays and by in vitro α-glucosidase inhibitory assay. The extract exhibited a remarkable anti-herpetic activity at 100 mg/mL, at non-cytotoxic concentration, through multiple mechanisms: it reduced the infectivity of viral particles released from infected cells possibly through its anti-ER α-glucosidase inhibitory activity and it inhibited the beginning stage of HSV infection acting as a virucide agent and/or preventing virus attachment to the host cell surface.

Analytical Profiling of Bioactive Phenolic Compounds in Argan (Argania spinosa) Leaves by Combined Microextraction by Packed Sorbent (MEPS) and LC-DAD-MS/MS.

INTRODUCTION: The argan tree (Argania spinosa) is an endemic species from south-western Morocco. Argan-based preparations have been widely used in Moroccan traditional medicine for their biological properties, as well as for several cosmetic purposes. Whereas kernel, pulp of fruit and trunk have been extensively studied for their nutritional and pharmacological effects, relatively little is known about argan tree leaves. OBJECTIVE: The main purpose of the present study is to investigate and characterise the bioactive phenolic fractions in both crude and aqueous extracts derived from argan tree leaves. METHODOLOGY: A qualitative profile of the antioxidant phenolic compounds in argan leaves was obtained by means of structural hypothesis based on UV spectra and mass spectrometric fragmentation patterns. Moreover, selected phenolics were quantified in argan leaves by using a fully validated method based on liquid chromatography coupled to diode array detection and tandem mass spectrometry (LC-DAD-MS/MS). All the extracts were purified by a fast and reliable microextraction by packed sorbent (MEPS) procedure, before analysing them by LC-MS/MS. RESULTS: Based on retention times, mass spectrometric fragmentation and UV spectra, 13 phenolic compounds were identified or tentatively elucidated from crude and aqueous extracts derived from Argania spinosa leaves, while seven compounds were quantified in both extracts. CONCLUSION: The obtained results could represent a first step towards a complete characterisation of the argan plant, its bioactive profiling and the valorisation of its by-products as a source of potentially beneficial bioactive molecules.

Antioxidant and α-glucosidase inhibitory activities of Achillea tenorii.

CONTEXT: There is a need for the discovery of novel natural remedies to prevent and treat metabolic disorders such as hyperglycemia, type II non-insulin-dependent diabetes mellitus, and obesity. Several Achillea species have been utilized for centuries all around the world and are generally considered effective as hypoglycemic. OBJECTIVE: Considering the ethnobotanical uses of Achillea genus, we evaluated the in vitro inhibitory activity of Achillea tenorii Grande (Asteraceae) extract on α-glucosidase, which is a valuable target to prevent and treat metabolic disorders. We also tested its antioxidant activity. Moreover, the phytochemical profile was discussed from a chemotaxonomic point of view. MATERIALS AND METHODS: In vitro α-glucosidase inhibition of crude ethanolic extract obtained from the aerial parts was assayed as well as the in vitro antioxidant activity (ABTS, DPPH, and FRAP-FZ tests) was measured. The extract was characterized from a phytochemical point of view by means of spectroscopic analysis. RESULTS: The extract results endowed with α-glucosidase inhibitory activity (IC50 32 µg/mL) with a particular mechanism of action definable as un-competitive, which differed from the mechanism observed for the best-known α-glucosidase inhibitor (acarbose and miglitol). In addition, a considerable antioxidant potential has been found for A. tenorii extract, which resulted mainly constituted by phenolic compounds such as caffeoylquinic acids and flavonoids. DISCUSSION AND CONCLUSIONS: These results suggest the potential of A. tenorii as a possible natural remedy to prevent and treat metabolic disorders of carbohydrates.

Saving the local tradition: ethnobotanical survey on the use of plants in Bologna district (Italy).

BACKGROUND: Traditional knowledge about plants is unfortunately subjected to a progressive loss, mainly due to globalization and depopulation of the rural areas. This work enhances the ethnobotanical knowledge from Northern Italy, specifically Bologna district, and contributes to preserving Italy's plant-based traditional knowledge and to valorize local resources also in view of an ecological transition. METHODS: The study was conducted between 2010 and 2016 in Bologna district encompassing 22 municipalities, which were grouped into three areas: hill, mountain, and plain. In total, 1172 key informants were interviewed, ranging in age from 50 to 85 years, and having strong links with traditional activities in the area. RESULTS: The final inventory included 374 taxa belonging to 91 families. Among these, 251 were wild native, 40 wild alien, 74 cultivated and 6 were products bought from the market. Hill, mountain, and plain provided information on 278, 213, and 110 taxa, respectively. The most cited families were Asteraceae, Lamiaceae, and Rosaceae. The information was systematized in 12 use categories (UC): medicinal (MED), food, cosmetic, domestic, superstitious-magical-religious (SMR), agropastoral, craft, repellent-insecticide, veterinary, toxic, games, other uses and information. The most relevant UC were in turn divided into subcategories. A descriptive table with all the results was also created. MED was the most relevant UC (310 taxa), and among the 17 MED subcategories, the most significant ones were: gastroenteric (160 taxa), respiratory (133 taxa), and dermatologic (122 taxa). Food was also relevant (197 taxa, and 16 subcategories), and the widest food subcategory was nutraceutical (98 taxa). In cosmetic, the most relevant subcategory was skin treatment (37 taxa). Within SMR, the majority of the plants were cited to heal a disease in a ritual or superstitious way (15 taxa), while for agropastoral, the majority of the taxa (29) were cited as feed. CONCLUSIONS: The data collected has highlighted a significant traditional use of plants in Bologna district. Some plants or uses emerged for the first time from an ethnobotanical study carried out in Italy. The inclusion of a large number of municipalities and informants enabled the collection of a wide spectrum of data, encompassing various uses, anecdotes, and historical curiosities, which are crucial to preserve from being forgotten.

Citrus Pomace as a Source of Plant Complexes to Be Used in the Nutraceutical Field of Intestinal Inflammation.

This study aims to recover the main by-product of Citrus fruits processing, the raw pomace, known also as pastazzo, to produce plant complexes to be used in the treatment of inflammatory bowel disease (IBD). Food-grade extracts from orange (OE) and lemon (LE) pomace were obtained by ultrasound-assisted maceration. After a preliminary phytochemical and biological screening by in vitro assays, primary and secondary metabolites were characterized by proton nuclear magnetic resonance (1H-NMR) and liquid chromatography coupled to diode array detection and electrospray ionization mass spectrometry (LC-DAD-ESI-MS) analyses. The intestinal bioaccessibility and antioxidant and anti-inflammatory properties were investigated by in vitro simulated gastro-intestinal digestion followed by treatments on a lipopolysaccharide (LPS)-stimulated human colorectal adenocarcinoma cell line (Caco-2). The tight junctions-associated structural proteins (ZO-1, Claudin-1, and Occludin), transepithelial electrical resistance (TEER), reactive oxygen species (ROS)-levels, expression of some key antioxidant (CAT, NRF2 and SOD2) and inflammatory (IL-1ß, IL-6, TNF-α, IL-8) genes, and pNFkB p65 nuclear translocation, were evaluated. The OE and LE digesta, which did not show any significant difference in terms of phytochemical profile, showed significant effects in protecting against the LPS-induced intestinal barrier damage, oxidative stress and inflammatory response. In conclusion, both OE and LE emerged as potential candidates for further preclinical studies on in vivo IBD models.

Exploring the Anti-Inflammatory and Antioxidant Potential, Metabolite Composition and Inorganic Profile of Cistus monspeliensis L. Aerial Parts and Roots.

This work focuses on Cistus monspeliensis L. aerial parts (AP) and roots (R) extracts, investigating the anti-inflammatory and antioxidant potential of the two organs in comparison. At dosages between 1.56 and 6.25 µg/mL, both extracts showed a protective effect against LPS inflammatory stimulus on a macrophage cell line (RAW 264.7). Interestingly, only R was able to significantly reduce both IL-1ß and IL-6 mRNA gene expression in the presence of LPS. Moreover, the treatment of a neuroblastoma cell line (SH-SY5Y) with AP and R at 6.25 µg/mL increased the cell survival rate by nearly 20% after H2O2 insult. However, only R promoted mitochondria survival, exhibited a significantly higher production of ATP and a higher activity of the enzyme catalase than the control. Both AP and R had similar primary metabolites; in particular, they both contained 1-O-methyl-epi-inositol. Labdane and methoxylated flavonoids were the most characteristic compounds of AP, while R contained mainly catechins, gallic acid, and pyrogallol derivatives. Considering the importance of elemental composition in plants, the inorganic profile of AP and R was also investigated and compared. No potentially toxic elements, such as Pb, were detected in any sample.

Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors.

Novel treatments in gastrointestinal stromal tumors (GISTs) are essential due to imatinib resistance and the modest results obtained with multi-target tyrosine kinase inhibitors. We investigated the possibility that the hydroalcoholic extract from the leaves of Arbutus unedo L. (AUN) could harbor novel chemotherapeutics. The bio-guided fractionation of AUN led to a subfraction, FR2-A, that affected the viability of both imatinib-sensitive and -resistant GIST cells. Cells treated with FR2-A were positive for Annexin V staining, a marker of apoptosis. A rapid PARP-1 downregulation was observed, although without the traditional caspase-dependent cleavage. The fractionation of FR2-A produced nine further active subfractions (FRs), indicating that different molecules contributed to the effect promoted by FR2-A. NMR analysis revealed that pyrogallol-bearing compounds, such as gallic acid, gallic acid hexoside, gallocatechin, myricetin hexoside, and trigalloyl-glucose, are the main components of active FRs. Notably, FRs similarly impaired the viability of GIST cells and peripheral blood mononuclear cells (PBMCs), suggesting a non-specific mechanism of action. Nevertheless, despite the lack of specificity, the established FRs showed promising chemotherapeutic properties to broadly affect the viability of GIST cells, including those that are imatinib-resistant, encouraging further studies to investigate whether pyrogallol-bearing compounds could represent an alternative avenue in GISTs.

Phytochemical Profile and In Vitro Bioactivities of Plant-Based By-Products in View of a Potential Reuse and Valorization.

Wastes and by-products of plant origin are of particular interest to develop a circular economy approach, which attempts to turn them into resources. In this work, thirty-seven neglected plant matrices, including agricultural residues, pest plants, and by-products from the herbal and food industry were extracted and tested for their in vitro anti-tyrosinase, antioxidant, and antibacterial activity against the phytopathogens Pseudomonas syringae pv. syringae ATCC 19310 and Clavibacter michiganensis subsp. nebraskense ATCC 27822. Antioxidant activity ranged from 0.3 to 5 mg of Tr. eq/mL of plant extract, and extract of Castanea sativa pericarp (Csp), Rosa damascena buds (post-distillation) (Rod), and Prunus amygdalus exocarp and mesocarp (Pam) were the most powerful ones. Csp was also capable of inhibiting tyrosinase (IC50 = 16.5 µg/mL), as well as three distillation by-products, namely: Cupressus sempervirens (Css) (IC50 = 95.5 µg/mL), Salvia officinalis (Sco) (IC50 = 87.6 µg/mL), and Helichrysum italicum (Hei) (IC50 = 90.1 µg/mL). Five residues from distillation showed antibacterial activity against C. michiganensis (MICs ranging from 0.125 to 1 mg/mL), namely: Salvia sclarea L. (Sas), Salvia rosmarinus Schleid (Sar), Sco, Hei, and Css. The 1H NMR fingerprinting of the bioactive matrices was acquired, detecting primary and secondary metabolites (rosmarinic acid, shikimic acid, sclareol, and hydroxycinnamic acids).

Metabolomics and genetics of reproductive bud development in Ficus carica var. sativa (edible fig) and in Ficus carica var. caprificus (caprifig): similarities and differences.

In figs, reproductive biology comprises cultivars requiring or not pollination, with female trees (edible fig) and male trees (caprifig) bearing different types of fruits. Metabolomic and genetic studies may clarify bud differentiation mechanisms behind the different fruits. We used a targeted metabolomic analysis and genetic investigation through RNA sequence and candidate gene investigation to perform a deep analysis of buds of two fig cultivars, 'Petrelli' (San Pedro type) and 'Dottato' (Common type), and one caprifig. In this work, proton nuclear magnetic resonance (1H NMR-based metabolomics) has been used to analyze and compare buds of the caprifig and the two fig cultivars collected at different times of the season. Metabolomic data of buds collected on the caprifig, 'Petrelli', and 'Dottato' were treated individually, building three separate orthogonal partial least squared (OPLS) models, using the "y" variable as the sampling time to allow the identification of the correlations among metabolomic profiles of buds. The sampling times revealed different patterns between caprifig and the two edible fig cultivars. A significant amount of glucose and fructose was found in 'Petrelli', differently from 'Dottato', in the buds in June, suggesting that these sugars not only are used by the ripening brebas of 'Petrelli' but also are directed toward the developing buds on the current year shoot for either a main crop (fruit in the current season) or a breba (fruit in the successive season). Genetic characterization through the RNA-seq of buds and comparison with the literature allowed the identification of 473 downregulated genes, with 22 only in profichi, and 391 upregulated genes, with 21 only in mammoni.

Design and Characterization of an Ethosomal Gel Encapsulating Rosehip Extract.

Rising environmental awareness drives green consumers to purchase sustainable cosmetics based on natural bioactive compounds. The aim of this study was to deliver Rosa canina L. extract as a botanical ingredient in an anti-aging gel using an eco-friendly approach. Rosehip extract was first characterized in terms of its antioxidant activity through a DPPH assay and ROS reduction test and then encapsulated in ethosomal vesicles with different percentages of ethanol. All formulations were characterized in terms of size, polydispersity, zeta potential, and entrapment efficiency. Release and skin penetration/permeation data were obtained through in vitro studies, and cell viability was assessed using an MTT assay on WS1 fibroblasts. Finally, ethosomes were incorporated in hyaluronic gels (1% or 2% w/v) to facilitate skin application, and rheological properties were studied. Rosehip extract (1 mg/mL) revealed a high antioxidant activity and was successfully encapsulated in ethosomes containing 30% ethanol, having small sizes (225.4 ± 7.0 nm), low polydispersity (0.26 ± 0.02), and good entrapment efficiency (93.41 ± 5.30%). This formulation incorporated in a hyaluronic gel 1% w/v showed an optimal pH for skin application (5.6 ± 0.2), good spreadability, and stability over 60 days at 4 °C. Considering sustainable ingredients and eco-friendly manufacturing technology, the ethosomal gel of rosehip extract could be an innovative and green anti-aging skincare product.

By-Product Extracts from Castanea sativa Counteract Hallmarks of Neuroinflammation in a Microglial Model.

Castanea sativa is very common in Italy, and the large amount of waste material generated during chestnut processing has a high environmental impact. Several studies demonstrated that chestnut by-products are a good source of bioactive compounds, mainly endowed with antioxidant properties. This study further investigates the anti-neuroinflammatory effect of chestnut leaf and spiny bur extracts, together with the deepest phytochemical characterisation (by NMR and MS) of active biomolecules contained in leaf extracts, which resulted in being more effective than spiny bur ones. BV-2 microglial cells stimulated with lipopolysaccharide (LPS) were used as a model of neuroinflammation. In BV-2 cells pre-treated with chestnut extracts, LPS signalling is partially blocked via the reduced expression of TLR4 and CD14 as well as the expression of LPS-induced inflammatory markers. Leaf extract fractions revealed the presence of specific flavonoids, such as isorhamnetin glucoside, astragalin, myricitrin, kaempferol 3-rhamnosyl (1-6)(2″-trans-p-coumaroyl)hexoside, tiliroside and unsaturated fatty acids, all of which could be responsible for the observed anti-neuroinflammatory effects. Interestingly, the kaempferol derivative has been identified in chestnut for the first time. In conclusion, the exploitation of chestnut by-products is suitable for the achievement of two goals: satisfaction of consumers' demand for new, natural bio-active compounds and valorisation of by-products.

Attenuation of neuroinflammation in microglia cells by extracts with high content of rosmarinic acid from in vitro cultured Melissa officinalis L. cells.

Plant cell culture is a biotechnology cultivation method that permit to cultivate plants in a short period of time and to obtain extracts with a high degree of standardization and high safety profile. The aim of our study was to evaluate the anti-inflammatory and neuroprotective activity of a standardized Melissa officinalis L. phytocomplex extract (MD) obtained with an in vitro plant cell culture. The MD has been chemically characterized and the content of total polyphenols was 5.17 ± 0.1 % w/w, with a content of rosmarinic acid (RA), its main constituent, of 4.02 ± 0.1 % w/w. MD was tested in an in vitro model of neuroinflammation, in which microglia cells (BV2) were stimulated with Lipopolysaccharides (LPS; 250 ng/mL) for 24 h and its pharmacological activity was compared with that of RA. MD (10 µg/mL) and RA (0.4 µg/mL) reduced pro-inflammatory factors (NF-kB, HDAC, IL-1ß) in LPS-stimulated BV2 cells and counteracted the toxic effect produced by activated microglia medium on neuronal cells. This work shows the efficacy of MD on reducing microglia-mediated neuroinflammation and promoting neuroprotection, highlighting the innovative use of in vitro plant cell cultures to obtain contaminant-free extracts endowed with marked activity and improved quali-quantitative ratio in the constituents' content.