Enhancing the nitric oxide inhibitory activity using a combination of plant essential oils and mixture design approach.

The synergistic effects of essential oils (EOs) from three aromatic plant species, Foeniculum vulgare subsp. piperitum (C.Presl) Bég. (FV), Origanum heracleoticum L. (OH) and Lavandula austroapennina N.G.Passal., Tundis & Upson. (LA), were evaluated for their inhibitory properties on nitric oxide production in RAW 264.7 macrophages stimulated with lipopolysaccharide (LPS). We utilized a Design of Experiments (DoE) methodology to optimize a formulation by combining three Essential Oils (EOs), while simultaneously taking into account two response variables, maximization of NO inhibition with minimum cytotoxicity. The optimal blend of components was predicted, and the statistical outcome's efficacy was experimentally verified. The combination corresponding to 87.7 % FV, 12.3 % LA and 0.0 % OH showed high inhibitory effect (76.3 %) with negligible cytotoxicity (4.5 %). This research provides new information on the interactions among fennel, oregano and lavender essential oils and shows how they can synergistically inhibit in vitro LPS-induced NO production.

Phenolic Compounds from Tropea Red Onion as Dietary Agents for Protection against Heavy Metals Toxicity.

The present study aims to highlight the cell protective effect of Tropea red onion (TRO) hydroalcoholic extract and some of its components against "non-essential" heavy metals. For this purpose, the cytoprotective roles of cyanidin, cyanidin-3-O-glucoside and quercetin against Cd, Hg and Pb and of TRO extract against Hg and Pb have been investigated, and data are reported here. To the best of our knowledge, this is the first detailed evaluation of the protective effect against cell damage induced by "non-essential" heavy metals through the simultaneous administration of cyanidin, cyanidin-3-O-glucoside and quercetin with CdCl2, HgCl2 or PbCl2 and the TRO extract against HgCl2 and PbCl2. Present data are also compared with our previous results from the TRO extract against Cd. The antioxidant capacity of the extract was also determined by the ferric reducing antioxidant power (FRAP) and the bovine brain peroxidation assay. Both of the assays indicated a good antioxidant capacity of the extract. Cell viability and the impact on necrotic cell death were examined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test and lactate dehydrogenase (LDH) release assay. After 24 h of exposure, Caco-2 cell viability decreased by approximately 50% at 0.25 µM for Cd, Hg and Pb and, after 72 h, the ranking order of "non-essential" heavy metal toxicity on cell viability was PbCl2 > CdCl2 > HgCl2. Cell viability was assessed by treating the cells with the biomolecules at doses of 25, 50 and 100 µg/mL for 24 and 72 h. The same analysis was carried out on Caco-2 cells treated with combinations of TRO extract, cyanidin, cyanidin-3-O-glucoside, or quercetin and "non-essential" heavy metals. Treatments with the bioactive metabolites did not significantly improve cell viability. The identical treatment of Caco-2 cells produced instead LDH release, suggesting a decrease in cell viability. Consistently with the finding that TRO extract showed a good antioxidant activity, we suggest that its higher cytotoxicity, compared to that of the individual assayed phytochemicals, may be derived by the combined antioxidant and chelating properties of all the molecules present in the extract. Therefore, from all the acquired experimental evidence, it appears that the TRO extract may be a better promising protective agent against the toxic effect of Cd, Hg and Pb compared to its bioactive metabolites.

Molecular Docking Studies and In Vitro Activity of Paliurus spina-christi Mill Extracts as Pancreatic Lipase Inhibitors.

Obesity is a risk factor for the onset of chronic diseases. One of the most promising approaches to treating obesity consists of reducing dietary fat absorption using extracts from plants because they contain phenolic compounds, especially flavonoids. Paliurus spina-christi, belonging to the Rhamnaceae family, is one of the five species belonging to the Paliurus genus. Herein, the aerial parts of the plant were extracted with methanol through the pressurized cyclic solid-liquid extraction using the Naviglio extractor®. The extracts were analyzed with High Performance Thin Layer Chromatography and investigated for their in vitro biological potential. The phytochemical analysis revealed that rutin has been shown to be the most abundant flavonoid component. The best antiradical activity was observed for the fruit extract with an IC50 value of 53.41 ± 1.24 µg/mL. This extract also has a better inhibitory capacity on lipid peroxidation evaluated at a different time of incubation. Potent lipase inhibitor activity of the extract from fruits was also demonstrated with in vitro experiments. This property can be attributed to a direct interaction of main components of P. spina-christi extract with the human pancreatic enzyme as demonstrated by the results of molecular docking experiments conducted on the crystallographic structures of lipase.

Targeting of mitochondrial fission through natural flavanones elicits anti-myeloma activity.

BACKGROUND: Mitochondrial alterations, often dependent on unbalanced mitochondrial dynamics, feature in the pathobiology of human cancers, including multiple myeloma (MM). Flavanones are natural flavonoids endowed with mitochondrial targeting activities. Herein, we investigated the capability of Hesperetin (Hes) and Naringenin (Nar), two aglycones of Hesperidin and Naringin flavanone glycosides, to selectively target Drp1, a pivotal regulator of mitochondrial dynamics, prompting anti-MM activity. METHODS: Molecular docking analyses were performed on the crystallographic structure of Dynamin-1-like protein (Drp1), using Hes and Nar molecular structures. Cell viability and apoptosis were assessed in MM cell lines, or in co-culture systems with primary bone marrow stromal cells, using Cell Titer Glo and Annexin V-7AAD staining, respectively; clonogenicity was determined using methylcellulose colony assays. Transcriptomic analyses were carried out using the Ion AmpliSeq™ platform; mRNA and protein expression levels were determined by quantitative RT-PCR and western blotting, respectively. Mitochondrial architecture was assessed by transmission electron microscopy. Real time measurement of oxygen consumption was performed by high resolution respirometry in living cells. In vivo anti-tumor activity was evaluated in NOD-SCID mice subcutaneously engrafted with MM cells. RESULTS: Hes and Nar were found to accommodate within the GTPase binding site of Drp1, and to inhibit Drp1 expression and activity, leading to hyperfused mitochondria with reduced OXPHOS. In vitro, Hes and Nar reduced MM clonogenicity and viability, even in the presence of patient-derived bone marrow stromal cells, triggering ER stress and apoptosis. Interestingly, Hes and Nar rewired MM cell metabolism through the down-regulation of master transcriptional activators (SREBF-1, c-MYC) of lipogenesis genes. An extract of Tacle, a Citrus variety rich in Hesperidin and Naringin, was capable to recapitulate the phenotypic and molecular perturbations of each flavanone, triggering anti-MM activity in vivo. CONCLUSION: Hes and Nar inhibit proliferation, rewire the metabolism and induce apoptosis of MM cells via antagonism of the mitochondrial fission driver Drp1. These results provide a framework for the development of natural anti-MM therapeutics targeting aberrant mitochondrial dependencies.

Chemical Profile of Essential Oils of Selected Lamiaceae Plants and In Vitro Activity for Varroosis Control in Honeybees (Apis mellifera).

The most significant ectoparasitic mite of honeybees, Varroa destructor, has a detrimental effect on bee health and honey output. The principal strategy used by the control programs is the application of synthetic acaricides. All of this has resulted in drug resistance, which is now a major worry for beekeeping. As a result, research on alternate products and techniques for mite management is now required. The aim of this study was to determine whether essential oils (EOs) extracted from botanical species of Lamiacae, typical of the Calabria region of Southern Italy, could reduce the population of the mite V. destructor. Among the best-known genera of the Lamiaceae family are oregano, rosemary and thyme, whose EOs were employed in this study. By steam distillation, the EOs were extracted from Origanum vulgare subsp. viridulum (Martrin-Donos) Nyman, Thymus capitatus Hoffmanns. and Link, Thymus longicaulis C.Presl and Salvia rosmarinus Schleid. plant species harvested directly on the Calabrian territory in their balsamic time. Each EO went to the test in vitro (contact toxicity) against V. destructor. Fifty adult female mites, five for each EO and the positive and negative control, were used in each experimental replicate. The positive controls comprised five individuals treated to Amitraz dilute in acetone, and the negative controls included five individuals exposed to acetone alone. To create the working solution to be tested (50 µL/tube), the EOs were diluted (0.5 mg/mL, 1 mg/mL, 2 mg/mL and 4 mg/mL) in HPLC-grade acetone. After 1 h of exposure, mite mortality was manually assessed. Origanum vulgare subsp. viridulum, Thymus capitatus and Thymus longicaulis were the EOs with the highest levels of efficiency at 2 mg/mL, neutralizing (dead + inactivated), 94%, 92% and 94% of parasites, respectively. Salvia rosmarinus EO gave a lower efficacy, resulting in a percentage of 38%. Interestingly, no adverse effects were highlighted in toxicity tests on honeybees. These results show that these OEs of the Lamiaceae family have antiparasitic action on V. destructor. Therefore, they could be used, individually or combined, to exploit the synergistic effect for a more sustainable control of this parasite mite in honeybee farms.

Plants and Their Derivatives as Promising Therapeutics for Sustainable Control of Honeybee (Apis mellifera) Pathogens.

The most important pollinator for agricultural crops is the Western honeybee (Apis mellifera). During the winter and summer seasons, diseases and stresses of various kinds endanger honeybee numbers and production, resulting in expenses for beekeepers and detrimental effects on agriculture and ecosystems. Researchers are continually in search of therapies for honeybees using the resources of microbiology, molecular biology, and chemistry to combat diseases and improve the overall health of these important pollinating insects. Among the most investigated and most promising solutions are medicinal plants and their derivatives. The health of animals and their ability to fight disease can be supported by natural products (NPs) derived from living organisms such as plants and microbes. NPs contain substances that can reduce the effects of diseases by promoting immunity or directly suppressing pathogens, and parasites. This literature review summarises the advances that the scientific community has achieved over the years regarding veterinary treatments in beekeeping through the use of NPs. Their impact on the prevention and control of honeybee diseases is investigated both in trials that have been conducted in the laboratory and field studies.

Interaction of Quercetin, Cyanidin, and Their O-Glucosides with Planar Lipid Models: Implications for Their Biological Effects.

Flavonoids are specialized metabolites produced by plants, as free aglycones or as glycosylated derivatives, which are particularly endowed with a variety of beneficial health properties. The antioxidant, anti-inflammatory, antimicrobial, anticancer, antifungal, antiviral, anti-Alzheimer's, anti-obesity, antidiabetic, and antihypertensive effects of flavonoids are now known. These bioactive phytochemicals have been shown to act on different molecular targets in cells including the plasma membrane. Due to their polyhydroxylated structure, lipophilicity, and planar conformation, they can either bind at the bilayer interface or interact with the hydrophobic fatty acid tails of the membrane. The interaction of quercetin, cyanidin, and their O-glucosides with planar lipid membranes (PLMs) similar in composition to those of the intestine was monitored using an electrophysiological approach. The obtained results show that the tested flavonoids interact with PLM and form conductive units. The modality of interaction with the lipids of the bilayer and the alteration of the biophysical parameters of PLMs induced by the tested substances provided information on their location in the membrane, helping to elucidate the mechanism of action which underlies some pharmacological properties of flavonoids. To our knowledge, the interaction of quercetin, cyanidin, and their O-glucosides with PLM surrogates of the intestinal membrane has never been previously monitored.

Identification of Pinosylvin in Pinus nigra subsp. laricio: A Naturally Occurring Stilbenoid Suppressing LPS-Induced Expression of Pro-Inflammatory Cytokines and Mediators and Inhibiting the JAK/STAT Signaling Pathway.

Stilbenoids, a group of phytoalexin polyphenols produced by plants as a defence mechanism in response to stress conditions, are known for their anti-inflammatory potential. Pinosylvin, a naturally occurring molecule traditionally found in pinus trees, was here identified in Pinus nigra subsp. laricio var. calabrica from Southern Italy through HPLC analysis. Both this molecule and its well-known analogue resveratrol, the most famous wine polyphenol, were compared for their in vitro potential anti-inflammatory activity. Pinosylvin significantly inhibited the release of pro-inflammatory cytokines (TNF-α and IL-6) and NO mediator in LPS-stimulated RAW 264.7 cells. Moreover, its ability to inhibit the JAK/STAT signaling pathway was assessed: Western blot analyses showed a downregulation of both phosphorylated JAK2 and STAT3 proteins. Finally, in order to verify whether this biological activity could be attributed to a direct interaction of pinosylvin with JAK2, a molecular docking study was performed, confirming the capability of pinosylvin to bind the active site of the protein.

Essential Oils for a Sustainable Control of Honeybee Varroosis.

The Varroa destructor parasite is the main obstacle to the survival of honey bee colonies. Pest control mainly involves the use of synthetic drugs which, used with the right criteria and in rotation, are able to ensure that infestation levels are kept below the damage threshold. Although these drugs are easy to use and quick to apply, they have numerous disadvantages. Their prolonged use has led to the emergence of pharmacological resistance in treated parasite populations; furthermore, the active ingredients and/or their metabolites accumulate in the beehive products with the possibility of risk for the end consumer. Moreover, the possibility of subacute and chronic toxicity phenomena for adult honeybees and their larval forms must be considered. In this scenario, eco-friendly products derived from plant species have aroused great interest over the years. In recent decades, several studies have been carried out on the acaricidal efficacy of plant essential oils (EOs). Despite the swarming of laboratory and field studies, however, few EO products have come onto the market. Laboratory studies have often yielded different results even for the same plant species. The reason for this discrepancy lies in the various study techniques employed as well as in the variability of the chemical compositions of plants. The purpose of this review is to take stock of the research on the use of EOs to control the V. destructor parasite. It begins with an extensive discussion of the characteristics, properties, and mechanisms of action of EOs, and then examines the laboratory and field tests carried out. Finally, an attempt is made to standardize the results and open up new lines of study in future.

Cachrys L. Genus: A Comprehensive Review on Botany, Phytochemistry and Biological Properties.

The Cachrys L. genus belongs to the Apiaceae family and it is widely distributed in the Mediterranean basin, with plant species being endemic to southern Europe, Asia, and northern Africa. Different studies, focused on the phytochemical composition of Cachrys spp. and the biological properties of their phytocomplexes, have been reported. These works mostly focused on the essential oils obtained from these plants, and pointed out that Cachrys species are a rich source of coumarins, mainly furanocoumarins. Other phytochemicals, such as terpenes, fatty acids, phytosterols, and flavonoids have been also identified. Moreover, a number of biological properties such as antioxidant, antimicrobial, anti-inflammatory, cytotoxic, and photocytotoxic effects have been assessed. Nevertheless, a review of the chemical and pharmacological properties of this genus is not available in the literature. The aim of this paper is to provide an overview of the reports concerning the identified phytochemicals and the biological effects reported for Cachrys spp., and to offer a comprehensive understanding of the potential of this genus as a source of bioactive compounds. The current taxonomy, the traditional uses, and the toxicological aspects of plants belonging to this genus are also reported, and the future research directions are discussed.

Phytochemical Profile and In Vitro Antioxidant and Photobiological Properties of Different Extracts from Prangos ferulacea Lindl.

Interesting photobiological properties have been demonstrated for some Cachrys species, including C. libanotis L., C. sicula L., and C. pungens Jan. The present study was designed to assess the photocytotoxic activity of Prangos ferulacea Lindl. (synonym of C. ferulacea (L.) Calest.). This plant was previously considered a Cachrys species but, at present, it is part of the Prangos genus. P. ferulacea is an orophilous plant present in the eastern Mediterranean and in western Asia. Three different extraction techniques were utilized. Obtained extracts were compared both for their phytochemical content and for their photobiological properties on human melanoma cells irradiated with UVA light. The apoptotic responses, together with the antioxidant activity, were also assessed. P. ferulacea extracts were able to affect cell viability in a concentration-dependent manner, with the sample obtained through supercritical CO2 extraction showing the highest activity (IC50 = 4.91 µg/mL). This research points out the interesting content in the photoactive compounds of this species, namely furanocoumarins, and could provide a starting point for further studies aimed at finding new photosensitizing agents useful in cancer photochemotherapy.

In vitro antioxidant and anti-denaturation effects of Buglossoides purpurocaerulea (L.) I. M. Johnst. fruit extract.

Fruits from Buglossoides purpurocaerulea (L.) I. M. Johnst. were extracted with methanol through maceration. The phytochemical composition of obtained raw extract was investigated with gas chromatography-mass spectrometry (GC-MS) and high-performance thin layer chromatography (HPTLC) analyses. The total phenolic and flavonoid contents were also quantified and in vitro antioxidant activity was assessed using the DPPH assay and the ß-carotene bleaching test. In order to verify the anti-arthritic potential, the raw extract was evaluated for its anti-denaturation effect on heat-treated bovine serum albumin (BSA), chosen as a protein model. Linoleic and α-linolenic acids were the most abundant identified apolar constituents. HPTLC analyses allowed the identification of rosmarinic acid. The sample showed a concentration-dependent antioxidant activity and it was also proven to be effective in preventing the heat-induced denaturation of BSA, with an IC50 value equal to 15.7 ± 0.5 µg/mL.

Phytochemical Composition and Pharmacological Efficacy Evaluation of Calamintha nepeta, Calamintha sylvatica, Lavandula austroapennina and Mentha piperita Essential Oils for the Control of Honeybee (Apis mellifera) Varroosis.

Varroa destructor is currently considered the parasite that causes the greatest damage and economic losses to honeybee farms. Its presence is often associated with that of viral and bacterial pathogens, which ultimately leads to colony collapse. Careful control of the parasitic load is therefore necessary to avoid the onset of these events. Although chemical treatments are often in easily and quickly administered formulations, in recent years, there have been increasingly frequent reports of the onset of drug resistance phenomena, which must lead to reconsidering their use. Furthermore, chemical compounds can easily accumulate in the food matrices of the hive, with possible risks for the final consumer. In such a condition, it is imperative to find alternative treatment solutions. Essential oils (EOs) prove to be promising candidates due to their good efficacy and good environmental biodegradability. In this study, the acaricidal efficacy of the EOs of Calamintha sylvatica Bromf., Calamintha nepeta Savi, Lavandula austroapennina N.G. Passal. Tundis & Upson and Mentha piperita L., extracted from botanical species belonging to the Lamiaceae family, was evaluated. The test chosen for the evaluation was residual toxicity by contact. The examined EOs were diluted in Acetone to a concentration of 2, 1 and 0.5 mg/mL. At the highest concentration, the EOs demonstrated an acaricidal activity equal to 52% for C. nepeta, 60% for C. sylvatica, 80% for L. austroapennina and 68% for M. piperita. Of the EOs tested, therefore, Lavender proves to be a good candidate for subsequent evaluations in semi-field and field studies.

Phytochemical Profile of Foeniculum vulgare Subsp. piperitum Essential Oils and Evaluation of Acaricidal Efficacy against Varroa destructor in Apis mellifera by In Vitro and Semi-Field Fumigation Tests.

Varroatosis is an important parasitic disease of Apis mellifera caused by the mite Varroa destructor (V. destructor). The parasite is able to transmit numerous pathogens to honeybees which can lead to colony collapse. In recent years, the effectiveness of authorized drug products has decreased due to increasing resistance phenomena. Therefore, the search for alternatives to commercially available drugs is mandatory. In this context, essential oils (EOs) prove to be a promising choice to be studied for their known acaricide properties. In this research work, the acaricide activity of EO vapours isolated from the epigeal part (whole plant) of fennel (Foeniculum vulgare sbps. piperitum) and its three fractions (leaves, achenes and flowers) against V. destructor was evaluated. The effectiveness of fumigation was studied using two methods. The first involved prolonged exposure of mites to oil vapour for variable times. After exposure, the five mites in each replicate were placed in a Petri dish with an Apis mellifera larva. Mortality, due to chronic toxicity phenomena, was assessed after 48 h. The second method aimed to translate the results obtained from the in vitro test into a semi-field experiment. Therefore, two-level cages were set up. In the lower compartment of the cage, a material releasing oil vapours was placed; in the upper compartment, Varroa-infested honeybees were set. The results of the first method showed that the increase in mortality was directly proportional to exposure time and concentration. The whole plant returned 68% mortality at the highest concentration (2 mg/mL) and highest exposure time (48 h control), while the leaves, achenes and flowers returned 64%, 52% and 56% mortality, respectively. In the semi-field experiment, a concentration up to 20 times higher than the one used in the in vitro study was required for the whole plant to achieve a similar mite drop of >50%. The results of the study show that in vitro tests should only be used for preliminary screening of EO activity. In vitro tests should be followed by semi-field tests, which are essential to identify the threshold of toxicity to bees and the effective dose to be used in field studies.

Cachrys spp. from Southern Italy: Phytochemical Characterization and JAK/STAT Signaling Pathway Inhibition.

Different phytochemical compounds have been demonstrated to modulate the JAK/STAT signaling pathway. Here, three Cachrys species from Southern Italy were investigated for both the phytochemical profile and the potential anti-inflammatory properties. The aerial parts were extracted with methanol through Naviglio Extractor®, an innovative solid-liquid extraction technique that allows to obtain high quality extracts by working with gradient pressure. Extracts were analyzed with GC-MS and standardized in furanocoumarin content, resulting rich in xanthotoxin, bergapten and isopimpinellin. Given the known ability of bergapten to inhibit the JAK/STAT signaling pathway by decreasing the levels of pro-inflammatory cytokines (TNF-α, IL-6) and inflammatory mediators (NO) in RAW 264.7 cells activated by LPS, Cachrys extracts were investigated for their biological properties. The results obtained in this study showed that Cachrys pungens extract, presenting the highest content in furanocoumarins (7.48 ± 0.48 and 2.94 ± 0.16 mg/50 mg of extract for xanthotoxin and bergapten, respectively), significantly decreased STAT3 protein levels, pro-inflammatory cytokines (TNF-α, IL-6) and increased IL-10 anti-inflammatory cytokine. Cachrys ferulacea significantly decreased JAK2 phosphorylation, being even more effective than bergapten. In conclusion, investigated extracts could be potential candidates for the search of novel anti-inflammatory agents acting via inhibiting the JAK/STAT signaling pathway.

Antioxidant activity and protective effect of the outer scales hydroalcoholic extract of Allium cepa L. var. Tropea on toxicity damage induced by Cadmium in Caco-2 cells.

The characterization of bioactive metabolites and the protective effects against Cadmium (Cd) of the hydroalcoholic extract from Allium cepa var. Tropea (Tropea red onion) in human Caco-2 colon adenocarcinoma cells and in vitro antioxidant effects were investigated. Tropea red onion extract showed high levels of bioactive compounds and a strong activity as radical scavenger and inhibitor of lipid peroxidation. The most abundant specialized metabolites were quercetin derivatives. In addition, the extract inhibited NO release in a dose-dependent manner. Exposure of Cd treated human Caco-2 cells to the Tropea red onion extract resulted in a higher cytoprotection with a significant falloff of cells damage produced by Cd. The effect of Tropea red onion evaluated with the MTT assay and LDH test, was time-dependent and more evident after 24h of treatment. Caco-2 cells treated with the Tropea red onion extract and CdCl2 at 24h showed a significant cytoprotection at concentrations equal to 50 µg/mL + 25 µM and 100 µg/mL + 25 µM while at 72h at a concentration of 25 µg/mL + 25 µM. We observed with MTT and LDH assays that treatment of cells with Tropea red onion extract at 24h was able to significantly prevent Cd-induced cytotoxicity alone.

Green Veterinary Pharmacology for Honey Bee Welfare and Health: Origanum heracleoticum L. (Lamiaceae) Essential Oil for the Control of the Apis mellifera Varroatosis.

Varroatosis, caused by the Varroa destructor mite, is currently the most dangerous parasitic disease threatening the survival of honey bees worldwide. Its adverse effect on the welfare and health of honey bees requires the regular use of specific acaricides. This condition has led to a growing development of resistance phenomena towards the most frequently used drugs. In addition, another important aspect that should not be understated, is the toxicity and persistence of chemicals in the environment. Therefore, the identification of viable and environmentally friendly alternatives is urgently needed. In this scenario, essential oils are promising candidates. The aim of this study was to assess the contact toxicity, the fumigation efficacy and the repellent effect of Origanum heracleoticum L. essential oil (EO) against V. destructor mite. In the contact tests, each experimental replicate consisted of 15 viable adult female mites divided as follows: 5 treated with EO diluted in HPLC grade acetone, 5 treated with acetone alone (as negative control) and 5 treated with Amitraz diluted in acetone (as positive control). The EO was tested at concentrations of 0.125, 0.25, 0.5, 1 and 2 mg/mL. For each experimental replicate, mortality was manually assessed after one hour. The efficacy of EO fumigation was evaluated through prolonged exposure at different time intervals. After each exposure, the 5 mites constituting an experimental replicate were transferred to a Petri dish containing a honey bee larva and mortality was assessed after 48 h. The repellent action was investigated by implementing a directional choice test in a mandatory route. During the repellency tests the behavior of the mite (90 min after its introduction in the mandatory route) was not influenced by the EO. In contact tests, EO showed the best efficacy at 2 and 1 mg/mL concentrations, neutralizing (dead + inactivated) 90.9% and 80% of the mites, respectively. In fumigation tests, the mean mortality rate of V. destructor at maximum exposure time (90 min) was 60% and 84% at 24 and 48 h, respectively. Overall, these results demonstrate a significant efficacy of O. heracleoticum EO against V. destructor, suggesting a possible alternative use in the control of varroatosis in honey bee farms in order to improve Apis mellifera welfare and health and, consequently, the hive productions.

Ranunculus species suppress nitric oxide production in LPS-stimulated RAW 264.7 macrophages.

Aerial parts and roots from three Ranunculus species were extracted with 70% ethanol. The phytochemical composition was investigated using GC-MS and multivariate data analysis. The total phenolic and flavonoid contents were also assessed. The in vitro inhibitory properties were evaluated as nitrite concentration in LPS-stimulated RAW 264.7 macrophage cell line. All the samples induced concentration-dependent inhibitory effects, with R. sceleratus aerial parts extract being the most interesting sample (IC50 = 22.08 ± 1.32 µg/mL), even more active than the reference compound indomethacin. A very good activity was also observed for R. sardous and R. ficaria aerial parts extracts (IC50 = 51.61 ± 3.12 µg/mL and 84.55 ± 3.40 µg/mL). A lesser but noteworthy potential was also demonstrated for the root extracts. The results suggest that Ranunculus hydroalcoholic extracts are able to inhibit nitrite accumulation and may be useful in preventing inflammatory diseases mediated by excessive production of NO.

Centranthus ruber (L.) DC. and Tropaeolum majus L.: Phytochemical Profile, In Vitro Anti-Denaturation Effects and Lipase Inhibitory Activity of Two Ornamental Plants Traditionally Used as Herbal Remedies.

Ornamental plants often gain relevance not only for their decorative use, but also as a source of phytochemicals with interesting healing properties. Herein, spontaneous Centranthus ruber (L.) DC. and Tropaeolum majus L., mainly used as ornamental species but also traditionally consumed and used in popular medicine, were investigated. The aerial parts were extracted with methanol trough maceration, and resultant crude extracts were partitioned using solvents with increasing polarity. As previous studies mostly dealt with the phenolic content of these species, the phytochemical investigation mainly focused on nonpolar constituents, detected with GC-MS. The total phenolic and flavonoid content was also verified, and HPTLC analyses were performed. In order to explore the potential antiarthritic and anti-obesity properties, extracts and their fractions were evaluated for their anti-denaturation effects, with the use of the BSA assay, and for their ability to inhibit pancreatic lipase. The antioxidant properties and the inhibitory activity on the NO production were verified, as well. Almost all the extracts and fractions demonstrated good inhibitory effects on NO production. The n-hexane and dichloromethane fractions from T. majus, as well as the n-hexane fraction from C. ruber, were effective in protecting the protein from heat-induced denaturation (IC50 = 154.0 ± 1.9, 270.8 ± 2.3 and 450.1 ± 15.5 µg/mL, respectively). The dichloromethane fractions from both raw extracts were also effective in inhibiting pancreatic lipase, with IC50 values equal to 2.23 ± 0.02 mg/mL (for C. ruber sample), and 2.05 ± 0.02 mg/mL (T. majus). Obtained results support the traditional use of these species for their beneficial health properties and suggest that investigated plant species could be potential sources of novel antiarthritic and anti-obesity agents.

Sub-Micromolar Inhibition of SARS-CoV-2 3CLpro by Natural Compounds.

Inhibiting the main protease 3CLpro is the most common strategy in the search for antiviral drugs to fight the infection from SARS-CoV-2. We report that the natural compound eugenol is able to hamper in vitro the enzymatic activity of 3CLpro, the SARS-CoV-2 main protease, with an inhibition constant in the sub-micromolar range (Ki = 0.81 µM). Two phenylpropene analogs were also tested: the same effect was observed for estragole with a lower potency (Ki = 4.1 µM), whereas anethole was less active. The binding efficiency index of these compounds is remarkably favorable due also to their small molecular mass (MW < 165 Da). We envision that nanomolar inhibition of 3CLpro is widely accessible within the chemical space of simple natural compounds.