Therapeutic potential of plant-derived extracellular vesicles as nanocarriers for exogenous miRNAs.

Cell-to-cell communication strategies include extracellular vesicles (EVs) in plants and animals. The bioactive molecules in a diet rich in vegetables and fruits are associated with disease-preventive effects. Plant-derived EVs (PDEVs) are biogenetically and morphologically comparable to mammalian EVs and transport bioactive molecules, including miRNAs. However, the biological functions of PDEVs are not fully understood, and standard isolation protocols are lacking. Here, PDEVs were isolated from four foods with a combination of ultracentrifugation and size exclusion chromatography, and evaluated as vehicles for enhanced transport of synthetic miRNAs. In addition, the role of food-derived EVs as carriers of dietary (poly)phenols and other secondary metabolites was investigated. EVs from broccoli, pomegranate, apple, and orange were efficiently isolated and characterized. In all four sources, 4 miRNA families were present in tissues and EVs. miRNAs present in broccoli and fruit-derived EVs showed a reduced RNase degradation and were ferried inside exposed cells. EVs transfected with a combination of ath-miR159a, ath-miR162a-3p, ath-miR166b-3p, and ath-miR396b-5p showed toxic effects on human cells, as did natural broccoli EVs alone. PDEVs transport trace amounts of phytochemicals, including flavonoids, anthocyanidins, phenolic acids, or glucosinolates. Thus, PDEVs can act as nanocarriers for functional miRNAs that could be used in RNA-based therapy.

Microbiologically influenced corrosion on naval carbon steel inside the hull of tugboats: a case study of prevention and control.

Microbiologically influenced corrosion (MIC) has a significant cost to many industries, including naval engineering. In this case-of-study, three tugboats developed pitting corrosion in the carbon steel of the inner hulls. Grade A naval steel was used for the hull sheets but the inner side (corroded) showed only two protective layers of paint. The maintenance employed seawater, which ended up in the bilge and made MIC possible. Bilge's waters were submitted to physicochemical, biological and molecular tests. DNA analyses confirmed the presence of Pseudomonas spp. and Desulfovibrio spp. in water samples and, consequently, a MIC mechanism was proposed to explain the corrosion process. In addition, a biocide treatment was evaluated and a new maintenance protocol was recommended. This work highlights the importance of the engineering design to prevent MIC in marine transports and provides some guidelines to treat it.

Antinociceptive and antiedema effects produced in rats by Brassica oleracea var. italica sprouts involving sulforaphane.

Natural products are recognized as potential analgesics since many of them are part of modern medicine to relieve pain without serious adverse effects. The aim of this study was to investigate the antinociceptive and anti-inflammatory activities of an aqueous extract of Brassica oleracea var. italica sprouts (AEBS) and one of its main reported bioactive metabolites sulforaphane (SFN). Antinociceptive activity of the AEBS (30, 100, and 300 mg/kg, i.p. or 1000 and 2000 mg/kg, p.o.) and SFN (0.1 mg/kg, i.p.) was evaluated in the plantar test in rats to reinforce its analgesic-like activity at central level using the reference drug tramadol (TR, 50 mg/kg, i.p.). The anti-inflammatory-like response was determined in the carrageenan-induced oedema at the same dosages for comparison with ketorolac (KET, 20 mg/kg, i.p.) or indomethacin (INDO, 20 mg/kg, p.o.). A histological analysis of the swollen paw was included to complement the anti-inflammatory response. Additionally, acute toxicity observed in clinical analgesics as the most common adverse effects, such as sedation and/or gastric damage, was also explored. As a result, central and peripheral action of the AEBS was confirmed using enteral and parenteral administration, in which significant reduction of the nociceptive and inflammatory responses resembled the effects of TR, KET, or INDO, respectively, involving the presence of SFN. No adverse or toxic effects were observed in the presence of the AEBS or SFN. In conclusion, this study supports that Brassica oleracea var. italica sprouts are a potential source of antinociceptive natural products such as SFN for therapy of pain alone and associated to an inflammation condition.

Dietary-Fibre-Rich Fractions Isolated from Broccoli Stalks as a Potential Functional Ingredient with Phenolic Compounds and Glucosinolates.

The Brassica oleracea industry generates large amounts of by-products to which value could be added because of the characteristics of their composition. The aim was to extract different fibre fractions from broccoli stalks to obtain potential new added-value ingredients. Using an ethanol and water extraction procedure, two fibre-rich fractions (total fibre fraction, TFB, and insoluble fibre fraction, IFB) were obtained. These fractions were analysed to determine the nutritional, (poly)phenols and glucosinolates composition and physicochemical properties, comparing the results with those of freeze-dried broccoli stalks (DBS). Although TFB showed a higher content of total dietary fibre, IFB had the same content of insoluble dietary fibre as TFB (54%), better hydration properties, higher content of glucosinolates (100 mg/100 g d.w.) and (poly)phenols (74.7 mg/100 g d.w.). The prebiotic effect was evaluated in IFB and compared with DBS by in vitro fermentation with human faecal slurries. After 48 h, the short-chain fatty acid (SCFA) production was higher with IFB than with DBS because of the greater presence of both uronic acids, the main component of pectin, and (poly)phenols. These results reveal that novel fibre-rich ingredients-with antioxidant, technological and physiological effects-could be obtained from broccoli stalks by using green extraction methods.

Anti-Leukemic Activity of Brassica-Derived Bioactive Compounds in HL-60 Myeloid Leukemia Cells.

Acute myeloid leukemia (AML) is a cancer of the myeloid blood cells mainly treated with chemotherapy for cancer remission, but this non-selective treatment also induces numerous side effects. Investigations with bioactive compounds from plant-derived foods against cancer have increased in the last years because there is an urgent need to search for new anti-leukemic agents possessing higher efficacy and selectivity for AML cells and fewer negative side effects. In this study, we analyzed the anti-leukemic activity of several phytochemicals that are representative of the major classes of compounds present in cruciferous foods (glucosinolates, isothiocyanates, hydroxycinnamic acids, flavonols, and anthocyanins) in the human acute myeloid leukemia cell line HL-60. Our results revealed that among the different Brassica-derived compounds assayed, sulforaphane (SFN) (an aliphatic isothiocyanate) showed the most potent anti-leukemic activity with an IC50 value of 6 µM in dose-response MTT assays after 48 h of treatment. On the other hand, chlorogenic acid (a hydroxycinnamic acid) and cyanidin-3-glucoside (an anthocyanin) also displayed anti-leukemic potential, with IC50 values of 7 µM and 17 µM after 48 h of incubation, respectively. Importantly, these compounds did not show significant cell toxicity in macrophages-like differentiated cells at 10 and 25 µM, indicating that their cytotoxic effects were specific to AML cancer cells. Finally, we found that these three compounds were able to induce the NRF2/KEAP1 signaling pathway in a dose-dependent manner, highlighting SFN as the most potent NRF2 activator. Overall, the present evidence shed light on the potential for using foods and ingredients rich in anticancer bioactive phytochemicals from Brassica spp.

Potential of Sulforaphane and Broccoli Membrane Vesicles as Regulators of M1/M2 Human Macrophage Activity.

Macrophages have emerged as important therapeutic targets in many human diseases. The aim of this study was to analyze the effect of broccoli membrane vesicles and sulphoraphane (SFN), either free or encapsulated, on the activity of human monocyte-derived M1 and M2 macrophage primary culture. Our results show that exposure for 24 h to SFN 25 µM, free and encapsulated, induced a potent reduction on the activity of human M1 and M2 macrophages, downregulating proinflammatory and anti-inflammatory cytokines and phagocytic capability on C. albicans. The broccoli membrane vesicles do not represent inert nanocarriers, as they have low amounts of bioactive compounds, being able to modulate the cytokine production, depending on the inflammatory state of the cells. They could induce opposite effects to that of higher doses of SFN, reflecting its hormetic effect. These data reinforce the potential use of broccoli compounds as therapeutic agents not only for inflammatory diseases, but they also open new clinical possibilities for applications in other diseases related to immunodeficiency, autoimmunity, or in cancer therapy. Considering the variability of their biological effects in different scenarios, a proper therapeutic strategy with Brassica bioactive compounds should be designed for each pathology.

Contribution of the diverse experimental models to unravelling the biological scope of dietary (poly)phenols.

The health benefits associated with (poly)phenols need to be supported by robust and insightful information on their biological effects. The use of in vitro, ex vivo, and in vivo models is crucial to demonstrate functionalities in specific targets. In this regard, bioaccessibility, bioavailability, and tissue/organ distribution need to be fully understood and established. In addition, the structure-function relationships, concerning both descriptive and mechanistic information, between specific compounds and therapeutic objectives, need to be supported by results obtained from in vivo studies. Nevertheless, these studies are not always possible or have some limitations, particularly concerning the mechanistic information explaining the health benefits provided that should be covered with complementary experimental models. Based on these premises, this review aims to overview the contribution of the separate experimental approaches to gain insights into the bioaccessibility, bioavailability, and bioactivity of (poly)phenols. To achieve this objective, recent evidence available on the linkage of healthy/functional foods with the incidence of non-communicable pathologies is presented. The different experimental approaches provide complementary information that allows advances to be applied to the knowledge gained on the functional properties and mechanistic facts responsible for the health attributions of polyphenols. © 2022 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Maqui berry (Aristotelia chilensis) extract improves memory and decreases oxidative stress in male rat brain exposed to ozone.

Introduction: Prolonged ozone exposure can produce a state of oxidative stress, which in turn causes alterations in the dynamics of the brain and affects memory and learning. Moreover, different investigations have shown that high flavonoid content berries show a great antioxidant activity. The relationship between the protective effect of the maqui berry extract and its antioxidant properties in the brain has not been studied in depth. Objectives: The present study evaluated whether the protection exerted by the aqueous extract of maqui berry in brain regions associated with cognitive performance is due to its antioxidant capacity. Methods: Sprague Dawley rats were exposed to 0.25 ppm ozone and administered with maqui berry extracts. At the end of the treatments, spatial learning and short- and long-term memory were evaluated, as well as oxidative stress markers. Results: The administration of 50 and 100 mg/kg of the aqueous extract of maqui berry was effective in preventing the cognitive deficit caused by chronic exposure to ozone. The antioxidant effect of the administration of maqui berry was analyzed in the prefrontal cortex, hippocampus, and amygdala. Oxidative stress markers levels decreased and the enzymatic activity of superoxide dismutase diminished in animals exposed to ozone treated with the 50 mg/kg dose of maqui berry. Discussion: These results show a relationship between protection at the cognitive level and a decrease in oxidative stress markers, which suggests that the prevention of cognitive damage is due to the antioxidant activity of the maqui berry.

Evidence on the Bioaccessibility of Glucosinolates and Breakdown Products of Cruciferous Sprouts by Simulated In Vitro Gastrointestinal Digestion.

Cruciferous vegetables are gaining importance as nutritious and sustainable foods, rich in phytochemical compounds such as glucosinolates (GSLs). However, the breakdown products of these sulfur-based compounds, mainly represented by isothiocyanates (ITC) and indoles, can contribute to human health. In the human digestive system, the formation of these compounds continues to varying extents in the different stages of digestion, due to the contact of GSLs with different gastric fluids and enzymes under the physicochemical conditions of the gastrointestinal tract. Therefore, the aim of the present work was to uncover the effect of gastrointestinal digestion on the release of glucosinolates and their transformation into their bioactive counterparts by applying a simulated in vitro static model on a range of brassica (red radish, red cabbage, broccoli, and mustard) sprouts. In this sense, significantly higher bioaccessibility of ITC and indoles from GSLs of red cabbage sprouts was observed in comparison with broccoli, red radish, and mustard sprouts, due to the aliphatic GSLs proportion present in the different sprouts. This indicates that the bioaccessibility of GSLs from Brasicaceae sprouts is not exclusively associated with the initial content of these compounds in the plant material (almost negligible), but also with the release of GSLs and the ongoing breakdown reactions during the gastric and intestinal phases of digestion, respectively. Additionally, aliphatic GSLs provided higher bioaccessibility of their corresponding ITC in comparison to indolic and aromatic GSLs.

The Differential Vertical Distribution of the Airborne Biological Particles Reveals an Atmospheric Reservoir of Microbial Pathogens and Aeroallergens.

The most abundant biological particles present in the air are bacteria, fungal propagules and pollen grains. Many of them are proved allergens or even responsible for airborne infectious diseases, which supports the increase of studies in recent years on their composition, diversity, and factors involved in their variability. However, most studies in urban areas are conducted close to ground level and a factor such as height is rarely taken into account. Thus, the information about how the composition of biological particles changes with this variable is scarce. Here, we examined the differential distribution of bacteria, fungi, and plants at four altitudes (up to ∼ 250 m) in a metropolitan area using high-throughput DNA sequencing. Most taxa were present at all levels (common taxa). However, a transitional layer between 80 and 150 m seemed to affect the scattering of these bioaerosols. Taxa not present at all altitudes (non-common) showed an upward tendency of diversity for bacteria and plants with height, while the opposite trend was observed for fungi. Certain patterns were observed for fungi and specific plant genera, while bacterial taxa showed a more arbitrary distribution and no patterns were found. We detected a wide variety of aeroallergens and potential pathogens at all heights, which summed a substantial portion of the total abundance for fungi and plants. We also identified potential connections between the biological particles based on their abundances across the vertical section.

Artemisia pollen is the main vector for airborne endotoxin.

BACKGROUND: Endotoxin (LPS) released from gram-negative bacteria causes strong immunologic and inflammatory effects and, when airborne, can contribute to respiratory conditions, such as allergic asthma. OBJECTIVES: We sought to identify the source of airborne endotoxin and the effect of this endotoxin on allergic sensitization. METHODS: We determined LPS levels in outdoor air on a daily basis for 4 consecutive years in Munich (Germany) and Davos (Switzerland). Air was sampled as particulate matter (PM) greater than 10 µm (PM > 10) and PM between 2.5 and 10 µm. LPS levels were determined by using the recombinant Factor C assay. RESULTS: More than 60% of the annual endotoxin exposure was detected in the PM > 10 fraction, showing that bacteria do not aerosolize as independent units or aggregates but adhered to large particles. In Munich 70% of annual exposure was detected between June 12th and August 28th. Multivariate modeling showed that endotoxin levels could be explained by phenological parameters (ie, plant growth). Indeed, days with high airborne endotoxin levels correlated well with the amount of Artemisia pollen in the air. Pollen collected from plants across Europe (100 locations) showed that the highest levels of endotoxin were detected on Artemisia vulgaris (mugwort) pollen, with little on other pollen. Microbiome analysis showed that LPS concentrations on mugwort pollen were related to the presence of Pseudomonas species and Pantoea species communities. In a mouse model of allergic disease, the presence of LPS on mugwort pollen was needed for allergic sensitization. CONCLUSIONS: The majority of airborne endotoxin stems from bacteria dispersed with pollen of only one plant: mugwort. This LPS was essential for inducing inflammation of the lung and allergic sensitization.

Functional Ingredients From Brassicaceae Species: Overview and Perspectives.

Brassicaceae vegetables are important crops consumed worldwide due to their unique flavor, and for their broadly recognized functional properties, which are directly related to their phytochemical composition. Isothiocyanates (ITC) are the most characteristic compounds, considered responsible for their pungent taste. Besides ITC, these vegetables are also rich in carotenoids, phenolics, minerals, and vitamins. Consequently, Brassica's phytochemical profile makes them an ideal natural source for improving the nutritional quality of manufactured foods. In this sense, the inclusion of functional ingredients into food matrices are of growing interest. In the present work, Brassicaceae ingredients, functionality, and future perspectives are reviewed.

Brassica Bioactives Could Ameliorate the Chronic Inflammatory Condition of Endometriosis.

Endometriosis is a chronic, inflammatory, hormone-dependent disease characterized by histological lesions produced by the presence of endometrial tissue outside the uterine cavity. Despite the fact that an estimated 176 million women are affected worldwide by this gynecological disorder, risk factors that cause endometriosis have not been properly defined and current treatments are not efficient. Although the interaction between diet and human health has been the focus of many studies, little information about the correlation of foods and their bioactive derivates with endometriosis is available. In this framework, Brassica crops have emerged as potential candidates for ameliorating the chronic inflammatory condition of endometriosis, due to their abundant content of health-promoting compounds such as glucosinolates and their hydrolysis products, isothiocyanates. Several inflammation-related signaling pathways have been included among the known targets of isothiocyanates, but those involving aquaporin water channels have an important role in endometriosis. Therefore, the aim of this review is to highlight the promising effects of the phytochemicals present in Brassica spp. as major candidates for inclusion in a dietary approach aiming to improve the inflammatory condition of women affected with endometriosis. This review points out the potential roles of glucosinolates and isothiocyanates from Brassicas as anti-inflammatory compounds, which might contribute to a reduction in endometriosis symptoms. In view of these promising results, further investigation of the effect of glucosinolates on chronic inflammatory diseases, either as diet coadjuvants or as therapeutic molecules, should be performed. In addition, we highlight the involvement of aquaporins in the maintenance of immune homeostasis. In brief, glucosinolates and the modulation of cellular water by aquaporins could shed light on new approaches to improve the quality of life for women with endometriosis.

The Role of Brassica Bioactives on Human Health: Are We Studying It the Right Way?

Brassica vegetables and their components, the glucosinolates, have been suggested as good candidates as dietary coadjutants to improve health in non-communicable diseases (NCDs). Different preclinical and clinical studies have been performed in the last decade; however, some concerns have been posed on the lack of established and standardized protocols. The different concentration of bioactive compounds used, time of intervention or sample size, and the lack of blinding are some factors that may influence the studies' outcomes. This review aims to analyze the critical points of the studies performed with Brassica-related biomolecules and propose some bases for future trials in order to avoid biases.

Anti-inflammatory and antinociceptive effects of an ethanol extract from Senna septemtrionalis.

Senna septemtrionalis (Viv.) H.S. Irwin & Barneby (Fabaceae) is a medicinal plant used as a folk remedy for inflammation and pain. The objective of this study was to evaluate the anti-inflammatory and antinociceptive actions of an ethanol extract of Senna septemtrionalis aerial parts (SSE). The in vitro anti-inflammatory effects of SSE were assessed using LPS-stimulated macrophages and the subsequent quantification of the levels of cytokines (IL-6, IL-1ß, and TNF-α) with ELISA kits, nitric oxide (NO), and hydrogen peroxide (H2O2). The in vivo anti-inflammatory actions of SSE were evaluated with the TPA-induced ear oedema test and the carrageenan-induced paw oedema test. The antinociceptive actions of SSE (10-200 mg/kg p.o.) were assessed using three models: two chemical assays (formalin-induced orofacial pain and acetic acid-induced visceral pain) and one thermal assay (hot plate). SSE showed in vitro anti-inflammatory actions with IC50 values calculated as follows: 163.3 µg/ml (IL-6), 154.7 µg/ml (H2O2) and > 200 µg/ml (IL-1ß, TNF-α, and NO). SSE showed also in vivo anti-inflammatory actions in the TPA test (40% of inhibition of ear oedema) and the carrageenan test (ED50 = 137.8 mg/kg p.o.). SSE induced antinociceptive activity in the formalin orofacial pain test (ED50 = 80.1 mg/kg) and the acetic acid-induced writhing test (ED50 = 110 mg/kg). SSE showed no antinociceptive actions in the hot plate assay. The pre-treatment with glibenclamide abolished the antinociceptive action shown by SSE alone. Overall, SSE exerted in vitro and in vivo anti-inflammatory actions, and in vivo antinociceptive effects by the possible involvement of ATP-sensitive K + channels.

Use of elicitation in the cultivation of Bimi® for food and ingredients.

BACKGROUND: Cruciferous foods rich in health-promoting metabolites are of particular interest to consumers as well as being a good source of bioactives-enriched ingredients. Several elicitors have been used to stimulate the biosynthesis and accumulation of secondary metabolites in foods; however, little is known about the response of new hybrid varieties, such as Bimi®, under field-crop production conditions. Therefore, this study was designed to evaluate the effect of salicylic acid (200 µmol L-1 , SA), methyl jasmonate (100 µmol L-1 , MeJA), and their combination on Bimi plant organs (inflorescences and aerial vegetative tissues - stems and leaves). For this, the composition of the glucosinolates present in the tissues was evaluated. Also, aqueous extracts of the plant material, obtained with different times of extraction with boiling water, were studied. RESULTS: The results indicate that the combined treatment (SA + MeJA) significantly increased the content of glucosinolates in the inflorescences and that MeJA was the most effective elicitor in leaves. Regarding the aqueous extracts, the greatest amount of glucosinolates was extracted at 30 min - except for the leaves elicited with MeJA, for which 15 min was optimal. CONCLUSION: The elicitation in the field enriched leaves in glucobrassicin (GB), 4-methoxyglucobrassicin (MGB), and neoglucobrassicin (NGB) and stems and inflorescences in glucoraphanin, 4-hydroxyglucobrassicin, GB, MGB, and NGB. In this way, this enhanced vegetable material favored the presence of bioactives in the extracts, which is of great interest regarding enriched foods and ingredients with added value obtained from them. © 2019 Society of Chemical Industry.

Naturally occurring melatonin: Sources and possible ways of its biosynthesis.

According to recent reports, the global market for melatonin is worth 700 million USD in 2018 and would reach 2,790 million USD by 2025, growing at a CAGR of 18.9% during 2019 to 2025. Having regard to the prevalence of sleep and circadian rhythm disorders and a clear tendency to increase the demand for melatonin, and the current lack of alternative green and cost-efficient technologies of its synthesis, the supply of this remedy will not be enough to guarantee melatonin supply and affordability on a global scale. The emergence of naturally occurring melatonin and its isomers in fermented foods has opened an exciting new research area; there are still, however, some obscure points in the efficient microbiological biosynthesis of melatonin. This review summarizes the research progress and recent evidence related to melatonin and its isomers in various foodstuffs. Additionally, one possible way to synthesize melatonin is also discussed. The evidence pointed out that the presence of melatonin and its isomers is not exclusive for grapes and grape-derived products, because it can be also found in sweet and sour cherries. However, different species of both Saccharomyces and non-Saccharomyces yeasts could be used to obtain melatonin and melatonin isomers in the process of alcoholic fermentation biotechnologically. The availability of L-tryptophan has been a key factor in determining the concentration of indolic compounds produced, and the utilization of probiotic lactic acid bacteria could help in the formation of melatonin isomers during malolactic fermentation. These approaches are environmentally friendly alternatives with a safer profile than conventional ones and could represent the future for sustainable industrial-scale melatonin production.

Preventing legionellosis outbreaks by a quick detection of airborne Legionella pneumophila.

Legionellosis is a severe pneumonic infection caused by inhaling bacteria of the genus Legionella. Most cases reported in the USA and Europe are associated with the species Legionella pneumophila. This Gram-negative bacterium can survive within a wide spectrum of temperatures, and be transmitted via aerosols from multiple aquatic sources: fountains, thermal spas and other water systems. Although the PCR is one of the most popular methods to verify its presence in environmental or clinical samples, the direct application of this technique to ambient air samples is unusual because of the scarce material in the specimens. Here, we have developed a two-PCR assay, carried out over the V3 and V5 hypervariable regions of the 16S rRNA gene, to detect specifically the pathogenic bacteria Legionella pneumophila in outdoor air samples with low concentration of DNA. The application of this protocol does not require culture and retrieves quick results to activate the corresponding public alerts to prevent legionellosis outbreaks.

Comprehensive analysis of different adhesives in aerobiological sampling using optical microscopy and high-throughput DNA sequencing.

The standardization and unification of the procedures to analyze and quantify the airborne pollen concentrations are very important topics. In this work, the effectiveness of the two most used adhesives in aerobiological sampling, silicone prepared with cyclohexane solvent (Silicone) and petroleum jelly (Vaseline), was compared under outdoor conditions. This comparison was carried out using the traditional method based on the identification and quantification by optical microscopy (OM) of the airborne pollen and the novel methodology by high-throughput sequencing analysis (HTS). Globally, the results from both methods of analysis (OM and HTS) showed a good agreement between the two adhesives tested regarding the abundance of the main pollen types present in the samples: Cupressaceae, Olea, Poaceae, Platanus, Quercus. We concluded that the results from both adhesives are comparable data. Furthermore, the comparisons between methodologies, OM vs. HTS, showed that both techniques can accurately identify the most abundant pollen types in the atmosphere for the studied periods, with a good agreement of their relative abundances especially when the airborne pollen diversity is low but showing some divergences as the number of pollen types increases.

Taraxacum officinale and Urtica dioica extracts inhibit dengue virus serotype 2 replication in vitro.

BACKGROUND: Urtica dioica, Taraxacum officinale, Calea integrifolia and Caesalpinia pulcherrima are widely used all over the world for treatment of different illnesses. In Mexico, these plants are traditionally used to alleviate or counteract rheumatism and inflammatory muscle diseases. In the present study we evaluated the activity of aqueous and methanolic extracts of these four plants, on the replication of dengue virus serotype 2 (DENV2). METHODS: Extraction process was carried out in a Soxtherm® system at 60, 85 and 120 °C; a chemical fractionation in silica gel chromatography was performed and compounds present in the active fractions were identified by HPLC-DAD-ESI/MSn. The cytotoxic concentration and the inhibitory effect of extracts or fractions on the DENV2 replication were analyzed in the BHK-21 cell line (plaque forming assay). The half maximal inhibitory concentration (IC50) and the selectivity index (SI) were calculated for the extracts and fractions. RESULTS: The methanolic extracts at 60 °C of T. officinale and U. dioica showed the higher inhibitory effects on DENV2 replication. After the chemical fractionation, the higher activity fraction was found for U. dioica and T. officinale, presenting IC50 values of 165.7 ± 3.85 and 126.1 ± 2.80 µg/ml, respectively; SI values were 5.59 and 6.01 for each fraction. The compounds present in T. officinale, were luteolin and caffeoylquinic acids derivatives and quercertin diclycosides. The compounds in the active fraction of U. dioica, were, chlorogenic acid, quercertin derivatives and flavonol glycosides (quercetin and kaempferol). CONCLUSIONS: Two fractions from U. dioica and T. officinale methanolic extracts with anti-dengue activity were found. The compounds present in both fractions were identified, several recognized molecules have demonstrated activity against other viral species. Subsequent biological analysis of the molecules, alone or in combination, contained in the extracts will be carried out to develop therapeutics against DENV2.