Pharmacological evaluation of the anxiolytic-like effects of an aqueous extract of the Raphanus sativus L. sprouts in mice.

Raphanus sativus L. (Brassicaceae), commonly known as radish, is consumed worldwide as a vegetable. However, its benefits on mental health are unknown. The aim of this study was to evaluate its anxiolytic-like effects and safety using different experimental models. An aqueous extract of R. sativus sprouts (AERSS) was pharmacologically evaluated by intraperitoneal route (i.p.) at 10, 30, and 100 mg/kg and orally (p.o.) at 500 mg/kg on behavior by using open-field and plus-maze tests. In addition, its acute toxicity (LD50) was determined by the Lorke's method. Diazepam (1 mg/kg, i.p.) and buspirone (4 mg/kg, i.p.) were the reference drugs. A significant and anxiolytic-like dosage of AERSS (30 mg/kg, i.p.) resembling the effects of reference drugs was chosen to explore the involvement of GABAA/BDZs site (flumazenil, 5 mg/kg, i.p.) and serotonin 5-HT1A receptors (WAY100635, 1 mg/kg, i.p.) as a possible mechanism of action. A 500 mg/kg, p.o. dosage of AERSS produced an anxiolytic-like response equivalent to 100 mg/kg, i.p. No acute toxicity was observed since a LD50 > 2000 mg/kg, i.p. The phytochemical analysis allowed the identification and quantification of major presence of sulforaphene (2500 µM), sulforaphane (15 µM), iberin (0.75 µM), and indol-3-carbinol (0.75 µM), as major constituents. Both the GABAA/BDZs site and serotonin 5-HT1A receptors were involved in the anxiolytic-like activity of AERSS, depending on the pharmacological parameter or the experimental assay tested. Our results demonstrate that the anxiolytic activity of R. sativus sprouts involves GABAA/BDZs site and serotonin 5-HT1A receptors supporting its health benefits in the treatment of anxiety beyond the satisfaction of basic nutritional needs.

A Systematic Approach to Agastache mexicana Research: Biology, Agronomy, Phytochemistry, and Bioactivity.

Mexico is the center of origin of the species popularly known as toronjil or lemon balm (Agastache mexicana Linton & Epling). Two subspecies have been identified and are commonly called purple or red (Agastache mexicana Linton & Epling subspecies. mexicana) and white (Agastache mexicana subspecies xolocotziana Bye, E.L. Linares & Ramamoorthy). Plants from these subspecies differ in the size and form of inflorescence and leaves. They also possess differences in their chemical compositions, including volatile compounds. Traditional Mexican medicine employs both subspecies. A. mexicana exhibits a broad range of pharmacological properties, such as anti-inflammatory, anxiolytic, and antioxidant. A systematic vision of these plant's properties is discussed in this review, exposing its significant potential as a source of valuable bioactive compounds. Furthermore, this review provides an understanding of the elements that make up the species' holistic system to benefit from lemon balm sustainably.

Antivirulence Activity of a Dietary Phytochemical: Hibiscus Acid Isolated from Hibiscus sabdariffa L. Reduces the Virulence of Pseudomonas aeruginosa in a Mouse Infection Model.

Hibiscus sabdariffa L. (Hs) calyxes, rich in organic acids, are included in diets in different countries. In recent years, some phytochemicals have been shown to reduce bacterial virulence at sublethal concentrations by interfering with quorum sensing (QS) systems. Therefore, in this study the antivirulence properties of Hs calyxes and two γ-lactones (hibiscus acid [HA] and its methyl ester) in Pseudomonas aeruginosa were analyzed. Acetone and methanol extracts of Hs showed anti-QS activity by inhibiting violacein production (60% to 80% with 250 µg/mL). In molecular docking analysis, the γ-lactones registered a good binding score, which suggests strong interaction with the active site of LasR protein. To verify their effect in vitro, they were isolated from Hs and evaluated in six QS-regulated phenotypes, as well as in ExoU toxin that is released by the type III secretion system (T3SS). At 500 µg/mL they reduced alkaline protease (29-52%) and elastase (15-37%) activity, biofilm formation (∼75%), and swarming (50%), but there was no effect on pyocyanin production, hemolytic activity, or type III secretion. In a mouse abscess/necrosis model, HA at sublethal concentrations (15 and 31.2 µg/mL) affected infection establishment and prevented damage and systemic spread. In conclusion, HA is the first molecule identified with antivirulence properties in Hs with the potential to prevent infections caused by P. aeruginosa.

Climate and categories: Two key elements for understanding the Mesoamerican hot-cold classification of illnesses and medicinal plants.

ETHNOPHARMACOLOGICAL RELEVANCE: The concepts of health and illness, and their causes, are fundamental for understanding medicinal plant choice and use by traditional people. The hot-cold system is widespread in Mesoamerican traditional medicine and guides many therapeutic decisions. AIM OF THE STUDY: This study explores a hypothesis that climate influences the hot-cold classification of illnesses and medicinal plants, and the perception of hazard of illnesses. In addition, we examine the classification categories within the system used in different regions of Mexico. MATERIALS AND METHODS: Studies from Mexico with quantitative and qualitative data on the hot-cold properties of medicinal plants and ailments were reviewed. The information was organized and then related to the climate type of the study areas. RESULTS: In temperate climates, most diseases were considered cold, and hot medicinal plants were dominant. Conversely, in warm-tropical climates, hot diseases dominated, and the majority of medicinal plants were cold; however, this evidence was weaker. The perception of hazard was congruent with the number of illnesses for temperate climates. There were additional classification categories within the hot-cold system for diseases and medicinal plants, and they were expressed in different terms in Spanish, English, and indigenous languages. Although similar terms and categories were used in the classification of diseases and medicinal plants, they can differ conceptually and vary between places and cultures. Publications are sometimes unclear if the terms used are emic or etic. The basic principle of using plants with the opposite property of the disease does not always apply strictly. CONCLUSIONS: Climate appears to influence the hot-cold classification of diseases and medicinal plants in Mexico, and the system is not strictly dual. Improved knowledge of the hot-cold system is necessary to understand Mesoamerican medicinal plant use and culture.

Phytochemical Analysis and Antioxidant and Anti-Inflammatory Capacity of the Extracts of Fruits of the Sechium Hybrid.

In addition to their own antioxidants, human cells feed on external antioxidants, such as the phenolic compounds of fruits and vegetables, which work together to keep oxidative stress in check. Sechium edule, an edible species of chayote, has phenolic compounds with antioxidant activity and antineoplastic activity. A Sechium hybrid shows one thousand times greater antineoplastic activity than edible species, but its antioxidant and anti-inflammatory activities and the content of phenolic compounds are unknown. The aim of this study was to determine the antioxidant and anti-inflammatory capacity of the extract of fruits of the Sechium hybrid in vitro and in vivo. Phytochemical analysis using HPLC showed that the extract of the Sechium hybrid has at least 16 phenolic compounds; galangin, naringenin, phloretin and chlorogenic acid are the most abundant. In an in vitro assay, this extract inhibited 2,2-diphenyl-L-picrylhydrazyl (DPPH) activity and protected the dimyristoylphosphatidylethanolamine (DMPE) phospholipid model cell membrane from oxidation mediated by hypochlorous acid (HClO). In vivo, it was identified that the most abundant metabolites in the extract enter the bloodstream of the treated mice. On the other hand, the extract reduces the levels of tumor necrosis factor alpha (TNFα), interferon gamma (IFNγ), and interleukin-6 (IL-6) but increases interleukin-10 (IL-10) and glutathione peroxidase levels. Our findings indicate that intake of the fruits of the Sechium hybrid leads to antioxidant and anti-inflammatory effects in a mouse model. Therefore, these results support the possibility of exploring the clinical effect of this hybrid in humans.

Antiquorum Sensing Activity of Seed Oils from Oleaginous Plants and Protective Effect During Challenge with Chromobacterium violaceum.

Seed oils from oleaginous plants are rich in fatty acids (FAs) that play important roles in the health of the consumers. Recent studies indicate that FA also can play an important role in communication and regulation of virulence in bacteria. Nevertheless, evidence demonstrating protection against bacterial infections mediated by their quorum sensing inhibition (QSI) activity is scarce. In this study, sunflower, chia, and amaranth oils, were assayed for their QSI capacity by inhibiting violacein production and alkaline exoprotease activity of Chromobacterium violaceum. In vitro assays revealed that the oils exhibited QSI activities, whereas in vivo they delayed death of mice inoculated intraperitoneally with the bacterium. Gas chromatography coupled with mass spectrometry analysis of the oils indicated the presence of saturated FA (SAFA) and unsaturated FA as main components. Through a structure-activity relationship study of free FAs, bactericidal effect was identified mainly for polyunsaturated FAs, whereas QSI activity was restricted to SAFA of chains 12-18 carbon atoms in length. These data correlate with a possible interaction suggested by molecular docking analysis of lauric, myristic, and stearic acids with the CviR protein. Our study highlights the antiquorum sensing potential of SAFA, which may be future antivirulence therapeutic agents for the treatment of bacterial infections.

Sechium edule (Jacq.) Swartz, a New Cultivar with Antiproliferative Potential in a Human Cervical Cancer HeLa Cell Line.

The Sechium edule Perla Negra cultivar is a recently-obtained biological material whose progenitors are S. edule var. nigrum minor and S. edule var. amarus silvestrys, the latter of which has been reported to have antiproliferative activity against the HeLa P-388 and L-929 cancer cell lines. The present study aimed to determine if the methanolic extract of the fruit of the Perla Negra cultivar had the same biological activity. The methanolic extract was phytochemically characterized by thin layer chromatography (TLC) and column chromatography (CC), identifying the terpenes and flavonoids. The compounds identified via high performance liquid chromatography (HPLC) were Cucurbitacins B, D, E, and I for the terpene fractions, and Rutin, Phlorizidin, Myricetin, Quercetin, Naringenin, Phloretin, Apigenin, and Galangin for the flavonoid fractions). Biological activity was evaluated with different concentrations of the methanolic extract in the HeLa cell line and normal lymphocytes. The methanolic extract inhibited the proliferation of HeLa cells (IC50 1.85 µg·mL-1), but the lymphocytes were affected by the extract (IC50 30.04 µg·mL-1). Some fractions, and the pool of all of them, showed inhibition higher than 80% at a concentration of 2.11 µg·mL-1. Therefore, the biological effect shown by the methanolic extract of the Perla Negra has some specificity in inhibiting tumor cells and not normal cells; an unusual feature among molecules investigated as potential biomedical agents.

Fruit extract from a Sechium edule hybrid induce apoptosis in leukaemic cell lines but not in normal cells.

The antiproliferative potential of a crude extract from the chayote hybrid H-837-07-GISeM® and its potential for apoptosis induction were assessed in leukaemic cell lines and normal mouse bone marrow mononuclear cells (BM-MNCs). The extract strongly inhibited the proliferation of the P388, J774, and WEHI-3 cell lines (with an IC50 below 1.3 µg·mL(-1)), reduced cell viability, and induced apoptotic body production, phosphatidylserine translocation, and DNA fragmentation. However, the extract had no effect on BM-MNCs. We postulate that these properties make the extract a good candidate for an anti-tumour agent for clinical use.

Inhibition of the growth and development of mosquito larvae of Culex quinquefasciatus (Diptera: Culicidae) treated with extract from leaves of Pseudocalymma alliaceum (Bignonaceae).

OBJECTIVE: To determine larvicidal activity of the essential oil, hydrolat and botanical extracts derived from leaves of Pseudocalymma alliaceum on mosquito larvae of Culex quinquefasciatus. METHODS: Groups of twenty larvae were used in the larvicidal assays. The mortality, relative growth rate, the larval and pupal duration and viability was estimated. The essential oil was analyzed by solid phase microextraction using gas chromatography coupled to mass spectrometry. RESULTS: Essential oil at 800 ppm showed larvicidal activity at 24 h with lethal values of LC50 and LC90 of 267.33 and 493.63 ppm. The hydrolat at 20% and 10% on 2nd stage larvae showed 100% effectiveness after 24 h. The aqueous extract at 10% had a relative growth index of 0.58, while the ethanolic and methanolic extract obtained values of 0.76 and 0.70 and control reached 0.99. Larvae treated with 10% of methanol, ethanol and aqueous extract showed a reduction in larval duration of 5.00, 2.20 and 4.35 days; ethanol extract at 1% provoke decrease of 2.40 days in the development and exhibited an increment of 3.30 days when treated with 0.01%. Aqueous, ethanol and methanol extracts at 10% reduced in 6.15, 3.42 and 5.57 days pupal development. The main compounds were diallyl disulfide (50.05%), diallyl sulfide (11.77%) and trisulfide di-2-propenyl (10.37%). CONCLUSIONS: The study demonstrated for the first time, the larvicidal activity of the essential oil and hydrolat of Pseudocalymma alliaceum; aqueous, ethanol and methanol extracts inhibited the normal growth and development of mosquito larvae, prolonging and delaying larval and pupal duration.

Amphypterygium adstringens anacardic acid mixture inhibits quorum sensing-controlled virulence factors of Chromobacterium violaceum and Pseudomonas aeruginosa.

BACKGROUND AND AIMS: Quorum sensing (QS) is a process of bacterial cell-cell communication that controls a large number of systems affecting pathogenicity. Interrupting this communication system can provide nonvirulent pathogenic bacteria. The aim of this study was to evaluate the anti-quorum sensing (anti-QS) potential of an anacardic acids mixture isolated from Amphipterygium adstringens, a medicinal plant known as "cuachalalate", to prevent the onset of bacterial infections as an alternate to antibiotics. METHODS: Initially we investigated the anti-QS activity of A. adstringens hexane extract (HE) by the inhibition of violacein production in Chromobacterium violaceum. From the active HE, an anacardic acid mixture (AAM) was obtained. The anti-quorum sensing activity of AAM was investigated by the rhamnolipid and pyocyanin production constraint as well as decrease of elastase activity, all being quorum sensing-controlled virulence factors expressed in the pathogenic bacteria Pseudomonas aeruginosa. RESULTS: HE induced a 91.6% of inhibition of the violecin production at 55 µg/mL concentration, whereas AAM showed 94% of inhibition at 166 µg/mL. In both cases, inhibition of violacein production did not affect the viability of the bacterium. AAM inhibited pyocyanin (86% at 200 µg/mL) and rhamnolipid (91% at 500 µg/mL) production in a dose/response form and decrease the elastase (75% at 500 µg/mL) activity in P. aeruginosa without affecting its development. CONCLUSIONS: Because an anacardic acids mixture isolated from A. adstringens demonstrated anti-QS, it could be further exploited for novel molecules to treat the emerging infections of antibiotic-resistant bacterial pathogens.

Flavonoids and antioxidant activity of flowers of Mexican Crataegus spp.

Flavonoids and antioxidant activity of extracts of flowers from some Mexican accessions of Crataegus were studied using six accessions with the purpose of contributing to the knowledge of the nutraceutical properties of the accessions of the Germplasm Bank of the Universidad Autonoma Chapingo. Flavonoids were identified by HPLC-MS. Among the flavonoids, the quercetin 3-O-glucoside (3), quercetin 3-O-rhamnoside (4), quercetin 3-O-rhamnosyl-(1 → 6)-glucoside (2) and quercetin 3-O-rhamnosyl-(1 → 2)-[rhamnosyl-(1 → 6)]-glucoside (1) were assigned. Flavonoid content and radical scavenging activity explain some of the medicinal properties attributed to flowers of Mexican hawthorns.

Anti-ulcer activity of Cyrtocarpa procera analogous to that of Amphipterygium adstringens, both assayed on the experimental gastric injury in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: The bark of Amphipterygium adstringens (Aa) is commonly mixed or adulterated with the bark of Cyrtocarpa procera (Cp) and sold in Mexican markets. Aa is a well known species in Mexico used as decoction to relieve ulcers. Scientific reports reinforcing the anti-ulcer activity of Aa have been previously described, but those describing the anti-ulcer properties of Cp as a substitute for Aa in folk medicine are scarce. AIM OF THE STUDY: To investigate anatomical and phytochemical differences between these species, as well as to assess the anti-ulcer effect of Cp extracts in comparison to the Aa extracts. MATERIAL AND METHODS: Anatomical micro-technique and physical and spectroscopic data were used to analyze differences between Cp and Aa. Regard to the pharmacological activity, it was assessed by using the ethanol-induced gastric damage model in rats. RESULTS: Whereas the bark anatomy of Aa was characterized by vertical canals in the periderm and the rare occurrence of fibers in its phloem, a periderm without vertical canals and abundant fibers in the phloem were distinctive features of Cp. Phytochemical analysis allowed the identification of tirucallane, masticadienonic and 3α-hydroxymasticadienonic acids as major components in Aa, while ß-amyrin and ß-sitosterol were obtained from Cp. Gastric lesions observed in the control group decreased in the presence of 100mg/kg of hexane, ethyl acetate and methanol extracts from the normal or regenerated bark of Cp, thus resembling the anti-ulcer effect of Aa. Nevertheless, major anti-ulcer potency was observed with the most active methanol extract from Cp obtained from normal [the effective dose fifty ED(50)=45.54 mg/kg] or regenerated (ED(50)=36.68 mg/kg) bark in comparison to Aa (ED(50)=115.64 mg/kg). CONCLUSION: Chemical and anatomical differences were found between these species, but since the anti-ulcer activity of Cp is similar to that shown by Aa our results reinforce the use of both species for the relief of gastric ulcer in folk medicine.

HPLC/MS analysis and anxiolytic-like effect of quercetin and kaempferol flavonoids from Tilia americana var. mexicana.

AIM OF THE STUDY: Around the world, Tilia species have been used in traditional medicine for their properties as tranquilizer. Furthermore, Mexican species of Tilia have been grouped as Tilia americana var. mexicana, but their specific content in flavonoids is poorly described. In this study, inflorescences of Mexican Tilia were collected in three different regions of Mexico to compare their flavonoid content and anxiolytic-like response. MATERIALS AND METHODS: Flavonoid content was analyzed by using an HPLC-MS technique. For anxiolytic-like response, Tilia inflorescences extracts (from 10 to 300 mg/kg, i.p.) were tested in experimental models (open-field, hole-board and plus-maze tests, as well as sodium pentobarbital-induced hypnosis) in mice. RESULTS: HPLC-MS analysis revealed specific peaks of flavonoid composition demonstrating some differences in these compounds in flowers and bracts depending on the region of collection. No differences in the neuropharmacological activity among these samples of Tilia were found. Moreover, their effects were associated with quercetin and kaempferol glycosides. CONCLUSIONS: Dissimilarities in the flavonoid composition of Mexican Tilias might imply that these species must be re-classified in more than one species, not as a unique Tilia americana var. mexicana. Since quercetin and kaempferol aglycons demonstrated anxiolytic-like response and that no difference in the pharmacological evaluation was observed between these three Mexican Tilias, we suggest that this pharmacological effect of Tilia inflorescences involves these flavonoids occurrence independently of the kind of glycosides present in the samples reinforcing their use in traditional medicine in several regions of Mexico.

Bactericidal and fungicidal activities of Calia secundiflora (Ort.) Yakovlev.

Calia secundiflora (Ortega) Yakovlev (Fabaceae) is considered a medicinal plant in Mexico but has scarcely been used because of the toxicity of its quinolizidine alkaloids. Several quinolizidine alkaloids have shown bactericidal, nematicidal, and fungicidal activities. The purpose of this study was to identify the alkaloids in the seeds and evaluate the activity of the organic extract on several phytopathogenic fungi and bacteria. An in vitro bioassay was conducted with species of the following phytopathogenic fungi: Alternaria solani, Fusarium oxysporum and Monilia fructicola; and of the following bacteria Pseudomonas sp., Xanthomonas campestris and Erwinia carotovora. Cytisine, lupinine, anagyrine, sparteine, N-methylcytisine, 5,6-dehydrolupanine, and lupanine were identified by liquid chromatography-mass spectrometry in the extract of seeds; the most abundant compound of the extract was cytisine. It was observed that the crude extract of Calia secundiflora was moderately active on bacteria and more potent on phytopathogenic fungi. In contrast cytisine showed the opposite effects.

Bioactivity-guided isolation of beta-sitosterol and some fatty acids as active compounds in the anxiolytic and sedative effects of Tilia americana var. mexicana.

Tilia species have been used as anxiolytics for many years. In a previous study anxiolytic-like effects of a hexane extract of Tilia americana var. mexicana inflorescences were observed in experimental models in mice. To get additional insights into the neuroactive actions of this particular Tilia species, in this study we report a bioactivity guided-fractionation of the extract and separation by column chromatographic methods to isolate three fatty acids and a triterpene identified as beta-sitosterol as major constituents. Our results revealed that the crude extract at 10 and 30 mg/kg I. P. and some pooled fractions at the same dosages potentiated sodium pentobarbital-induced sleeping time and caused a significant increase in the time spent at the open-arm sides in the plus-maze test. A reduction in the exploratory behavioral pattern manifested as ambulatory activity, as well as head dipping and rearing tests was also observed. Further fractionation and purification yielded four major fractions containing fatty acids and beta-sitosterol as the active compounds. A dose-response curve of beta-sitosterol in the range 1 to 30 mg/kg doses indicated that this compound produced an anxiolytic-like action from 1 to 10 mg/kg and a sedative response when the dose was increased to 30 mg/kg, these effects resemble those produced by diazepam (0.1 mg/kg). Our results suggest that hexane extract of Tilia americana var. mexicana produces depressant actions on the central nervous system, at least in part, because of the presence of beta-sitosterol and some fatty acids that remain to be identified.

Effect of 6-nonadecyl salicylic acid and its methyl ester on the induction of micronuclei in polychromatic erythrocytes in mouse peripheral blood.

The bark of Amphipterygium adstringens is widely used in the traditional Mexican medicine for treating ailments such as gastric ulcers, gastritis and stomach cancer. The 6-nonadecyl salicylic acid (anacardic acid) was isolated from the bark of this species. In previous papers have been informed that the anacardic acids possess anti-tumour, antimicrobial, antiacne, antibacterial and many others medicinal properties. Now we describe cytotoxic and genotoxic effects of this compound and its methyl ester. The cytotoxic and genotoxic effects of 6-nonadecyl salicylic acid (6NDSA) and its methyl ester (ME6NDSA) on CD1 male mice were determined with micronucleus assay at 24, 48 and 72h after oral administration of doses of 0.75, 2.5, 5.0 and 10.0mg/kg. Peripheral blood samples were drawn from the caudal vein and analyzed by Giemsa-stained technique. The results obtained showed that the ratios of polychromatic erythrocytes (PCE) to normochromatic erythrocytes (NCE) in mice treated with 10mg/kg of 6NDSA were statistically lower after 24h compared with its negative control animals, and that after 72h, PCE/NCE ratios were reduced in animals treated with 6NDSA at all tested dose levels. The methyl ester ME6NDSA showed no such cytotoxic activity. Neither of the test compounds increased the frequency of micronucleated polychromatic erythrocytes from which it appears that administration of 6NDSA and ME6NDSA may not lead to chromosome damage at the evaluated doses.