UHPLC-MS/MS and GC-MS Metabolic Profiling of a Medicinal Flourensia Fiebrigii Chemotype.

The comparative metabolic profiling and their biological properties of eight extracts obtained from diverse parts (leaves, flowers, roots) of the medicinal plant Flourensia fiebrigii S.F. Blake, a chemotype growing in highland areas (2750 m a.s.l.) of northwest Argentina, were investigated. The extracts were analysed by GC-MS and UHPLC-MS/MS. GC-MS analysis revealed the presence of encecalin (relative content: 24.86 %) in ethereal flower extract (EF) and this benzopyran (5.93 %) together sitosterol (11.35 %) in the bioactive ethereal leaf exudate (ELE). By UHPLC-MS/MS the main compounds identified in both samples were: limocitrin, (22.31 %), (2Z)-4,6-dihydroxy-2-[(4-hydroxy-3,5-dimethoxyphenyl)methylidene]-1-benzofuran-3-one (21.31 %), isobavachin (14.47 %), naringenin (13.50 %), and sternbin, (12.49 %). Phytocomplexes derived from aerial parts exhibited significant activity against biofilm production of Pseudomonas aeruginosa and Staphylococcus aureus, reaching inhibitions of 74.7-99.9 % with ELE (50 µg/mL). Notably, the extracts did not affect nutraceutical and environmental bacteria, suggesting a selective activity. ELE also showed the highest reactive species scavenging ability. This study provides valuable insights into the potential applications of this chemotype.

Lemon Oils Attenuate the Pathogenicity of Pseudomonas aeruginosa by Quorum Sensing Inhibition.

The chemical composition of three Citrus limon oils: lemon essential oil (LEO), lemon terpenes (LT) and lemon essence (LE), and their influence in the virulence factors production and motility (swarming and swimming) of two Pseudomonas aeruginosa strains (ATCC 27853 and a multidrug-resistant HT5) were investigated. The main compound, limonene, was also tested in biological assays. Eighty-four compounds, accounting for a relative peak area of 99.23%, 98.58% and 99.64%, were identified by GC/MS. Limonene (59-60%), γ-terpinene (10-11%) and ß-pinene (7-15%) were the main compounds. All lemon oils inhibited specific biofilm production and bacterial metabolic activities into biofilm in a dose-dependent manner (20-65%, in the range of 0.1-4 mg mL-1) of both strains. Besides, all samples inhibited about 50% of the elastase activity at 0.1 mg mL-1. Pyocyanin biosynthesis decreases until 64% (0.1-4 mg mL-1) for both strains. Swarming motility of P. aeruginosa ATCC 27853 was completely inhibited by 2 mg mL-1 of lemon oils. Furthermore, a decrease (29-55%, 0.1-4 mg mL-1) in the synthesis of Quorum sensing (QS) signals was observed. The oils showed higher biological activities than limonene. Hence, their ability to control the biofilm of P. aeruginosa and reduce the production of virulence factors regulated by QS makes lemon oils good candidates to be applied as preservatives in the food processing industry.

Argentinean Puna Plants with In Vitro Antioxidant and Anti-Inflammatory Activities as a Potential Nutraceutical.

Parastrephia lucida (Compositae), Tessaria absinthioides (Compositae), and Ephedra multiflora (Ephedraceae), three plant species from the Argentinean Puna (3600 m.a.s.l.) were selected for their anti-inflammatory and antioxidant properties to prepare mixtures to evaluate their use as nutraceuticals. Seven binary and ternary herbal mixtures made of ethanol 20% extracts of the selected plant species were prepared (Mixtures A to G), and they were analyzed for their effect on proinflammatory enzymes and their antioxidant activity in two cellular systems and in cell free systems. Toxicity tests were also carried out, and they were analyzed by high-performance liquid chromatography with a diode-array detector (HPLC-DAD) to quantify chemical markers. Mix A (equal parts of the three selected plant species) showed an important inhibitory capacity of different proinflammatory enzymes. Its potency on COX-2 was also higher than that of ibuprofen. Mix A and Mix G (P. lucida and T. absinthioides 1:1) showed a high antioxidant capacity in cellular and in cell-free systems. Toxicity assays further demonstrated their safety. This work shows the potential use of herbal mixtures made of medicinal plant species from the Argentinean Puna as nutraceutical or dietary supplements with antioxidant and anti-inflammatory activities. PRACTICAL APPLICATION: P. lucida, T. absinthioides, and E. multiflora are three plant species that are commonly used by Argentinean Puna inhabitants with medicinal purposes. Their proven safety, their antioxidant activity as well as their capacity to inhibit different proinflammatory enzymes make them attractive candidates to be used in combination as part of a dietary supplement aimed to prevent or palliate gastrointestinal and systemic inflammatory diseases. The use of native plant species as an alternative to more common and commercial plant species would have a positive impact on local communities' economies.

Effect of Wine Wastes Extracts on the Viability and Biofilm Formation of Pseudomonas aeruginosa and Staphylococcus aureus Strains.

In this work, we intended to inhibit the biofilm synthesis and the metabolism of Gram-positive and Gram-negative bacteria using two highly available wastes (stem and marc) obtained after the manufacturing of Torrontes wine at Cafayate, Argentina. Wine wastes contain a significant amount of bioactive compounds, mainly phenolic compounds, which makes them a potential source of compounds with beneficial properties to human health, as they could inhibit the virulence of pathogenic bacteria or protect the tissue against oxidative stress. Marc and stem extracts of Torrontes wine were evaluated for their ability to inhibit the metabolism and biofilm production of Pseudomonas aeruginosa and Staphylococcus aureus strains. The phytochemical composition and antioxidant activity of these extracts were also determined. The methanol and ethyl acetate extracts, which contained the highest amount of total polyphenolic, exhibited the highest scavenging capacity of ABTS and nitric oxide and the strongest Fe3+ reducing power and exhibited the highest level of inhibition of the biofilm formation and of the metabolic activity in bacterial biofilm. We also noticed a positive correlation between phenolic compounds content, the antioxidant activity, and the anti-biofilm capacity of the winemaking wastes. These results display the potentiality of wine wastes to prevent or reduce the formation of biofilm. Moreover, their abundance makes them an attractive and affordable source of antibiofilm and antioxidant agents.

Tetraglochin andina Ciald.: A medicinal plant from the Argentinean highlands with potential use in vaginal candidiasis.

ETHNOPHARMACOLOGICAL RELEVANCE: The Argentinean medicinal plant Tetraglochin andina Ciald, formerly classified as T. cristatum (Britton) Rothm is used in traditional medicine by inhabitants from Argentinean northwestern highlands (Puna) to treat candidiasis and as anti-inflammatory. AIM OF THE STUDY: To assess the potential of the crude drug as an anti-Candida agent with anti-inflammatory properties. The bioactivity and phytochemical composition of a dry extract of the plant was investigated. MATERIAL AND METHODS: The pharmacognostic description of the crude drug is carried out for the first time, including macroscopic and microscopic examinations of the different organs, physicochemical and extractive values (petroleum ether-, ethanol- and water-soluble). The dry extract from T. andina was evaluated as antifungal against pathogenic Candida sp. and Saccharomyces cerevisiae isolated from vaginal infections and reference strains, by the macrodilution and microdilution assays. The normal vaginal microbiome in women is characterized by the dominance of lactic acid-producing bacteria, mainly Lactobacillus spp. The effect of T. andina extract on Lactobacillus strains was also assayed. The inhibitory effect on proinflammatory enzymes (cyclooxygenase, lipoxygenase and phospholipase A2) and antioxidant capacity was studied. The chemical profile was analyzed by HPLC-ESI-MS. RESULTS: The hydroalcoholic extract inhibited the growth of all yeasts with Minimal Inhibitory Concentration (MIC) values between 12.5 and 400 µg GAE/mL and the MIC values on Lactobacillus were higher than the MIC values against Candida isolates ( > 400 µg GAE/mL). These results indicate that the hydroalcoholic extract could be used without affecting the normal microbiota of vaginal fluid. The extract showed antioxidant activity and could modulate the inflammatory process by three pathways (sPLA2, COX-2, LOX). The plant extract contained high total phenolic levels (386.9±1.7 mg GAE/g dry extract) and flavonoid levels (260.4±2.7 mg GAE/g dry extract). Fifty phenolic compounds were identified by HPLC-ESI-MS. They were mainly hydrolysable and condensed tannins. The dry extract was chemically and biologically stable during one year at room temperature or 4 °C. CONCLUSIONS: The presence of anti-Candida and anti-inflammatory activities in Tetraglochin andina extracts give support to their traditional use for treating conditions associated with microorganism infections and inflammatory process in humans. This plant preparation could be used to design phytopharmaceutical preparations to inhibit yeast growth and moderate the inflammatory and oxidative process.

The Native Fruit Geoffroea decorticans from Arid Northern Chile: Phenolic Composition, Antioxidant Activities and In Vitro Inhibition of Pro-Inflammatory and Metabolic Syndrome-Associated Enzymes.

The native tree Geoffroea decorticans (chañar) grows in the arid lands of northern Chile. It has been used as a food plant since prehistoric times. Phenolic-enriched extracts (PEEs) of Chilean chañar fruits were assessed for their chemical composition, antioxidant properties and inhibition of pro-inflammatory and metabolic syndrome-associated enzymes. Phenolic profiles were determined by HPLC-DAD-MS/MS. The PEEs of G. decorticans showed a strong effect towards the enzymes COX-1/COX-2, with inhibition percentages ranging from inactive to 92.1% and inactive to 76.0% at 50 µg PEE/mL, respectively. The IC50 values of the PEEs towards lipoxygenase and phospholipase A2 inhibitory activity were between 43.6-96.8 and 98.9-156.0 µg PEE/mL, respectively. Samples inhibited α-glucosidase (IC50 0.8-7.3 µg PEE/mL) and lipase (9.9 to >100 µg PEE/mL). However, samples did not inhibit α-amylase. The HPLC-DAD-MS analysis of the PEEs allowed the tentative identification of 53 compounds, mainly flavonol glycosides and procyanidins. The procyanidin content of the Chilean G. decorticans pulp was positively correlated with the antioxidant activity and the inhibition of the enzyme α-glucosidase. These results indicate that the Chilean chañar fruit contains bioactive polyphenols with functional properties.

Inhibition of pro-inflammatory enzymes by medicinal plants from the Argentinean highlands (Puna).

ETHNOPHARMACOLOGICAL RELEVANCE: Human groups in the Argentinean Andes highlands (Puna) selected native plants as anti-inflammatory agents. The indications of use are mainly to relieve pain, as infusions, ethanolic extracts or plasters. AIM OF THE STUDY: The objective of the study was to assess the effect of hydroalcoholic extracts from native highland plants as anti-inflammatory agents according to the traditional indications of use. The chemical profile of the three most active species was analyzed by HPLC-ESI-MS to get an insight into the constituents and the effects observed according to the ethnopharmacological information. MATERIALS AND METHODS: Hydroalcoholic extracts from 13 Argentinean Puna plants used as anti-inflammatory were evaluated as inhibitors of the pro-inflammatory enzymes phospholipase A2 (sPLA2), lipoxygenase (LOX), hyaluronidase, and for their capacity to stabilize red blood cells membrane. In addition, the extracts were evaluated to determine their reducing power, iron chelating capacity and ABTS•+ radical scavenging effect. The chemical profiles of the most active species were analyzed by HPLC-ESI-MS. RESULTS: Among the species investigated, Ephedra multiflora was the most active as LOX inhibitor (IC50:132µg/mL), by reducing the non-heme iron group and by scavenging radicals. The IC50 values of the reference compounds caffeic acid and naproxen were 57.0 and 14.0µg/mL, respectively. Parastrephia lucida showed the highest sPLA2 inhibitory effect (63% of inhibition at 200µg/mL). Under the same experimental conditions, the IC50 of the reference compound acetylsalicylic acid was 65±1µg/mL. Tessaria absinthioides exhibited the best inhibition towards hyaluronidase with an IC50 of 93.2±4.3µg/mL. Under the same experimental conditions, the reference compounds quercetin and indomethacin presented IC50 values of 340.0±17.0 and 502.0±10.0µg/mL, respectively. Among the most active species, 13 compounds were tentatively identified by HPLC-ESI-MS in E. multiflora and P. lucida, and 12 compounds in T. absinthioides. The constituents included caffeoyl- and feruloylquinic acid derivatives, flavonoids, simple phenolics and sesquiterpene glycosides. CONCLUSIONS: Six out of the 13 species investigated showed a moderate to strong effect towards the enzyme sPLA2 (>40% inhibition at 200µg/mL) while three species presented a strong activity against LOX with IC50<250µg/mL and three were very active against hyaluronidase. Most of the crude drug extracts were able to stabilize the red blood cells membrane, preventing their lysis. The compounds identified in the extracts explain, at least in part, the activity found in the samples. The effect observed for the most active species supports their traditional use as anti-inflammatory agents. However, more studies should be undertaken to disclose the potential of the Puna plants as anti-inflammatory crude drugs.

Chemical and functional characterization of seed, pulp and skin powder from chilto (Solanum betaceum), an Argentine native fruit. Phenolic fractions affect key enzymes involved in metabolic syndrome and oxidative stress.

The aim of this work was to assess the nutritional and functional components of powder obtained by lyophilization of whole fruits, seeds, pulp and skin from chilto (Solanum betaceum Cav) cultivated in the ecoregion of Yungas, Argentina. The powders have low carbohydrate and sodium content and are a source of vitamin C, carotenoid, phenolics, potassium and fiber. The HPLC-ESI-MS/MS analysis of the fractions enriched in phenolics allowed the identification of 12 caffeic acid derivatives and related phenolics, 10 rosmarinic acid derivatives and 7 flavonoids. The polyphenols enriched extracts before and after simulated gastroduodenal digestion inhibited enzymes associated with metabolic syndrome, including α-glucosidase, amylase and lipase and exhibited antioxidant activity by different mechanisms. None of the analyzed fruit powders showed acute toxicity or genotoxicity. The powders from the three parts of S. betaceum fruit may be a potential functional food and the polyphenol enriched extract of seed and skin may have nutraceutical properties.

Biological activities of polyphenols-enriched propolis from Argentina arid regions.

BACKGROUND: Propolis is a bioactive natural product collected by honeybees (Apis mellifera) from plant sources. PURPOSE: This study was undertaken to determine the effect of propolis extracts from arid region of Argentina, on the activity/expression of pro-inflammatory enzymes, and as potential free radical scavenger, antifungal and anthelmintic agent as well as to get a first insight into the polyphenolic profile of the active fractions. STUDY DESIGN/METHODS: Two propolis samples were collected in different time from hives located in Tucumán, Argentina. They are representative of the collection time of the raw material for phytotherapeutical purposes. Ethanolic extracts from both propolis were obtained. The PEEs were analyzed for total polyphenol (TP), non-flavonoid phenols (NFP) and flavonoid (FP) content followed by HPLC-DAD analysis and identification of components by HPLC-MS/MS(n). The potentiality as anti-inflammatory (LOX, COX, iNOS enzymes), antioxidant, antifungal and nematicidal was determined. RESULTS: PEEs contain high levels of TP, NFP and FP, including cinnamic acid, caffeic acid prenyl ester, caffeoyl dihydrocaffeate and caffeic acid 3,4-dihydroxyphenethyl ester, liquiritigenin, 2',4'-dihydroxychalcone and 2',4'-dihydroxy-3'-methoxychalcone. The PEEs in vitro reduced the activity of LOX and COX-2. Pretreatment of RAW 264.7 cells with PEEs before the induction of inflammatory state, inhibited NO overproduction and the iNOS protein expression was significantly decreased. The PEEs exhibited antioxidant, antifungal (Candida sp.) and nematicidal effect (C. elegans). CONCLUSION: These findings show the potential use of characterized PEEs from arid regions of Argentina as phytomedicine.

Zuccagnia punctata: A Review of its Traditional Uses; Phytochemistry, Pharmacology and Toxicology.

Zuccagnia punctata Cav. (Fabaceae, Caesalpiniaceae) is a plant with a long history of use in Argentine traditional medicines; it belongs to a monotypic genus, and is an endemic species of Argentina. This review provides a comprehensive overview of the traditional uses, phytochemistry, pharmacological activity and toxicology of Z. punctata. A wide range of traditional uses are cited in the literature such as antibacterial, antifungal, anti-inflammatory, and antitumor, among - others. Pharmacological studies to date have demonstrated significant activities that support the traditional uses of this plant. No human clinical trials had been - completed up to the time of this review and no toxic effect had been detected in animals. Compounds from different chemical groups have been isolated such las phenolic compounds and essential oils. Plant extracts and phytochemicals isolated exhibit a broad range of activities, anti-inflamnatory, antibacterial, antifungal, antigenotoxic, antioxidant, antiulcer, and nematicidal. The main bioactive phytochemicals in the aerial parts (leaf, stem and flower) were identified as 2', 4'-dihydroxy-3'-methoxychalcone and 2', 4'-dihydroxychalcone and were proposed as chemical markers. Consequently, standardized dry extracts of aerial parts of Z. puntacta could be used in herbal medicinal products. Also, they could be included in phytotherapeutic preparations such as capsules, creams, and gels, and for microencapsulation.

Anti-inflammatory, Antioxidant and Antimicrobial Activity Characterization and Toxicity Studies of Flowers of "Jarilla", a Medicinal Shrub from Argentina.

Zuccagnia punctata Cav. (Fabaceae) is an Argentine medicinal aromatic shrub (jarilla pispito, puspus, lata and jarilla macho). The chalcones were identified as pigments responsible for the yellow color of the flowers. Hydroethanolic extracts were obtained both from fresh flowers and from flowers dried by lyophilization. The extracts were standardized by their phenolic and flavonoids content. Their fingerprints by HPLC-DAD indicated the presence of two chalcones as major compounds (2',4'-dihydroxychalcone and 2',4'-dihydroxy-3'-methoxychalcone). Both extracts showed the same total phenolic, non-flavonoid phenolic and flavonoid phenolic content and their phenolic profiles were similar. The polyphenolic extracts exhibited antioxidant (free radical scavenging and inhibitory activity on lipoperoxidation) and anti-inflammatory (inhibition of lipoxygenase and cyclooxygenase enzymes) activities. The flower extracts were active against six Candida species with MIC values between 60 and 120 µg GAE x mL(-1) and were also active on methicillin-resistant Staphylococcus aureus (MIC: 250 µg GAE x mL(-1)) and Enterococcus faecalis (MIC: 500 µg GAE x mL(-1)). The extracts were neither toxic (Artemia salina test) nor mutagenic (Ames test). Jarilla flowers could be considered as a new dietary supplement that could help to prevent pathologies associated with oxidative stress and the polyphenolic extract obtained from them could be considered as a standardized phytotherapeutic product with antimicrobial, antioxidant and anti-inflammatory activities. The aim of this work was to determine the pigments responsible for the yellow color of the flowers of Z. punctata and to evaluate the functional properties of the polyphenolic extract of the flowers. The toxicity (Artemia salina) and mutagenic activity (Ames test) of the extract were also evaluated.

Anti-inflammatory activity of copao (Eulychnia acida Phil., Cactaceae) fruits.

Copao (Eulychnia acida Phil., Cactaceae) is an endemic species occurring in northern Chile. The edible fruits of this plant are valued for its acidic and refreshing taste. Phenolic-enriched extracts from copao fruit pulp and epicarp, collected in the Elqui and Limari river valleys, were assessed by its in vitro ability to inhibit the pro-inflammatory enzymes lipoxygenase (LOX) and cyclooxygenases (COX-1 and COX-2). At 100 µg/mL, pulp extracts showed better effect towards LOX than epicarp extract, while COX-2 inhibition was observed for both epicarp and pulp samples. In general, the extracts were inactive towards COX-1. A positive correlation was observed between the anti-inflammatory activity and the main phenolic compounds found in this fruit. Copao fruits from the Limari valley, a main place of collection and commercialization, showed major activity, adding evidence on the possible health-beneficial effects of this native Chilean fruit.

Effect of seasonal variations and collection form on antioxidant activity of propolis from San Juan, Argentina.

Propolis was included in the Argentine Food Code as a functional food. The chemical parameters and antioxidant properties of propolis samples from the same colonies of Apis mellifera in San Juan (Cuyo region, Western Argentine) were compared every month for 1 year using two collection methods. Chemical parameters were analyzed by the spectrophotometric method and fingerprinting using high-performance liquid chromatography with ultraviolet detection. The antioxidant activities of propolis samples were measured using model systems including the analysis of the scavenging activities for 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals and the beta-carotene bleaching assay. The results showed that propolis had a higher free radical scavenging and lipid peroxidation inhibitory capacity than butylated hydroxytoluene and quercetin, antioxidants used in the pharmaceutical and food industries. The concentration required to scavenge 50% of free radicals (SC(50)) values differed depending on the sample collection month. Samples collected in November had the highest antioxidant capacity. In all cases, SC(50) values of propolis samples obtained by scraping were similar to those collected from a wire mesh (5 microg/mL for ABTS and 20-30 microg/mL for DPPH radicals). A significant positive correlation was found between the antioxidant capacity and flavonoid content of each analyzed extract. The chemical profiles were very similar. Galangin (3,5,7-trihydroxyflavone), an antioxidant compound, was detected in all samples as a major compound. The chromatographic profile suggests that of Baccharis sp., which would be one of the botanical sources of propolis from western Argentina, and the content of galangin can be used as a parameter for evaluating propolis quality. Our results suggest that Argentine propolis from Cuyo is a promising source of bioactive compounds as ingredients for developing functional foods with a beneficial impact on oxidative stress.