Maytenus Ilicifolia Research Articles

Actividad antibacteriana in vitro de Maytenus ilicifolia Martius sobre Streptococcus mutans y Staphylococcus aureus

Maytenus ilicifolia es un arbusto empleado como medicación natural en Paraguay, Brasil, Argentina y Uruguay. Entre sus propiedades se cita la actividad antimicrobiana. El objetivo del estudio experimental in vitro fue evaluar el efecto antibacteriano del extracto etanólico de Maytenus ilicifolia Martius frente a Streptococcus mutans y Staphylococcus aureus.Se determinó el efecto de tres concentracionesC1 47,76 mg/mL, C2 24,4 mg/mL y C3 12,8 mg/mL, frente a cepas bacterianas de Streptococcus mutans ATCC 25175 y de Staphylococcus aureus ATCC 6538, mediante el método de difusión en pozo en agar Mueller Hinton.Se realizó un ensayo por triplicado para cada cepa con las diferentes concentraciones del extracto, incluyendo un control positivo de Gluconato de Clorhexidina 0,12% y un control negativo de solución fisiológica 0,9%.La C1 47,76 mg/mL tuvo efecto antibacteriano sobre ambas cepas. La media de halo de inhibición para Streptococcus mutans fue de 11,68 mmcorrespondiendo al 67,2% de efecto inhibitorio en relación con el control positivo. La media de halo de inhibición para Staphylococcus aureus fue de 9,39 mm, correspondiendo al 51,82% de efecto inhibitorio en relación con el Gluconato de Clorhexidina 0,12%. Se concluyó que el extracto etanólico de Maytenus ilicifolia Martius posee efecto antibacteriano para Streptococcus mutans y Staphylococcus aureus.

Evaluation of the Effects of Maytenus ilicifolia on the Activities of Cytochrome P450 3A and P-glycoprotein

Maytenus ilicifolia is a Brazilian popular medicine commonly used to treat ulcer and gastritis. Despite the absence of toxicity regarding its consumption, possible interactions when co-administrated with conventional drugs, are unknown. This study aimed to evaluate the effects of M. ilicifolia extracts on Cytochrome P450 3A (CYP3A) and P-glycoprotein (P-gp) activities. The extracts were obtained by infusion (MI) or turbo-extraction using hydro-acetonic solvent (MT70). The content of polyphenols in each extract was determined. To assess the modulation of M. ilicifolia on P-gp activity, the uptake of fexofenadine (FEX) by Caco-2 cells was investigated in the absence or presence of MI or MT70. The effect on CYP3A activity was evaluated by the co-administration of midazolam (MDZ) with each extract in male Wistar rats. The pharmacokinetic parameters of the drug were determined and compared with those from the control group. The content of total phenolic compounds, tannins, and flavonoids on MT70 extract was about double of that found in MI. In the presence of the extracts, the uptake of the P-gp marker (FEX) by Caco-2 cells increased from 1.7 ± 0.4 ng.mg-1 protein (control) to 3.5 ± 0.2 ng.mg-1 protein (MI) and 4.4 ± 0.5 ng.mg-1 protein (MT70), respectively. When orally co-administrated with MDZ (substrate of CYP3A), the extracts augmented the AUC(0-∞) (Control: 911.7 ± 215.7 ng.h.mL-1; MI: 1947 ± 554.3 ng.h.mL-1; MT70: 2219.0 ± 506.3 ng.h.mL-1) and the Cmax (Control: 407.7 ± 90.4 ng.mL-1; MI: 1770.5 ± 764.5 ng.mL-1; MT70: 1987.2 ± 544.9 ng.mL-1) of the drug in rats indicating a 50% reduction of the oral Cl. No effect was observed when midazolam was given intravenously. The results suggest that M. ilicifolia can inhibit the intestinal metabolism and transport of drugs mediated by CYP3A and P-gp, respectively, however, the involvement of other transporters and the clinical relevance of such interaction still need to be clarified.

Medicinal plants used in street markets in different regions of Brazil

In Brazil, markets and street markets emerged in 1841 as a solution for the regional supply of products. Street markets and municipal markets constitute a privileged space for traditional expressions and cultures about their ethnobotanical heritage since these places are centers of large numbers of information, underlying an environment of intense cultural exchanges. Bibliographic surveys were carried out on medicinal plants used in Street markets in different regions of Brazil. Where the objectives were to identify the families and species of medicinal plants used in different regions of Brazil, the common names and frequency of citations were identified regarding the mode of use of medicinal plants in the Street markets and it was verified which parts of the most cited plants in Street markets in different regions of Brazil were verified, the most cited therapeutic indications were verified. The most cited medicinal plants were “stonebreaker” plants Phyllanthus niruri, “elderberries” Sambucus nigra, “macela” Achyrocline satureioides and “espinheira santa” Maytenus ilicifolia. The modes of use of these plants were more cited as medicinal uses 71% and ritualistic baths 27%. The most used parts were leaves 60.1%, flower 13.5%, and bark 9.0%, the families that had a higher citation were Asteraceae, Lamiaceae, and Malvaceae. Regarding the therapeutic indications, stomach diseases, flu-like symptoms, skin problems, and healing, inflammation, and diuretics were more cited. It is concluded then that the use of medicinal plants is used for various therapeutic indications.

Validated spectrophotometric method for quantification of total triterpenes in plant matrices.

Triterpenes are ubiquitous secondary metabolites present in plants. They can be found in both forms, as genins or conjugated as glycosides. Although distinct analytical methods to quantify these compounds in vegetal tissues are available in the literature, limitations like high cost, complexity on sample preparation, and selectivity are often challenging issues. This study aimed to develop and to validate a simple and rapid spectrophotometric method to detect and quantify total triterpenes in plant matrices. The assay was conducted directly into glass tubes using vanillin, acetic acid, and sulphuric acid as reagents, and β-sitosterol as reference standard. The samples were analyzed at 548nm assessing the quality parameters of selectivity, linearity, precision, accuracy, limit of detection (LOD), limit of quantification (LOQ), and robustness. The method was selective, with precision and accuracy varying from 0.56% to 4.98% and 96.63% to 113.87%, respectively. The values of the limit of detection and quantification were 0.042μg.mL-1 and 0.14μg.mL-1, correspondingly. The correlation coefficient (r) at the concentration range of 3.08μg.mL-1to 24.61μg.mL-1 was 0.9998. The total of triterpenes found in of B. holophylla and M. ilicifolia leaves were 132.36 ± 20.36mg EβS.g-1 of dry extract and 53.91 ± 2.6mg EβS.g-1 of dry extract, respectively. The method was reliable to quantify total triterpenes extracted from Maytenus ilicifolia and Bauhinia holophylla. Graphical abstract.

First Report of Diaporthe oxe on Peroba Rosa in Brazil

“Peroba rosa” (Aspidosperma polyneuron Muell. Arg.) is a native tree species to the forests of Brazil, Venezuela, Argentina, Colombia, Peru, and Paraguay. The species has a high commercial value and, due to logging and destruction of its original ecosystems, is at extinction risk. Currently, the plant is listed for ex-situ and in-situ conservation in Brazil and Venezuela. In 2014, samples of peroba rosa seeds were collected from native trees in Parana State, southern Brazil, and analyzed to evaluate their sanitary quality (Mazarotto et al. 2019). Following germination under greenhouse conditions with average temperature of 24°C, Diaporthe sp. (TR-57) was transmitted from seeds to seedlings, causing necrotic spots on cotyledons, malformation of plants, and eventually their death. In PDA medium, this isolate showed sparse mycelium, white to olive, ochre reverse, presence of semi-immersed dark pycnidia, and alpha (7 × 3 µm) and beta conidia (28 × 2 µm), resembling Diaporthe oxe Gomes, Glienke & Crous (Gomes et al. 2013). The presence of perithecia was not observed. For species identification, the ITS and TEF-1α regions were partially sequenced using the primer pairs ITS4/ITS5 and EF1-728F/EF1-986R, respectively. The obtained sequences were compared phylogenetically with 90 congeneric species. The ITS (GenBank accession no. KX385041) and TEF-1α (KX385079) sequences had respectively 98 and 100% similarity with D. oxe (KC343166, KC343893) (Gomes et al. 2013). This isolate was deposited in the Collection of Forest Fungi and Oomycetes, Embrapa Florestas, Colombo, Parana State, Brazil. For the pathogenicity test in seedlings, disks (5 mm diameter) of active mycelium in PDA were removed and deposited on the adaxial side of the leaves with previous circular needle injuries (4 mm²). This procedure was repeated for 10 young leaves in three seedlings (1 year old), with one disk per leaf. The control consisted of a PDA disk deposited on the injured leaf. The seedlings were incubated in a humid chamber for 72 h and then kept in a greenhouse for 30 days. A darkening of the leaf blade was detected reaching an area of approximately 8 mm² and a white mycelial growth of the pathogen over the lesions in 20% of the leaves. The control group had no symptoms. The symptomatic leaves were plated in PDA for 7 days at 24°C. D. oxe was reisolated from 100% of the inoculated leaves, and identification was confirmed by molecular sequencing, fulfilling Koch’s postulates. In Brazil, D. oxe was isolated as an endophyte from Maytenus ilicifolia (Gomes et al. 2013) and, in Uruguay, was pathogenic to Pyrus communis (Sessa et al. 2017). This study represents the first report of D. oxe associated with peroba rosa seeds as pathogens, not only in Brazil but also worldwide.

Anticancer Activities of the Quinone-Methide Triterpenes Maytenin and 22-β-hydroxymaytenin Obtained from Cultivated Maytenus ilicifolia Roots Associated with Down-Regulation of miRNA-27a and miR-20a/miR-17-5p.

Natural triterpenes exhibit a wide range of biological activities. Since this group of secondary metabolites is structurally diverse, effects may vary due to distinct biochemical interactions within biological systems. In this work, we investigated the anticancer-related activities of the quinone-methide triterpene maytenin and its derivative compound 22-β-hydroxymaytenin, obtained from Maytenus ilicifolia roots cultivated in vitro. Their antiproliferative and pro-apoptotic activities were evaluated in monolayer and three-dimensional cultures of immortalized cell lines. Additionally, we investigated the toxicity of maytenin in SCID mice harboring tumors derived from a squamous cell carcinoma cell line. Both isolated molecules presented pronounced pro-apoptotic activities in four cell lines derived from head and neck squamous cell carcinomas, including a metastasis-derived cell line. The molecules also induced reactive oxygen species (ROS) and down-regulated microRNA-27a and microRNA-20a/miR-17-5p, corroborating with the literature data for triterpenoids. Intraperitoneal administration of maytenin to tumor-bearing mice did not lead to pronounced histopathological changes in kidney tissue, suggesting low nephrotoxicity. The wide-ranging activity of maytenin and 22-β-hydroxymaytenin in head and neck cancer cells indicates that these molecules should be further explored in plant biochemistry and biotechnology for therapeutic applications.

Impact of the Training of Professionals from São Paulo Public Health System in Phytotherapy Practice

Abstract: Introduction: Following the introduction in 2006 of the National Policy on Integrative and Complementary Practices, and The National Policy on Medicinal Plants and Herbal Medicines, the Municipal Secretariat for the Environment of the City Hall of Sao Paulo initiated the Medicinal Plants Course, later expanded to The Medicinal Plants and Herbal Medicines Course for the training of health professionals. This study aimed to evaluate the impact of the 2014 and 2015 edition of the course “Medicinal Plants and Herbal Medicines” on health professional practices. Methods: An exploratory and descriptive study was conducted with a qualitative-quantitative approach. The quantitative process (Phase I) consisted of a questionnaire being emailed to health professionals who were involved in the course, divided into seven categories: biomedical professional, dental surgeon, nurse, pharmacist, physiotherapist, physician and nutritionist; and, out of 165 questionnaires, 114 responses were received (69.1%). The Qualitative and Quantitative process (Phase II), comprised semi-structured, in-person individual interviews to obtain detailed information on Phytotherapy practices, with 73 health professionals being interviewed, before and after the training, comparing their practices afterwards. Results: The course had a positive impact on the acceptance and application of Phytotherapy by health professionals with a significant increase (p<0.001) in the expansion of activities related to Phytotherapy (herbal tea “meetings”, medicinal herb gardens and capacity training), regarding the use of herbal products, such as Matricaria chamomilla (chamomile), Maytenus ilicifolia (espinheira-santa), Valeriana officinalis (valeriana). An increase in the knowledge of Phytotherapy risks was also observed, although there was no increase in the reporting of adverse reactions. The study confirmed the importance of the inclusion of Medicinal Plants and Herbal Medicines Courses as part of undergraduate and postgraduate school, as well as the technical training and continuing education for SUS health professionals. Conclusion: The positive impact found out in almost all evaluated aspects, such as the increase in the knowledge and in the prescription of herbal medicines, confirms the importance of such courses. The results suggest the training promoted a positive impact on the Phytotherapy practice of the Public Health System professionals in São Paulo.

Impact of the Training of Professionals from São Paulo Public Health System in Phytotherapy Practice

Abstract: Introduction: Following the introduction in 2006 of the National Policy on Integrative and Complementary Practices, and The National Policy on Medicinal Plants and Herbal Medicines, the Municipal Secretariat for the Environment of the City Hall of Sao Paulo initiated the Medicinal Plants Course, later expanded to The Medicinal Plants and Herbal Medicines Course for the training of health professionals. This study aimed to evaluate the impact of the 2014 and 2015 edition of the course “Medicinal Plants and Herbal Medicines” on health professional practices. Methods: An exploratory and descriptive study was conducted with a qualitative-quantitative approach. The quantitative process (Phase I) consisted of a questionnaire being emailed to health professionals who were involved in the course, divided into seven categories: biomedical professional, dental surgeon, nurse, pharmacist, physiotherapist, physician and nutritionist; and, out of 165 questionnaires, 114 responses were received (69.1%). The Qualitative and Quantitative process (Phase II), comprised semi-structured, in-person individual interviews to obtain detailed information on Phytotherapy practices, with 73 health professionals being interviewed, before and after the training, comparing their practices afterwards. Results: The course had a positive impact on the acceptance and application of Phytotherapy by health professionals with a significant increase (p<0.001) in the expansion of activities related to Phytotherapy (herbal tea “meetings”, medicinal herb gardens and capacity training), regarding the use of herbal products, such as Matricaria chamomilla (chamomile), Maytenus ilicifolia (espinheira-santa), Valeriana officinalis (valeriana). An increase in the knowledge of Phytotherapy risks was also observed, although there was no increase in the reporting of adverse reactions. The study confirmed the importance of the inclusion of Medicinal Plants and Herbal Medicines Courses as part of undergraduate and postgraduate school, as well as the technical training and continuing education for SUS health professionals. Conclusion: The positive impact found out in almost all evaluated aspects, such as the increase in the knowledge and in the prescription of herbal medicines, confirms the importance of such courses. The results suggest the training promoted a positive impact on the Phytotherapy practice of the Public Health System professionals in São Paulo.

Development of effervescent medical powder containing Maytenus ilicifolia extract Mart. ex Reissek for treatment of gastric disorders

Brazil is the country with the biggest biodiversity on the planet, but it does not avail itself enough of this potential to produce herbal medicines. Maytenus ilicifolia Mart. Ex Reissek is popularly known as Espinheira-Santa and in Brazil is widely used in popular medicine to treat a number of diseases such as gastritis and dyspepsia, and also displays laxative, diuretic, analgesic, antiseptic and healing properties and antineoplastic and antimicrobial activities. Among the reasons that hinder a better exploration of such biodiversity, the absence of an innovative culture draws a lot of attention, besides the inherent difficulties in the characteristics of the research and the development of this product modality. The aim of this study was developing an effervescent powder containing medicinal extract of M. ilicifolia. The methodologies to determine quality of the plant and extract including the organoleptic characteristics, the moisture content and volatiles, total ashes, extractive content, pH, and the bulk density. In our experiments, we demonstrated that the plant has been identified and has displayed the appropriate organoleptic characteristics to the preparation of the extract; we also demonstrated that the extract has been incorporated into a formulation that maintained its organoleptic characteristics, pH and content, as well as the formation of stable gases. With these experiments, we demonstrate for the first time that the developed formulation can serve as a prototype for the development of pharmaceutics effervescent product containing standardized extract of M. ilicifolia.

A New Species of Macrolechriops Champion (Coleoptera: Curculionidae: Conoderinae: Lechriopini) Reared from the Fruits of Maytenus ilicifolia Mart. ex Reissek (Celastraceae)

Macrolechriops moreyi Anzaldo and O'Brien, new species, from Brazil and Uruguay is described and figured. Most examined specimens were reared from the fruits of Maytenus ilicifolia Mart. ex Reissek (Celastraceae), a widely grown shrub with many medicinal uses. The genus MacrolechriopsChampion, 1906 is discussed, and images of type specimens are provided for the other six species currently placed in Macrolechriops to aid future studies on the Lechriopini.

Effect of cover on the development and production of secondary compounds of <i>Maytenus ilicifolia</i> and <i>Ilex paraguariensis</i> in agroforestry systems

The Midwest regions of Santa Catarina state, in Brazil, include a great diversity of forest essences with medicinal application; however, many of these resources are poorly exploited or managed in an extractive manner. This research aimed to assess the effects of canopy cover in agroforestry systems on development and production of secondary composites of espinheira-santa (Maytenus ilicifolia) and yerba mate (Ilex paraguariensis). Two experimental areas were planted following the same spacing and arrangement for the plants. Experimental blocks were set up by pruning the pioneer plants, with three variable levels of canopy cover (0-20 %, 20-40 % and 40-60 %). Plants of espinheira-santa and yerba mate were monitored for development and their marketable vegetative parts were harvested, processed and aqueous extracts were prepared for the analysis of presence of secondary compounds. The yerba mate showed the highest production of marketable biomass under high cover. By contrast, the biosynthesis of the secondary compound, theobromine, was greater in the lowest cover, while chlorogenic acid and caffeine were higher in the cover (40-60 %). Espinheira-santa showed no difference in either development or yield among the assessment treatments and, the production of phenols was lower in the cover above 40 %.

Effects of the extraction method and chromatographic separation solvent in the antioxidant activity of different species of "espinheira-santa".

Bioactivity of "espinheira-santa" leaves can be evaluated through solvent purification processes. Species of the same genus may contain different compounds, which may be separated by conventional chromatography using different solvents. The extraction process also influences the contents of these compounds. In order to optimize the above process research is required on variables such as solvents, temperature and extraction time. The objective of this work was to evaluate the antioxidant activity (ABTS, DPPH and FRAP) of extracts and purified fractions from leaves of Maytenus ilicifolia and M. aquafolium. Percolation at 28°C for 7days, and ultrasound at 50, 60 and 70°C for 15min were the extraction methods tested. Open column chromatography (silica gel) was used in the fractionation of extracts, and hexane, chloroform and methanol were the elution solvents used. The evaluation methods of the antioxidant activity were DPPH, FRAP and ABTS. Plant species showed no significant differences between methods for DPPH, but the fractions obtained with chromatography had significant differences, the crude ethanolic extract (CEE) and the methanolic been in general superior. "espinheira-santa" is a plant with antioxidant potential, but the extraction methods are not optimal. Still, it can be said that the separation by chromatography provided a better visualization of the nature of the components of that plant that best stood out in relation to the antioxidant activity.

CYP712K4 Catalyzes the C-29 Oxidation of Friedelin in the Maytenus ilicifolia Quinone Methide Triterpenoid Biosynthesis Pathway.

The native Brazilian plant Maytenus ilicifolia accumulates a set of quinone methide triterpenoids with important pharmacological properties, of which maytenin, pristimerin and celastrol accumulate exclusively in the root bark of this medicinal plant. The first committed step in the quinone methide triterpenoid biosynthesis is the cyclization of 2,3-oxidosqualene to friedelin, catalyzed by the oxidosqualene cyclase friedelin synthase (FRS). In this study, we produced heterologous friedelin by the expression of M. ilicifolia FRS in Nicotiana benthamiana leaves and in a Saccharomyces cerevisiae strain engineered using CRISPR/Cas9. Furthermore, friedelin-producing N. benthamiana leaves and S. cerevisiae cells were used for the characterization of CYP712K4, a cytochrome P450 from M. ilicifolia that catalyzes the oxidation of friedelin at the C-29 position, leading to maytenoic acid, an intermediate of the quinone methide triterpenoid biosynthesis pathway. Maytenoic acid produced in N. benthamiana leaves was purified and its structure was confirmed using high-resolution mass spectrometry and nuclear magnetic resonance analysis. The three-step oxidation of friedelin to maytenoic acid by CYP712K4 can be considered as the second step of the quinone methide triterpenoid biosynthesis pathway, and may form the basis for further discovery of the pathway and heterologous production of friedelanes and ultimately quinone methide triterpenoids.

Neopestalotiopsis species presenting wide dye destaining activity: report of a mycelium-associated laccase

Wastewaters from textile dyeing industries represent an ecological concern, notably due to the known toxicity of azo dyes to the local microbiome and human health. Although physicochemical approaches are the rule for the treatment of industrial effluents, biological strategies such as enzyme-mediated dye destaining is a promising alternative. Notwithstanding a broad range of microorganisms, including fungi, algae, yeast, and bacteria, display dye-destaining properties, most of the literature has focused in ligninolytic fungi, leaving other classes of organisms somehow ignored. In this study, six endophytic strains isolated from Maytenus ilicifolia were studied for their destaining activity. The phylogenetic and morphological analysis allowed the identification of strain LGMF1504 as Neopestalotiopsis sp. LGMF1504 that decolorized several commercial dyes as the result of a mycelium-associated laccase. The enzyme expression was modulated by carbon and nitrogen content in the culture medium, it was weakly affected by the presence of aromatic compounds and metal ions while some common laccase mediators improved the destaining activity onto dye substrates. The best culture condition observed for laccase activity was a basic culture medium containing 5 g L−1 starch and 15 g L−1 ammonium tartrate. The laccase activity showed low substrate specificity and almost unaltered performance in a wide range of pH values and NaCl concentrations, suggesting the potential of Neopestalotiopsis sp. LGMF1504 for biodegradation approaches.

A newly validated HPLC–DAD-UV method to study the effects of medicinal plants extracts, fractions and isolate compounds on gastric emptying in rodents

In this study, we developed and validated a chromatographic method by High-performance Liquid Chromatography coupled to Diode-Array Ultraviolet Detector for quantification of acetaminophen in small volumes of plasma. This analytical method is particularly suitable for studies of gastric emptying time and pharmacokinetics in small rodents. Orally administered acetaminophen is promptly and completely absorbed in the intestines, with negligible absorption from the stomach. Owing to these kinetic features, acetaminophen can be used to study gastric emptying. The newly-validated analytical method was employed to investigate whether plants used as anti-dyspeptics in the Brazilian traditional medicine affect the gastric emptying. Lyophilized aqueous extracts from leaves of Baccharis trimera (Less) DC., Asteraceae (carqueja), Maytenus ilicifolia Mart. Ex Reissek, Celastraceae (espinheira-santa) and Lippia alba (Mill.) N.E.Br. ex Britton & P. Wilson, Verbenaceae (erva-cidreira) were administered by gavage to female Wistar rats 30min before a single oral dose of acetaminophen (50mg/kg). L. alba extract (50mg/kg) did not alter acetaminophen plasma concentration versus time curve an indication that it has no effect on gastric emptying. Extracts of B. trimera (50mg/kg) and M. ilicifolia (30 and 50mg/kg, but not 10mg/kg), however, slightly delayed gastric emptying. M. ilicifolia given by intraperitoneal route (30mg/kg) also retarded gastric emptying. In conclusion, the newly-validated analytical method is adequate for studying the effects of medicinal plants on gastric emptying.

Ecological Insights to Track Cytotoxic Compounds among Maytenus ilicifolia Living Individuals and Clones of an Ex Situ Collection.

Biodiversity is key for maintenance of life and source of richness. Nevertheless, concepts such as phenotype expression are also pivotal to understand how chemical diversity varies in a living organism. Sesquiterpene pyridine alkaloids (SPAs) and quinonemethide triterpenes (QMTs) accumulate in root bark of Celastraceae plants. However, despite their known bioactive traits, there is still a lack of evidence regarding their ecological functions. Our present contribution combines analytical tools to study clones and individuals of Maytenus ilicifolia (Celastraceae) kept alive in an ex situ collection and determine whether or not these two major biosynthetic pathways could be switched on simultaneously. The relative concentration of the QMTs maytenin (1) and pristimerin (2), and the SPA aquifoliunin E1 (3) were tracked in raw extracts by HPLC-DAD and 1H-NMR. Hierarchical Clustering Analysis (HCA) was used to group individuals according their ability to accumulate these metabolites. Semi-quantitative analysis showed an extensive occurrence of QMT in most individuals, whereas SPA was only detected in minor abundance in five samples. Contrary to QMTs, SPAs did not accumulate extensively, contradicting the hypothesis of two different biosynthetic pathways operating simultaneously. Moreover, the production of QMT varied significantly among samples of the same ex situ collection, suggesting that the terpene contents in root bark extracts were not dependent on abiotic effects. HCA results showed that QMT occurrence was high regardless of the plant age. This data disproves the hypothesis that QMT biosynthesis was age-dependent. Furthermore, clustering analysis did not group clones nor same-age samples together, which might reinforce the hypothesis over gene regulation of the biosynthesis pathways. Indeed, plants from the ex situ collection produced bioactive compounds in a singular manner, which postulates that rhizosphere environment could offer ecological triggers for phenotypical plasticity.

Effect of Trichoderma spp. on the Propagation of Maytenus ilicifolia Mart. ex Reissek

The interest in using biotechnology tools that contribute to reducing the need for chemical inputs in agroforestry production has increased in recent years, aiming at higher quality for the environment and for society. This interest is also applied to medicine species consumed in natura. The objective of this study was to evaluate the effect of Trichoderma spp. on germination and initial growth of Maytenus ilicifolia seedlings. Laboratory and greenhouse tests were carried out using Trichoderma spp. isolates obtained from three different strains identified as Trichoderma asperelloides (T1 & T2 strains) and Trichoderma virens (T10 strain). In laboratory, M. ilicifolia seeds without aryl were inoculated in solutions containing fungal spores and distributed in substrate paper in four replicates of 25 seeds/treatment. The first germination count, accumulated germination, percentages of accumulated dead seeds and firm seeds were evaluated at 7, 14, 21 and 28 days of incubation. The same isolates were used to evaluate the effect on the initial growth of seedling in a greenhouse through a completely randomized design with 40 replicates, considering a seedling as each replicate. Trichoderma spp. isolates were inoculated on the substrate used as the basis for seeding. Seeds of M. ilicifolia were used from the same batch of the laboratory test, but without removing the aryl. The variables of total height, diameter at root collar and number of leaves were evaluated at 90, 120, 150 and 180 days after seeding. A positive effect of Trichoderma inoculation on seed germination and vigor was observed in the laboratory, with emphasis on the T2 isolate. Growth promoting effects on the M. ilicifolia seedlings were not observed in greenhouse. We suggest to better investigate the interaction between the tested Trichoderma isolates and M. ilicifolia seeds in the presence of aryl.

Functional characterization of an oxidosqualene cyclase (PdFRS) encoding a monofunctional friedelin synthase in Populus davidiana.

An oxidosqualene cyclase (PdFRS) from Populus davidiana was characterized as a monofunctional friedelin synthase by its heterologous expression in yeast and overexpression in plants. Triterpenesare one of the largest classes of plant chemical compounds composed of threeterpeneunits, which form the basic skeleton of all sterols and saponins. Friedelin (friedelan-3-one), a pentacyclic triterpene, occurs in many plant families and is particularly present in rich amounts in cork tissues from trees. The biosynthesis of friedelin occurs through the oxidosqualene cyclase (OSC) enzyme that generates friedelin from 2,3-oxidosqualene after the maximum rearrangement of a triterpene skeleton. Populus davidiana is called Korean aspen and grows in northern East Asia. From 57,322 unique sequences generated from the P. davidiana transcriptome database, one complete coding sequence (PdFRS) was obtained from a contig, which showed 74% identity to Betula platyphylla β-amyrin synthase and 73% identity with friedelin synthase from Maytenus ilicifolia. The open reading frame (ORF) region of the PdFRS sequence was 2280bp long and composed a 759 amino acid protein with a predicted molecular mass of 87.81kDa. qPCR analysis revealed that methyl jasmonate treatments strongly upregulated PdFRS gene expression and resulted in enhanced friedelin accumulation in leaves. Heterologous expression of the PdFRS gene in yeast resulted in the production of friedelin triterpene as a single product, which was confirmed by comparison with the mass fragmentation pattern from an authentic friedelin standard by GC/MS analysis. Transgenic P. davidiana overexpressing the PdFRS gene was constructed via Agrobacterium-mediated transformation. Overexpression of PdFRS in transgenic P. davidiana lines resulted in enhanced friedelin production.

Toxicity assessment of plant extracts to Cleruchoides noackae Lin and Huber (Hymenoptera: Mymaridae)

Extracts of Matricaria chamomilla, Maytenus ilicifolia and Echinodorus grandiflorus have been integrated as viable pest management strategy for controlling bronze bug Thaumastocoris peregrinus (Hemipetra: Thaumastocoridae) due to their insecticidal activities. However, the side effects of these extracts on Cleruchoides noackae (Hymenoptera: Mymaridae), the main biological control agent against the bronze bug of eucalyptus, have never been studied. The objective of this study was to verify the effect of 5% aqueous extracts of M. chamomilla, M. ilicifolia and E. grandiflorus on C. noackae. Tests with eggs treated before being offered to the parasitoid female (pre-parasitism) and tests on which the eggs were treated 1 and 7 days after the female had performed oviposition (post-parasitism) were carried out. There was no difference between the treatments and the control group regarding parasitoid emergence in none of the tests. Although, there was increase in egg viability and number of emerged parasitoids when comparing the eggs treated 1 and 7 days after parasitism for all treatments. In conclusion, none of the plant extracts caused any harmful effect on C. noackae under lab conditions, indicating they may be an environment-friendly option to control the bronze bug of eucalyptus.

Friedelin in Maytenus ilicifolia Is Produced by Friedelin Synthase Isoforms.

Triterpenes are interesting compounds because they play an important role in cell homeostasis and a wide variety exhibiting defense functions is produced by plant secondary metabolism. Those same plant secondary metabolites also exhibit biological properties with promising therapeutic potential as anti-inflammatory and antitumor agents. Friedelin is a triterpene ketone with anti-inflammatory and gastroprotective activities and it is a precursor of relevant antitumor quinonemethides. Although many triterpene synthases have been described, only two friedelin synthases were characterized and there is no information about their genomic features and alleles. In the present work, we aimed to identify the gene and new isoforms of friedelin synthase in Maytenus ilicifolia leaves to be functionally characterized in Saccharomyces cerevisiae. The gene sequence analysis elucidated the exon/intron structure and confirmed the presence of single nucleotide polymorphisms with four non-synonymous mutations outside the active site of the enzyme. Therefore, two new isoforms were observed and the heterologous production of the enzymes in yeast showed similar production of friedelin. This first description of different alleles of the gene of friedelin synthase in M. ilicifolia can guide their validation as markers for friedelin-producer specimens.