In vitro evaluation of the effects of Viscum album homeopathic preparations on murine melanoma cells

Viscum album extract (VA) is a complementary treatment in cancer, with in vitro and in vivo cytotoxic effects on several tumor types when applied in phytochemical doses. However, highly diluted ethanolic homeopathic preparations' effects and mechanisms need further study. Aims:To assess the in vitro effects of highly diluted VA from the subspecies V. album abietis and V. album album at different potency levels in different dilution ratios on murine melanoma cells. Methodology:The VA mother tinctures (MT)from Abies alba (MTA) and Quercus robur (MTQ) were prepared with summer and winter samples, harvested in Switzerland. They were submitted to homeopathic ethanolic maceration and a subsequent dynamization process. MTA, MTQ and the following respective potencies were tested in B16F10 murine cells: 3x, 12x, 30x, 6cH, 12cH, 200cH, 2LM, 3LM, and 5LM. Dynamized water, dynamized and non-dynamized ethanol, and carboplatin were used as control groups. The mitochondrial activity and cell viability analysis were performed at 1, 24, 48, and 72 hours by in vitro incubation. MTA and MTQ harvested in summer, as well as 12x, 200cH and 5LM potencies were also tested to cell apoptosis and necrosis markers, reactive oxygens species (ROS) production, inflammatory cytokines profile, cell morphology, and migratory capacity. Results and discussion: MTA and MTQ induced a decrease in cell metabolism and higher cytotoxicity within 1 hour, with significant morphological changes and increased production of ROS and inflammatory cytokines. Both homeopathic dilutions 12x and 5LM showed an influence on cell metabolism, cell replication, and oxidative stress modulation with inflammatory cytokines, mitosis, and migration pattern changes. On the other hand, Quercus robur and Abies alba 200cH showed increased on cytotoxicity and ROS levels, respectively. Conclusion:The in vitro effects of Viscum album homeopathic solutions in melanoma cells highlight the promising antitumoral potential and reinforce the need for further research to better understanding their mechanisms of action.

Improvement of ethanol production from birch and spruce pretreated with 1-H-3-methylmorpholinium chloride

Background: Pretreatment is the critically important step for the production of ethanol from lignocelluloses. In this study, hardwood birch (Betula pendula) and softwood spruce (Norway spruce) woods were pretreated with a newly synthesized morpholinium ionic liquid, 1-H-3-methylmorpholinium chloride ([HMMorph][Cl]), followed by enzymatic hydrolysis and fermentation to ethanol. Results: [HMMorph][Cl] was synthesized using inexpensive raw materials, i.e., hydrochloric acid and N-methyl morpholine, following a simple process. The influence of pretreatment time (2, 3, 5, and 8 h) and temperature (120 and 140°C) in terms of hydrolysis efficiency was investigated. Glucose yields from enzymatic hydrolysis were improved from 13.7% to 45.7% and 12.9% to 51.8% after pretreatment of birch and spruce woods, respectively, under optimum pretreatment conditions (i.e., at 140°C for 3 h) as compared to those from pristine woods. Moreover, the yields of ethanol production from birch and spruce were increased to 34.8% and 44.2%, respectively, while the yields were negligible for untreated woods. Conclusions: This study demonstrated the ability of [HMMorph][Cl] as an inexpensive agent to pretreat both softwood and hardwood.

Bioactividad de extractos de seis especies vegetales nodrizas de bosques de pinabete (Abies guatemalensis Rehder) de Ixchiguán, San Marcos, Guatemala / Bioactivity of extracts from six nurse plants species from fir (Abies guatemalensis Rehder) forests from Ixchiguan, San Marcos, Guatemala

Plantas nodrizas son aquellas especies vegetales útiles para la sobrevivencia y convivencia de otras especies. Abies guatemalensis Rehder (pinabete), es una especie endémica en peligro de extinción y para asegurar su sobrevivencia es necesario estudiar sus plantas nodrizas. Se colectaron seis especies de plantas nodrizas provenientes de bosque Los Cuervos, Ixchiguán, San Marcos, se secaron a la sombra en un horno, hasta obtener un porcentaje de humedad < 10%. Se obtuvieron extractos etanólicos por percolación y concentración empleando rotavapor; los aceites senciales fueron extraídos por hidrodestilación con Neoclevenger. Para los análisis de actividad biológica se realizaron ensayos contra bacterias, hongos, levaduras, larvas de insectos, nauplios de Artemia salina y actividad antioxidante. Los aceites esenciales presentaron un porcentaje de rendimiento < 0.3%. Cinco de los seis extractos etanólicos demostraron baja actividad antimicrobiana y larvicida (CIM: 1 mg/mL). Respecto a la actividad antioxidante por DPPH, Acaena elongata (CI50 de 0.21 [0.019, 0.23] mg/mL) y Rubus trilobus (CI50 0.32 [0.31, 0.33] mg/mL) presentaron los mejores resultados y Buddleja megalocephala (CI50 0.75 [0.72, 0.77] mg/mL) presentó actividad moderada. Estos hallazgos estimulan a continuar la investigación de plantas nodrizas para identificar las moléculas responsables de la actividad antioxidante y definir su posible aplicación como antioxidantes para la prevención o tratamiento de patologías humanas o para la preservación de alimentos o uso cosmético, permitiendo que las comunidades conserven y aprovechen de manera sostenible dichas especies.

Nurse plants are species useful for the survival and coexistence of other plants. Abies guatemalensis Rehder (fir) is an endemic species in danger of extinction and in order to ensure its survival, it is necessary to study their nurse plants. Six species of nurse plants were collected from Los Cuervos forest, Ixchiguán, San Marcos, they were dried in the shade and in an oven until a moisture content < 10% was obtained. Ethanol extracts were obtained by percolation followed by concentration in rotavapor; essential oils were obtained by hydrodistillation with Neoclevenger. Assays for biological activity were established against bacteria, fungi, yeasts, insect larvae, nauplii of Artemia salina and antioxidant activity. The essential oils yields were < 0.3%. Extracts have little antimicrobial and larvicidal activity with a MIC 1 mg/mL of the ethanol extracts. The free radical trapping activity assay evaluated by DPPH, yielded good results: Acaena elongata (IC50 of 0.21 [0.019, 0.23] mg/mL) and Rubus trilobus (IC50 0.32 [0.31, 0.33] mg/mL) and moderate activity in Buddleja megalocephala (IC50 0.75 [0.72, 0.77] mg/mL). These findings stimulate research on nurse plants to identify the molecules responsible for the antioxidant activity and to define its possible application for the prevention or treatment of human pathologies or for the preservation of food or cosmetic use, allowing the communities to preserve and take advantage on a sustainable way.

Modelo de negocio de restauración productiva de Abies guatemalensis Rehder utilizando plantas nodrizas para su establecimiento / Business model of productive restoration of Abies guatemalensis Rehder using nurse plants for its establishment

El presente reporte de caso tuvo como objetivo generar el modelo de negocios para el pinabete (Abies guatemalensis Rehder) en restauración, no se pretende hacer un análisis profundo de la producción de árboles de pinabete, únicamente mostrar que la producción se puede combinar con la conservación. En la región alta del altiplano occidental de Guatemala hay muchas tierras degradadas fuera de bosques, donde la sucesión vegetal ha promovido el crecimiento de especies arbustivas, útiles como nodrizas en el establecimiento de pinabete para protegerlas de heladas y lograr establecimiento mayor a 80 %, lo cual ya se ha puesto en práctica por varios silvicultores locales. Se realizó un análisis técnico financiero sobre la producción de pinabete con doble propósito: especie comercial y para la restauración de áreas degradadas. El modelo de negocios se estableció para 0.25 ha, con un precio de venta por árbol de Q. 300, con y sin incentivos forestales, considerando establecer previamente plantas arbustivas. Los resultados indican que con una inversión de Q. 45,227 durante ocho años a una tasa de descuento del 13 %, el Valor Actual Neto (VAN) toma valores entre Q. 398,576 a 742,020 con una Tasa Interna de Rendimiento (TIR) entre 30 y 50 %. A la vez se preparó un plan de negocios utilizando la metodología Canvas, que establece un plan estratégico que permite visualizar lo que actualmente se está realizando y qué situaciones hay que mejorar. El modelo propuesto puede utilizarse de una forma mixta para la conservación al dejar al menos un 30 % de los árboles, la otra parte se puede utilizar para comercialización a partir del octavo al 12do año de forma sostenible.

The present case study aimed to generate the business model for the Guatemalan fir (Abies guatemalensis Rehder) under restoration process. In the highlands of the altiplano region in western Guatemala there are many degraded lands outside of forests, where the ecological succession has promoted the growth of shrub species that help to the establishment of the fir and to protect it from frost. This condition also helped the specie to achieve more than 80 % of survival rate. The method has been implemented by several local silvicultors. A technical and financial analysis was carried out on the production of fir with two objectives: production of commercial Christmas trees and for the restoration of degraded areas. The business model was established for an area of 0.25 ha, with an estimated market price per tree of Q. 300, with and without forestry incentives, assuming previously establish shrub plants. The results indicate that with an investment of Q. 45,227 for eight years at a discount rate of 13 %, the Net Present Value (NPV) takes values between Q. 398,576 to 742,020 with an Internal Rate of Return (IRR) ranging between 30 and 50 %. At the same time, a business plan was prepared using the Canvas methodology, which establishes a strategic plan that allows visualizing what is currently being done and what areas need improvement. The proposed model can be used in a mixed way for conservation by leaving at least 30 % of the trees; the other part can be used for trading the trees from the eighth to the twelfth years old in a sustainable way.

Características in situ y de propagación de seis especies arbustivas utilizadas como plantas nodrizas en la región occidental de Guatemala / In situ features and propagation of six shrub used as plantas nurses in western Guatemala

En la restauración de ecosistemas, una técnica es utilizar arbustos de la sucesión como nodriza. En la parte alta de San Marcos, Guatemala, se ha recurrido al menos a seis de ellos en el establecimiento de pino y pinabete: arrayán (Baccharis vaccinoides Kunth), salvia (Buddleia megalocephala Donn. Sm.), mozote (Acaena elongata L.), chicajol (Stevia polycephala Bertol), mora (Rubus trilobus Ser.) y malacate (Symphoricarpos microphyllus Kunth). Se carece de la documentación de su comportamiento y características que puedan servir para utilizarlas ampliamente como plantas protectoras. En esta investigación se estudió características in situ y reproductivas de estas especies. Entre los principales resultados está que la altura de plantas es de 1.17 a 2.64 m y cobertura de 0.34 a 2.77 m2, con una amplia variación entre y dentro de especies. La cantidad de frutos/planta va desde 90 en S. microphyllus a 59,400 en B. vaccinioides. La propagación vegetativa en campo estuvo entre 48 a 92 %, mayor que cuando se realizó en vivero, donde B. vaccinoides, S. microphyllus y R. trilobus tuvieron mejor respuesta. La propagación por semilla muestra que las seis especies pueden reproducirse adecuadamente. Estos resultados permiten su reproducción en viveros para su uso en restauración ecológica.

In the restoration of ecosystems, one technique is to use shrubs of succession as nurse plants. In the highlands of San Marcos, Guatemala, at least six of these shrubs have been used in the establishment of pine and fir: arrayán (Baccharis vaccinoides Kunth), salvia (Buddleia megalocephala Donn Sm.), mozote (Acaena elongata L.) chicajol (Stevia polycephala Bertol), mora (Rubus trilobus Ser.) and malacate (Symphoricarpos microphyllus Kunth). There is a lack of documentation about their behavior and their characteristics that may make them useful to serve as protective plants. In this research in situ and reproductive characteristics of these species was studied. The main results reveal that the plants’ height is 1.17 to 2.64 m and have a coverage of 0.34 to 2.77 m2, with a wide variation between and within species. The amount of fruit per plant ranges from 90 in S. microphyllus to 59,400 in B. vaccinioides. Vegetative propagation in the field was 48-92% higher than when it was performed in a nursery, where B. vaccinoides, S. microphyllus and R. trilobus had better responses. Seed propagation shows that the six species can adequately reproduce. These results may allow for reproduction in nurseries to be used in ecological restoration.

Plantas asociadas a los bosques de Abies guatemalensis (Pinaceae) del occidente de Guatemala / Plants associated to Abies guatemalensis (Pinaceae) forests in Western Guatemala

The fragments of Abies guatemalensis forests in Western Guatemala are the reservoirs of plant species that have been poorly documented, missing the opportunity to expand the knowledge of the local flora and its use in conservation planning. To assess this, a floristic study was done in areas between 2 950-3 360masl in Western Guatemala between 2010-2011. Ten locations were sampled: in each a 500m² plot was surveyed, and plants were classified in four strata by plant height (0.05-30m). A total of 119 species, 92 genera and 50 families in four divisions were found. The families with more species were Asteraceae, Poaceae, Rosaceae, Lamiaceae, Apiaceae and Solanaceae, and the most abundant genera were Salvia, Alchemilla and Bidens. The number of species found by strata was: 33 (low herbaceous), 49 (high herbaceous), 30 (shrubs) and seven in the tree strata. Regarding geographical distribution, the biggest species group detected was from central Mexico to Central America with 67%, which compared to the forests of A. guatemalensis in central and Southern Mexico, showed high floristic affinity, especially at the family and genus level. However, even having families and genera in common in the general structure of the fir forests, their floristic particularities should be taken into account when making management and conservation plans, because these are influenced by soil, latitude and microclimate conditions.

Hay una carencia de información detallada sobre la composición y estructura de las comunidades montanas guatemaltecas. El objetivo del estudio fue contribuir al conocimiento de la flora de bosques de abeto (Abies guatemalensis), para esto se hizo el levantamiento florístico en bosques de abeto del occidente de Guatemala. Se encontraron 119 especies, 92 géneros, 50 familias en cuatro divisiones. Las familias más numerosas fueron: Asteraceae, Poaceae, Rosaceae, Lamiaceae, Apiaceae y Solanaceae y los géneros más abundantes Salvia, Alchemilla y Bidens. Las especies se ubicaron en cuatro estratos, 33 en el herbáceo inferior, 49 en el herbáceo superior, 30 en arbustos y siete en árboles. Se hace énfasis en la contribución del estudio al conocimiento de la flora de bosques de A. guatemalensis y la necesidad de otros similares en los demás bosques de esta especie, que sirva para fomentar su conocimiento y conservación. Se consideraron seis grupos de distribución geográfica, el principal es del centro de México a Centroamérica con 67% de especies. Se realiza una comparación fitogeográfica y de composición florística, con otras áreas de Abies de Guatemala y México. Se propone que a pesar de haber familias y géneros comunes, que proporcionan la estructura general entre los bosques de abeto, se deben considerar las particularidades florísticas de cada área, en el manejo y conservación influidas por suelo, latitud y microclima.

Calli cultures from Abies equi-trojani (Aschers et Sinten) and changes in antioxidant defense system enzymes.

This study was conducted to explain difficulties of indirect regeneration of forest trees in tissue culture conditions. For this purpose, changes of antioxidant defense system enzymes; superoxide dismutase (SOD), peroxidase (POD) activities were determined during calli formation on young apical shoots of Abies equi-trojani (Aschers et Sinten). Young apical shoots were collected from naturally growing trees and cultured on two different media; Murashige and Skoog (MS) and McCown Woody plant medium (WPM) supplemented with growth regulators benzyl amino purine (BAP), 2,4-dichloro phenoxy acetic acid (2.4-D), kinetin (Kn) in various concentrations for callus induction. WPM media containing 1 mg ml(-1) BAP and 1 mg ml(-1) 2,4-D gave the best calli induction ratio (74%) between tested combinations. POD and SOD enzyme activities were measured both on young shoot explants and 10 day-old calli derived from these explants. POD and SOD enzyme activities were higher being 81.02% and 74.82%, respectively on calli when compared to shoots. The results showed that culture stress tolerated with increased antioxidant enzyme activities could be considered as protective physiological responses in calli cells.

Biomass distribution in declining sacred-fir seedling

En los bosques localizados en el oeste y suroeste de la Ciudad de México se ha observado una declinación del oyamel Abies religiosa (H.B.K.) Schl. et Cham. durante los últimos 25 años. La caracterización de este fenómeno desde diferentes puntos de vista es importante si se pretende incrementar el conocimiento sobre el problema y proponer soluciones viables. Por esta razón se estudiaron los cambios en los patrones de distribución de biomasa en plántulas de 7 años de edad, como resultado de la declinación. Se estableció un experimento completamente al azar con tres niveles de daño como tratamientos: ligero, intermedio y severo, y ocho repeticiones por tratamiento. La biomasa de los componentes de las plántulas se redujo significativamente a medida que se incrementó el daño. La raíz fue el componente más afectado, seguido de las ramillas. Sin embargo, estas últimas acumularon biomasa con el tiempo, produciendo un desbalance entre la biomasa aérea y subterránea. Aun cuando este desbalance se compensa por la caída de follaje, trae como consecuencia una reducción de la cantidad de carbono fijado

Screening for Antifungal Endophytic Fungi Against Six Plant Pathogenic Fungi

A total of 187 endophytic fungi were isolated from 11 plant species, which were collected from 11 locations in Korea. Their antifungal activities were screened in vivo by antifungal bioassays after they were cultured in potato dextrose broth and rice solid media. Antifungal activity against plant pathogenic fungi such as Magnaporthe grisea (rice blast), Corticium sasaki (rice sheath blight), Botrytis cinerea (tomato gray mold), Phytophthora infestans (tomato late blight), Puccinia recondita (wheat leaf rust), and Blumeria graminis f. sp. hordei (barley powdery mildew) was determined in vivo by observing the inhibition of plant disease development. Twenty (11.7%) endophytic fungi fermentation broths were able to control, by more than 90%, at least one of the six plant diseases tested. Among 187 liquid broths, the F0010 strain isolated from Abies holophylla had the most potent disease control activity; it showed control values of more than 90% against five plant diseases, except for tomato late blight. On the other hand, fourteen (7.5%) solid culture extracts exhibited potent disease control values of more than 90% against one of six plant diseases. The screening results of this study strongly suggested that metabolites of plant endophytic fungi could be good potential sources for screening programs of bioactive natural products.