High variability of perezone content in rhizomes of Acourtia cordata wild plants, environmental factors related, and proteomic analysis

With the aim of exploring the source of the high variability observed in the production of perezone, in Acourtia cordata wild plants, we analyze the influence of soil parameters and phenotypic characteristics on its perezone content. Perezone is a sesquiterpene quinone responsible for several pharmacological effects and the A. cordata plants are the natural source of this metabolite. The chemistry of perezone has been widely studied, however, no studies exist related to its production under natural conditions, nor to its biosynthesis and the environmental factors that affect the yield of this compound in wild plants. We also used a proteomic approach to detect differentially expressed proteins in wild plant rhizomes and compare the profiles of high vs. low perezone-producing plants. Our results show that in perezone-producing rhizomes, the presence of high concentrations of this compound could result from a positive response to the effects of some edaphic factors, such as total phosphorus (Pt), total nitrogen (Nt), ammonium (NH4), and organic matter (O. M.), but could also be due to a negative response to the soil pH value. Additionally, we identified 616 differentially expressed proteins between high and low perezone producers. According to the functional annotation of this comparison, the upregulated proteins were grouped in valine biosynthesis, breakdown of leucine and isoleucine, and secondary metabolism such as terpenoid biosynthesis. Downregulated proteins were grouped in basal metabolism processes, such as pyruvate and purine metabolism and glycolysis/gluconeogenesis. Our results suggest that soil parameters can impact the content of perezone in wild plants. Furthermore, we used proteomic resources to obtain data on the pathways expressed when A. cordata plants produce high and low concentrations of perezone. These data may be useful to further explore the possible relationship between perezone production and abiotic or biotic factors and the molecular mechanisms related to high and low perezone production.This work was supported by the Programa de Mejoramiento del Profesorado PROMEP/103.5/13/6626 and Consejo Nacional de Ciencia y Tecnología CONACyT-Mexico for Ph.D. scholarship 392123/254165. The University of Alicante lab is a member of Proteored, PRB3 and is supported by grant PT17/0019, of the PE I+D+I 2013-2016, funded by ISCIII and ERDF. Roque Bru-Martínez received financial support from the University of Alicante (VIGROB-105)

Diagnóstico y caracterización molecular de aislamientos de mycosphaerella sp. provenientes de plantaciones de banano y plátano de diferentes regiones de colombia.

Mycosphaerella fijiensis Morelet, agente causal de la Sigatoka negra, afecta dramáticamente la producción comercial de banano y plátano en la mayoría de las regiones productoras del mundo. En Colombia, la sigatoka negra se observó por primera vez en 1981 en la zona bananera del Urabá y desde entonces se disemino a todas las regiones, Atlantico, Pacifico, Centro y Oriente del país. En el 2000 la enfermedad fue encontrada en el Choco, zona del Pacifico Colombiano cubierta completamente por selva y que presenta cultivos de plátano en pequeñas parcelas al borde del rio Atrato, la única vía de acceso a la región. En este trabajo se realizó una prueba molecular de diagnóstico a 21 cepas de Mycosphaerella spp. Aisladas de diferentes zonas y se estudió la diversidad genética del patógeno en algunas regiones de Colombia utilizando marcadores RAPD’s. En total se obtuvieron 26 bandas polimórficas con los cebadores OPM01, OPM5 Y OPM20. El análisis de distancias genéticas sugiere que las cepas del Choco provienen de la zona del Urabá antioqueño y que en la zona de Santa Marta se presenta una subpoblación del patógeno diferente de las cepas del resto del país

Diagnóstico y caracterización molecular de aislamientos de Mycosphaerella sp. Provenientes de plantaciones de banano y plátano de diferentes regiones de Colombia.

Mycosphaerella fijiensis Morelet, agente causal de la Sigatoka negra, afecta dramáticamente la producción comercial de banano y plátano en la mayoría de las regiones productoras del mundo. En Colombia, la sigatoka negra se observó por primera vez en 1981 en la zona bananera del Urabá y desde entonces se disemino a todas las regiones, Atlantico, Pacifico, Centro y Oriente del país. En el 2000 la enfermedad fue encontrada en el Choco, zona del Pacifico Colombiano cubierta completamente por selva y que presenta cultivos de plátano en pequeñas parcelas al borde del rio Atrato, la única vía de acceso a la región. En este trabajo se realizó una prueba molecular de diagnóstico a 21 cepas de Mycosphaerella spp. Aisladas de diferentes zonas y se estudió la diversidad genética del patógeno en algunas regiones de Colombia utilizando marcadores RAPD’s. En total se obtuvieron 26 bandas polimórficas con los cebadores OPM01, OPM5 Y OPM20. El análisis de distancias genéticas sugiere que las cepas del Choco provienen de la zona del Urabá antioqueño y que en la zona de Santa Marta se presenta una subpoblación del patógeno diferente de las cepas del resto del país

<em>In Vitro</em> Seed Germination and Seedling Development of Two Avocado Varieties

Avocado (Persea americana Mill.) is a tree native to central and eastern Mexico. A basal angiosperm of the Lauraceae family, it produces an oil-rich fruit that is appreciated worldwide for its nutritional value. Mexico is the world’s leading avocado producer. Production is based mainly on the use of rootstocks of Persea americana var. drymifolia, a “Mexican native”. The agronomic characteristics of the rootstock are key to avocado production. This work reports a germination method to obtain seedlings in vitro from two avocado varieties, P. americana var. drymifolia and West Indian P. americana var. americana. With this system, germination success rates of 100% were obtained in a maximum of five days, with homogeneous seedling development. This system could provide rootstock that improves the characteristics of agronomic programs and the selection of genetic material for avocado production

Production of the Anti-Inflammatory Compound 6-O-Palmitoyl-3-O-β-D-glucopyranosylcampesterol by Callus Cultures of Lopezia racemosa Cav. (Onagraceae)

Lopezia racemosa Cav. is a plant used in Mexican traditional medicine to heal inflammatory diseases. From this plant we isolated the novel compound 6-O-palmitoyl- 3-O-β-D-glucopyranosylcampesterol (1) and 6-O-palmitoyl-3-O-β-D-glucopyranosyl-β-sitosterol (2), previously reported to have cytotoxic activity on several cancer cell lines. We evaluated the anti-inflammatory activity of 1 in vivo by mouse ear edema induced with 12-O-tetradecanoylphorbol-13-acetate (TPA) and 57.14% inhibition was observed. The aim of our study was to obtain callus cultures derived from this plant species with the ability to produce the compounds of interest. Callus cultures were initiated on MS basal medium amended with variable amounts of naphthaleneacetic acid (NAA), or 2,4-dichlorophenoxyacetic acid (2,4-D), combined or not with 6-benzylaminopurine (BAP). Ten treatments with these growth regulators were carried out, using in vitro germinated seedlings as source of three different explants: hypocotyl, stem node, and leaf. Highest yield of 1 was observed on callus derived from leaf explants growing in medium containing 1.0 mg/L 2,4-D and 0.5 mg/L BAP. Selected callus lines produced less 1 than wild plants but the in vitro cultured seedlings showed higher production. So we conclude that it could be attractive to further investigate their metabolic potential

Establishment and Phytochemical Analysis of a Callus Culture from Ageratina pichinchensis (Asteraceae) and Its Anti-Inflammatory Activity

A protocol was established to produce bioactive compounds in a callus culture of Ageratina pichinchensis by using 1 mg L&minus;1 NAA with 0.1 mg L&minus;1 KIN. The phytochemical study of the EtOAc extract obtained from the callus biomass, allowed the isolation and characterization of eleven secondary metabolites, of which dihydrobenzofuran (5) and 3-epilupeol (7), showed important anti-inflammatory activity. Compound 5 inhibits in vitro the secretion of NO (IC50 = 36.96 &plusmn; 1.06 &mu;M), IL-6 (IC50 = 73.71 &plusmn; 3.21 &mu;M), and TNF-&alpha; (IC50 = 73.20 &plusmn; 5.99 &mu;M) in RAW (Murine macrophage cells) 264.7 macrophages, as well as the activation of NF-&kappa;B (40% at 150 &mu;M) in RAW-blue macrophages, while compound 7 has been described that inhibit the in vivo TPA-induced ear edema, and the in vitro production of NO, and the PLA2 enzyme activity. In addition, quantitative GC-MS analysis showed that the anti-inflammatory metabolites 5 and 7 were not detected in the wild plant. Overall, our results indicated that A. pichinchensis can be used as an alternative biotechnological resource for obtaining anti-inflammatory compounds. This is the first report of the anti-inflammatory activity of compound 5 and its production in a callus culture of A. pichinchensis

A Cytotoxic and Anti-inflammatory Campesterol Derivative from Genetically Transformed Hairy Roots of Lopezia racemosa Cav. (Onagraceae)

The genetically transformed hairy root line LRT 7.31 obtained by infecting leaf explants of Lopezia racemosa Cav with the Agrobacterium rhizogenes strain ATCC15834/pTDT, was evaluated to identify the anti-inflammatory and cytotoxic compounds reported previously for the wild plant. After several subcultures of the LRT 7.31 line, the bio-guided fractionation of the dichloromethane–methanol (1:1) extract obtained from dry biomass afforded a fraction that showed important in vivo anti-inflammatory, and in vitro cytotoxic activities. Chemical separation of the active fraction allowed us to identify the triterpenes ursolic (1) and oleanolic (2) acids, and (23R)-2α,3β,23,28-tetrahydroxy-14,15-dehydrocampesterol (3) as the anti-inflammatory principles of the active fraction. A new molecule 3 was characterized by spectroscopic analysis of its tetraacetate derivative 3a. This compound was not described in previous reports of callus cultures, in vitro germinated seedlings and wild plant extracts of whole L. racemosa plants. The anti-inflammatory and cytotoxic activities displayed by the fraction are associated to the presence of compounds 1–3. The present study reports the obtaining of the transformed hairy roots, the bioguided isolation of the new molecule 3, and its structure characterization

Spontaneous Regeneration of Plantlets Derived from Hairy Root Cultures of <i>Lopezia racemosa</i> and the Cytotoxic Activity of Their Organic Extracts

A histological analysis was performed with the aim of elucidating the spontaneous regeneration process of the hairy root lines LRT 2.3 and LRT 6.4, derived from Lopezia racemosa leaf explants and genetically transformed with the Agrobacterium rhizogenes strain ATCC15834/pTDT. The analysis showed both lines regenerate via indirect somatic embryogenesis; LRT 6.4 also regenerated by direct organogenesis. The morphogenic characteristics of the regenerated plantlets from both lines showed the typical characteristics, described previously, including a higher number of axillary shoot formation, short internodes, and plagiotropic roots compared with wild-type seedlings. The regeneration process occurred without the addition of plant growth regulators and was linked to the sucrose concentration in the culture medium. Reducing the sucrose concentration from 3% to 2%, 1%, and 0.5% increased the regeneration rate in LRT 6.4; the effect was less pronounced in LRT 2.3. The cytotoxic activity of different organic extracts obtained from roots and shoots were evaluated in the cancer cell lines HeLa (cervical carcinoma), HCT-15 (colon adenocarcinoma), and OVCAR (ovary carcinoma). The hexane and dichloromethane extracts from roots of both lines showed cytotoxic activity against the HeLa cell line. Only the dichloromethane extract from the roots of PLRT 2.3 showed cytotoxic activity against the OVCAR cell line. None of the methanol extracts showed cytotoxic activity, nor the shoot extracts from any solvent

LC-MS Fingerprinting Development for Standardized Precipitate from <i>Agastache mexicana</i>, Which Induces Antihypertensive Effect through NO Production and Calcium Channel Blockade

The aim of this work was to evaluate the vasorelaxant and antihypertensive effects of a standardized precipitate of the hydroalcoholic extract from Agastache mexicana (PPAm), comprising ursolic acid, oleanolic acid, acacetin, luteolin and tilianin, among others. In the ex vivo experiments, preincubation with L-NAME (nonspecific inhibitor of nitric oxide synthases) reduced the relaxation induced by PPAm; nevertheless, preincubation with indomethacin (nonspecific inhibitor of cyclooxygenases) did not generate any change in the vasorelaxation, and an opposed effect was observed to the contraction generated by CaCl2 addition. Oral administration of 100 mg/kg of PPAm induced a significant acute decrease in diastolic (DBP) and systolic (SBP) blood pressure in spontaneously hypertensive rats, without changes in heart rate. Additionally, PPAm showed a sustained antihypertensive subacute effect on both DBP and SBP for 10 days compared to the control group. On the other hand, human umbilical vein cells treated with 10 µg/mL of PPAm showed a significant reduction (p Am induces a significant antihypertensive effect in acute- and subacute-period treatments, due to its direct vasorelaxant action on rat aortic rings through NO production and Ca2+ channel blockade