Changes in Growth and Heavy Metal and Phenolic Compound Accumulation in Buddleja cordata Cell Suspension Culture under Cu, Fe, Mn, and Zn Enrichment.

Buddleja cordata cell suspension cultures could be used as a tool for investigating the capabilities of this species to tolerate heavy metals (HMs) and for assessing the effects of HMs on the accumulation of phenolic compounds in this species. It grows in a wide range of habitats in Mexico, including ultramafic soils, and mobilizes some HMs in the soil. The mobilization of these HMs has been associated with phenolic substances. In addition, this species is used in Mexican traditional medicine. In the present study, a B. cordata cell suspension culture was grown for 18 days in a culture medium enriched with Cu (0.03-0.25 mM), Fe (0.25-1.5 mM), Mn (0.5-3.0 mM), or Zn (0.5-2.0 mM) to determine the effects of these HMs on growth and HM accumulation. We also assessed the effects of the HMs on phenolic compound accumulation after 1 and 18 days of HM exposure. Cells were able to grow at almost all tested HM concentrations and accumulated significant amounts of each HM. The highest accumulation levels were as follows: 1160 mg Cu kg-1, 6845 mg Fe kg-1, 3770 mg Mn kg-1, and 6581 mg Zn kg-1. Phenolic compound accumulation was affected by the HM exposure time and corresponded to each HM and its concentration. Future research should analyze whole plants to determine the capabilities of Buddleja cordata to accumulate abnormally high amounts of HM and to evaluate the physiological impact of changes in the accumulation of phenolic compounds.

Cordyceps mexicana sp. nov., parasitizing Paradirphia sp. moths: A new sister species of the Cordyceps militaris complex, distributed in central Mexican Quercus-Pinus mixed forests.

Cordyceps s.l. is a paraphyletic group of ascomycete fungi that exhibit multifunctional lifestyles, that is, as saprotrophs, endophytes, and pathogens of insects, spiders, fungi, and grasses. The family Cordycipitaceae includes macroscopically similar species that have been erroneously considered to be conspecific with Cordyceps militaris. In this study, we describe a new species within the C. militaris complex that is distributed in the Quercus-Pinus forests of central Mexico on the basis of its morphology, phylogenetic relationships, and life cycle. Phylogenetically, Cordyceps mexicana is a well-supported new sister species of the C. militaris complex. It is distinguished by the morphology of its conidiophore, host association, and geographic distribution. This species parasitizes pupae of Paradirphia sp. (Lepidoptera: Saturniidae: Hemileucinae) and might be macroscopically confused with C. militaris. Its stromata are large, can measure up to 10 cm in length, and the fertile part is always bright yellow. This species develops whitish mycelial cords that emerge from the stromata and grow toward the host. Microscopically, it develops asci with filiform ascospores disarticulating in part-spores. Its life cycle and geographic distribution are also discussed.

Arnica montana Cell Culture Establishment, and Assessment of Its Cytotoxic, Antibacterial, α-Amylase Inhibitor, and Antioxidant In Vitro Bioactivities.

Arnica montana cell suspension culture could be a sustainable source of a vegetal material producer of secondary metabolites (SMs) possessing biological effects. Different plant growth regulator concentrations (0-5 mg/L) were tested in foliar explants to induce a callus that was used to establish a cell suspension culture. Growth kinetics was carried out for 30 days. A methanolic extract obtained from biomass harvested at 30 days of growth kinetics was fractionated, and three fractions were tested for bioactivities. We induced a callus with 1 mg/L of picloram and 0.5 mg/L of kinetin in foliar explants, which allowed for the establishment of a cell suspension culture, and the latter had the highest total SMs contents at day 30. Three fractions showed differences in total SMs contents, with the highest values per gram as follows: 270 mg gallic acid equivalent for total phenolic content, 200 mg quercetin equivalent for total flavonoid content, 83 mg verbascoside equivalent for total phenolic acid content, and 396 mg parthenolide equivalent for total sesquiterpene lactone content. The best bioactivities were 2-6 µg/mL for the 50% inhibition of 2,2-diphenyl-1-picrylhydrazyl radical, 30% cellular viability of lymphoma cells at 40 µg/mL, 17% inhibition against Escherichia coli and Staphylococcus aureus at 8 µg/disk, and α-amylase inhibition at 12% with 10 µg/mL. The total SMs contents were correlated with bioactivities.

Photoprotective Activity of Buddleja cordata Cell Culture Methanolic Extract on UVB-irradiated 3T3-Swiss Albino Fibroblasts.

The research on compounds exhibiting photoprotection against ultraviolet radiation (UVR) is a matter of increasing interest. The methanolic extract of a cell culture of Buddleja cordata has potential photoprotective effects as these cells produce phenolic secondary metabolites (SMs). These metabolites are attributed with biological activities capable of counteracting the harmful effects caused by UVR on skin. In the present work, the methanolic extract (310-2500 µg/mL) of B. cordata cell culture showed a photoprotective effect on UVB-irradiated 3T3-Swiss albino fibroblasts with a significant increase in cell viability. The greatest photoprotective effect (75%) of the extract was observed at 2500 µg/mL, which was statistically comparable with that of 250 µg/mL verbascoside, used as positive control. In addition, concentrations of the extract higher than 2500 µg/mL resulted in decreased cell viability (≤83%) after 24 h of exposure. Phytochemical analysis of the extract allowed us to determine that it was characterized by high concentrations of total phenol and total phenolic acid contents (138 ± 4.7 mg gallic acid equivalents and 44.01 ± 1.33 mg verbascoside equivalents per gram of extract, respectively) as well as absorption of UV light (first and second bands peaking at 294 and 330 nm, respectively). Some phenylethanoid glycosides were identified from the extract.

In vivo anti-arthritic effect and repeated dose toxicity of standardized methanolic extracts of Buddleja cordata Kunth (Scrophulariaceae) wild plant leaves and cell culture.

ETHNOPHARMACOLOGICAL RELEVANCE: Buddleja cordata Humb. Bonpl. & Kunth, known by the population as Tepozán blanco, is a shrub plant used in traditional herbal medicine in Mexico for the treatment of tumors, cancer, sores, skin burns, rheumatic pains and diseases related to inflammatory processes such as arthritis; authors adjudicate this etno-medicinal effect to the presence of secondary metabolites in the plant such as verbascoside, however due to its low concentration in recent years biotechnological tools are applied as cell culture to biosynthesize these pharmacological active metabolites in greater quantities. AIM OF THE STUDY: Evaluate the possible toxic effect after a daily administration of MeOH extracts from wild plant leaves (Bc-Wp), and cell culture (Bc-Cc) of B. cordata for 28 days, and after their anti-edematous and antioxidant activities in vivo, as well their effect on the cytokines profile during experimental arthritis induced by complete Freund's adjuvant. MATERIALS AND METHODS: Both extracts were evaluated in CD1 male mice first in a toxicity test of repeated dose administrations (1 g/kg) for 28 days, after which pharmacological activity of both extracts was measure during experimental induced arthritis where three doses were tested, at the end of the study edema formation, body weight gain and antioxidant activity were measure in edema and ganglionic tissues. Finally, dose that exerted the best protective effect (250 mg/kg) was evaluated to quantify its effect over ganglionic tissue concentration of lymphocytes T CD4+, and cytokines (IL-1ß, TNF-α and IL-10), as well histological analysis in arthritic mice. RESULTS: Both extracts (Bc-Wp and Bc-Cc) did not exert lethality, however body weight gain and food in-take were lower than in healthy mice administered with vehicles, also extract-treated animals showed a decrease in serum lipid concentration and only Bc-Wp extract treated animals decrease serum alkaline phosphatase after 28 daily administration compared to healthy un-treated mice. During experimental arthritis best inhibition effect over edema development was observed in those animals administered with both extracts at dose of 250 mg/kg (Bc-Wp and Bc-Cc) on day 28, compared to CFA un-treated mice. Also both extracts reduce oxidative damage over lipids and proteins at the same dose, in both ganglionic and edema tissue, as well antioxidant enzymatic response was reduced in both tissues compared to arthritic un-treated group. In ganglionic tissue of arthritic mice, CD4+ lymphocytes concentration was reduced by Bc-Wp and Bc-Cc treatment (250 mg/kg) respectively, as well IL-1ß, and TNF-α levels. Only arthritic animals treated with Bc-Cc extract at 250 mg/kg generated a significant increase of IL-10 doubling the levels compared to CFA un-treated group. Histological analysis of popliteal ganglion showed that both extracts decrease the incidence of lytic lesions, lipid inclusions and leukocyte infiltration. CONCLUSION: Methanolic extracts of wild Buddleja cordata and its cell cultures did not generated lethality after a daily administration for 28 days at 1 g/kg, but it was observed that both showed a lipid-lowering effect. Also at dose of 250 mg/kg both extracts exerted anti-edematous, protection against the oxidation of lipid and proteins, regulation on antioxidant enzymatic response, down-regulation on lymphocytes CD4+ producers of IL-1ß and TNF-α, an increase in IL-10 levels, which caused a decrease in leukocyte infiltration in ganglionic tissue during experimental arthritis.

Procedure for Estimating the Tolerance and Accumulation of Heavy Metals Using Plant Cell Cultures.

The tolerance index (TI) and the bioaccumulation factor (BF) for the estimation of accumulation and tolerance of different heavy metals in cell suspension cultures are reviewed. Procedures for measuring these parameters are described for the purposes of phytoremediation research.

Anti-staphylococcal properties of four plant extracts against sensitive and multi-resistant bacterial strains isolated from cattle and rabbits.

The aim of this study is to investigate the biopotency of methanolic extracts of Vitex mollis, Psidium guajava, Dalbergia retusa, and Crescential alata leaves against various staphylococcal strains isolated from cattle and rabbits. Methicillin-resistant S. aureus strains were isolated from cattle, while other strains were isolated from rabbits using standard methodology. The total phytochemical phenolic and saponins contents were obtained being the main groups of the antinutritional factors. The antimicrobial activity of the extracts against the standard culture of S. aureus (control) and S. aureus isolated from cattle and rabbits were investigated comparatively relative to that of oxacillin. It was found that both the control S. aureus and the isolated S. aureus are susceptible to all the four plant extracts, and sensitive to oxacillin. Of all the S. aureus including the control, MRSA2 is the most susceptible to all the extracts at 1000 µg/mL, except that of V. mollis where it is the least susceptible. Among all the plant extracts, P. guajava is the most active against MRSA2 and SOSA2. Therefore, the isolates from cattle (MRSA1 and MRSA2) are more susceptible to all the plant extracts than the isolates from rabbits. Among all the rabbit isolates, CoNS3 is the least susceptible to the extracts. Since all the plant extracts exhibit remarkable inhibitory activities against all the S. aureus strains, they are promising towards the production of therapeutic drugs.

Anti-staphylococcal properties of Eichhornia crassipes, Pistacia vera, and Ziziphus amole leaf extracts: Isolates from cattle and rabbits.

The desideratum aim of the present context was to assess the biopotency of methanolic extracts of Eichhornia crassipes (E. crassipes), Pistacia vera (P. vera), and Ziziphus amole (Z. amole) leaves against various staphylococcal strains, and to quantify the phenolics as well as saponin content in them. The antibacterial activity of various concentrations (62.5-1000 µg/mL) of plant extracts was tested against control clinical strains (Staphylococcus aureus ATCC 25923, S. aureus ATCC 29213, and S. aureus ATCC 43300), methicillin-resistant S. aureus (MRSA1 and MRSA2), oxacillin sensitive S. aureus (SOSA1 and SOSA2), and coagulase-negative Staphylococcus epidermidis (CoNS1, CoNS2, and CoNS3) using disc diffusion assay. Leaf extracts of the three plants exhibited pronounced growth inhibitory characteristics against staphylococci in a dose dependent manner. E. crassipes extract depicted the highest relative percentage inhibition values against control clinical strains (68.6 ± 0.5%), while P. vera (68.6 ± 0.3%) and Z. amole (74.79 ± 0.3%) extracts showed pronounced relative inhibition values against staphylococcal strains isolated from cattle. Total phenols and saponin content of leaf extracts were investigated by standard in vitro methods. The methanolic extracts of these plants were found to comprise substantial content of phenolics and saponin at varying levels. The highest value of phenolics was estimated in P. vera extract (60.0 ± 1.3 mg gallic acid/g extract), followed by Z. amole (33.6 ± 1.4 mg gallic acid/g extract), and E. crassipes (23.0 ± 1.3 mg gallic acid/g extract). Saponin content for P. vera, Z. amole, and E. crassipes extracts were estimated as 41.0 ± 1.3, 35.8 ± 1.3, and 25.0 ± 1.2 mg diosgenin/g extract, respectively. The outcome of this study suggested the exploitation of methanolic extract of P. vera, Z. amole, and E. crassipes leaves for their possible application in ethnomedicine, particularly as drugs preparation against staphylococcal infections. In conclusion, the study indicates the biopotency of these plants against pathogenic MRSA present in cattle, and SOSA as well as CoNS bacteria present in rabbits, which could be a serious issue for livestock.