Methyl Jasmonate and Salicylic Acid Enhanced the Production of Ursolic and Oleanolic Acid in Callus Cultures of Lepechinia Caulescens.

BACKGROUND: The production of triterpenes from plants for pharmacological purposes varies in concentration, due to genetic and environmental factors. In vitro culture enables the control and increase of these bioactive molecules. OBJECTIVE: To evaluate the effect of plant growth regulators and elicitors in the induction of calli and the production of ursolic acid (UA) and oleanolic acid (OA) in Lepechinia caulescens. MATERIALS AND METHODS: Leaf explants were exposed for the induction of calli at different concentrations and combinations of 2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzylaminopurine (BAP). Methyl jasmonate (MJ) and salicylic acid were used as elicitors. High-performance liquid chromatography method was used to quantify UA and OA content in each treatment. RESULTS: Treatment with 3.0 mg/L of 2,4-D and 0.1 mg/L of BAP produced the best results for calli induction and production of UA (1.57 mg/g dry weight [DW]) and OA (1.13 mg/g DW). Both elicitors facilitated the accumulation of triterpenes. CONCLUSION: The combination of auxins and cytokinins showed favorable results for the induction of calli. Variation concerning the accumulation of UA and OA was observed between treatments. MJ increased the production of triterpenes five times after 8 h of exposure, compared to control treatment. There is a greater accumulation of UA (16.58 mg/g DW) and OA (1.94 mg/g DW) in leaves of wild plants. SUMMARY: Callus cultures of Lepechinia caulescens were obtained from leaf explants treated with 2,4-dichlorophenoxyacetic acid and 6-bencylaminopurineResulting cultures were elicited with methyl jasmonate (MJ) and salicylic acid to increase the production of the triterpenes, ursolic acid (UA), and oleanolic acid (OA)The cultures elicited with MJ increased the production of UA and OA five times, as compared to the control. Abbreviations used: 2,4-D: 2,4-dichlorophenoxyacetic acid, BAP: 6-benzylaminopurine, DW: Dry weight, MJ: Methyl jasmonate, OA: Oleanolic acid, PGRs: Plant growth regulators, UA: Ursolic acid, SA: Salicylic acid.

Chemical and anti-ulcer evaluation of Jodina rhombifolia (Hook. & Arn.) Reissek extracts / Estudo químico e da atividade antiúlcera de Jodina rhombifolia (Hook. & Arn.) Reissek

Jodina rhombifolia (Hook. & Arn.) Reissek (Santalaceae) is a medicinal plant popularly used as an anti-ulcer medicine. The plant native from Southern Brazil was chemically investigated and tested for its in vivo gastric anti-ulcer property by chloride acid/ethanol model. The chromatographic analysis of the hydroethanol extract of its leaves revealed the presence of C-glycosylflavonoids. From the n-butanol fraction of the hydroethanol extract of its aerial parts, vicenin-2 was isolated as the main component and identified by spectroscopic methods; and, a direct comparison with authentic samples was made. This fraction afforded three other C-glycosylflavonoids: vitexin, orientin and swertisin; all of them identified by direct comparison with authentic samples. We found that the oral administration of aqueous and hydroethanolic extracts led to a significant decrease in the ulcer index.

Jodina rhombifolia (Hook. & Arn.) Reissek (Santalaceae) é uma espécie vegetal nativa do Sul do Brasil e usada externamente na medicina popular para o tratamento de úlcera na pele. Neste trabalho foi estudada a atividade antiúlcera gástrica em ratos usando como indutor ácido clorídrico/etanol. Os extratos hidroetanólico e aquoso apresentaram atividade antiúlcera no modelo utilizado. A análise cromatográfica do extrato hidroetanólico das folhas indicou a presença de C-glicosilflavonóides. Da fração n-butanólica obtida por extração seqüencial do extrato hidroetanólico foi isolada e identificada por métodos espectroscópicos a vicenina-2 como substância majoritária. Desta fração também foram identificados por CCD bidimensional, com auxílio de substâncias de referência, outros três C-glicosídeos: vitexina, orientina e swertisina.

Metabolic profiling of the Mexican anxiolytic and sedative plant Galphimia glauca using nuclear magnetic resonance spectroscopy and multivariate data analysis.

Galphimia glauca is popularly employed in Mexico for the treatment of central nervous system disorders. Pharmacological and phytochemical studies have resulted in the identification of the anxiolytic and sedative principle consisting of a mixture of nor-secofriedelanes, named the galphimine series (1 - 9). These active constituents were found in plants collected in the vicinity of a restricted region in Central Mexico, where this species is abundant. A metabolic profiling carried out by means of 1H-NMR spectroscopy and multivariate data analysis was applied to crude extracts from wild plant populations, collected from six different locations as a quality control assessment, in order to differentiate their chemical profile. Principal component analysis (PCA) of the 1H-NMR spectra revealed clear variations among the populations, with two populations out of the six studied manifesting differences, when the principal components PC-1 and PC-2 were analyzed. These two PCs permitted the differentiation of the various sample populations, depending on the presence of galphimines. This information consistently correlated with the corresponding HPLC analysis. The neuropharmacological effects of the crude extracts were evaluated by using ICR mice in the elevated plus maze, as well as the sodium pentobarbital-induced hypnosis models. Both assays demonstrated anxiolytic and sedative responses only among those sample populations which had previously been differentiated by PC-1. Partial least square regression-discriminant analysis (PLS-DA) also confirmed a strong correlation between the observed effects and the metabolic profiles of the plants. The overall results of this study confirm the benefits of using metabolic profiling for the in silico analysis of active principles in medicinal plants.

Isolation of nor-secofriedelanes from the sedative extracts of Galphimia glauca.

Preparative-scale recycling HPLC was used for the complete resolution of a complex mixture of nor-secofriedelanes into five major peaks (I-V) from the sedative methanolic extracts prepared from the aerial parts of Galphimia glauca. Argentation chromatography was used to show peaks I, II, IV, and V to be mixtures of isomers around the E-ring double bond, represented by the endocyclic C-20, C-21 double-bond isomers, galphimines A (3), B (1), D (4), and E (2), and the C-20, C-29 exocyclic forms, galphimines F-I (5-8). Galphimine C (9), isolated from peak III, corresponded to the C-19, C-20 double-bond isomer of the previously known major sedative constituent galphimine B. The characterization of all the new triterpenes (3-9) was performed primarily by high-field NMR spectroscopy. Comparison between experimental and calculated (1)H-(1)H vicinal coupling constants and the analysis of molecular mechanics structures revealed that the ring B of these compounds exists in a boatlike conformation. The absolute configuration for the stereogenic carbinol center at C-4 was established by the application of the Mosher ester derivatization technique carried out in NMR tubes.

Genetic transformation of Glaphimia glauca by Agrobacterium rhizogenes and the production of norfriedelanes.

Transformed root cultures of Galphimia glauca (Malpighiaceae) were established by infecting cotyledons and hypocotyls with Agrobacterium rhizogenes ATCC 15 834. Cotyledon-derived cell lines were grown in liquid B5 nutrient medium without phytohormones and have shown the typical hairy roots phenotype over two years of continuous subculturing. PCR analysis was used to confirm the integration of rol A and rol C genes into the plant genome. The transformed cultures synthesized three major norfriedelanes, the new glaucacetalins A-C (1-3), which were secreted into the nutrient medium. The structural elucidation of these in vitro produced metabolites was performed by the application of high resolution NMR techniques that proved them to be triterpenoids related to the known galphimines, the sedative principles of this plant species. These results suggest the possibility of further biotechnological exploration of sedative friedelane biosynthesis by in vitro plant organ cultures.