Chemical characterization and antioxidant potential of ecuadorian propolis.

The chemical composition and the antioxidant potential of Ecuadorian propolis samples (n = 19) collected in different provinces were investigated. HPLC-DAD-ESI/MSn and GC-EI-MS analysis of the methanol extracts enabled us to define six types of Ecuadorian propolis based on their secondary metabolite composition. 68 compounds were identified, 59 of which are reported for the first time in Ecuadorian propolis. The detected compounds include flavonoids, diterpenes, triterpenes, organic acid derivatives, alkylresorcinol derivatives and nemorosone. Plants belonging to genera Populus, Mangifera and Clusia seemed to be vegetable sources employed by bees to produce Ecuadorian propolis. Total phenolic content and antioxidant activity of propolis extracts were determined by the Folin-Ciocalteu assay and 2,2-diphenyl-1-picrylhydrazyl and ferric reducing/antioxidant potential assays, respectively. As expected, the variable chemical composition affected the differences in terms of antioxidant potential.

Counter-current chromatography with off-line detection by ultra high performance liquid chromatography/high resolution mass spectrometry in the study of the phenolic profile of Lippia origanoides.

Lippia origanoides (Verbenaceae) is an important Brazilian medicinal plant, also used for culinary purposes. Most chemical studies with this plant have been focused on its volatile composition. In this work, we combined High-Speed Counter-current Chromatography (HSCCC) and High Performance Liquid Chromatography coupled to Ultra Violet detection and High Resolution Mass Spectrometry (HPLC-UV-HRMSn) methodologies to access the non-volatile chemical composition of L. origanoides. The crude ethanol extract of L. origanoides (LOEF) was first analyzed by HPLC-UV-HRMSn and allowed the identification of 7 major compounds. Among them, eriodictyol, naringenin and pinocembrin, were determined and are phytochemical markers of this plant. However, owing to the complexity of this plant matrix, LOEF was fractionated by HSCCC (hexane-ethanol-water, 4:3:1) as a tool for preparative pre-purification, affording a flavonoid-rich fraction. A column screening with the chromatographic stationary phases ZIC-HILIC, monolithic and particulate RP18 was performed. The best column separation was achieved with a Purospher STAR RP18e, which was used for HPLC-DAD-HRMSn studies. By this approach 12 compounds were further identified in addition to the major ones identified in the raw extract. Two of them, 6,8-di-C-hexosyl-luteolin and 6,8-di-C-glucosyl-apigenin, are being reported for the first time in the family Verbenaceae. This work shows the integration of HSCCC as a preparative tool for the fractionation and purification of natural products from a complex plant extract with other analytical techniques, with the purpose of showing each technique's potential.

Countercurrent chromatography separation of saponins by skeleton type from Ampelozizyphus amazonicus for off-line ultra-high-performance liquid chromatography/high resolution accurate mass spectrometry analysis and characterisation.

Ampelozizyphus amazonicus Ducke (Rhamnaceae), a medicinal plant used to prevent malaria, is a climbing shrub, native to the Amazonian region, with jujubogenin glycoside saponins as main compounds. The crude extract of this plant is too complex for any kind of structural identification, and HPLC separation was not sufficient to resolve this issue. Therefore, the aim of this work was to obtain saponin enriched fractions from the bark ethanol extract by countercurrent chromatography (CCC) for further isolation and identification/characterisation of the major saponins by HPLC and MS. The butanol extract was fractionated by CCC with hexane - ethyl acetate - butanol - ethanol - water (1:6:1:1:6; v/v) solvent system yielding 4 group fractions. The collected fractions were analysed by UHPLC-HRMS (ultra-high-performance liquid chromatography/high resolution accurate mass spectrometry) and MSn. Group 1 presented mainly oleane type saponins, and group 3 showed mainly jujubogenin glycosides, keto-dammarane type triterpene saponins and saponins with C31 skeleton. Thus, CCC separated saponins from the butanol-rich extract by skeleton type. A further purification of group 3 by CCC (ethyl acetate - ethanol - water (1:0.2:1; v/v)) and HPLC-RI was performed in order to obtain these unusual aglycones in pure form.

Spray-dried extract from the Amazonian adaptogenic plant Ampelozizyphus amazonicus Ducke (Saracura-mirá): Chemical composition and immunomodulatory properties.

Ampelozizyphus amazonicus Ducke is a medicinal plant used in the Amazon region to prepare a drink with tonic, immunomodulatory and adaptogenic properties. Due to the growing interest in dietary supplements with these properties and, to provide a new functional ingredient, barks from A. amazonicus were extracted. The extract was spray dried without drying adjuvants, resulting in a powder (SARF), which was characterized by its physico-chemical properties and proximate, mineral and saponin contents. The SARF saponins were characterized by ultra-high-performance liquid chromatography/high resolution accurate mass spectrometry (HPLC-HRMSn) analysis. The SARF particles tended to have a spherical shape and a unimodal size distribution. The particles also had good rehydration characteristics and high saponin content (33%). The effect of SARF on antibody production was investigated, and we found that SARF increased the basal levels of anti-ovalbumin, anti-LPS and anti-dextran IgM antibodies, and the anti-dextran IgG antibodies in unimmunized mice. No increase in antibody titers was observed after SARF treatment in immunized mice. These results suggest that SARF could be an interesting new functional ingredient for food applications or pharmaceutical products.