An extraction method using 80% EtOH was selected and applied to obtain the total extracts from leaves, flowers, fruits, twigs, and roots of Vitex rotundifolia L.f. based on the antioxidant activity-guided experiments. Subsequently, total extract from each part of V. rotundifolia was successfully partitioned into fractions, which were evaluated for their antioxidant and anti-inflammatory properties via DPPH, ABTS, and NO assays, respectively. Among them, EtOAc (E) and n-butanol (B) fractions showed the potent antioxidant activity and the methylene chloride (MC) fractions of roots, leaves, and fruits that exhibited strong scavenging activity on DPPH and ABTS radicals. In the anti-inflammatory assay, n-hexane (H) and MC fractions of leaves potently inhibited NO production in LPS-stimulated RAW264.7 cells, followed by E fractions derived from fruits, flowers, twigs, and roots, along with B fractions from flowers and twigs. Additionally, a comprehensive HPLC-decoupled MS profiling was established and validated using seven isolated marker compounds (1–7), which were identified by analysis of their UV, NMR, and MS data. The established method was also applied for quantification of these marker compounds in each organ collected from different locations, and to assess their antioxidant capacity by a screening DPPH-HPLC method. Principal component analysis suggested the botanical organs from this plant correlated with the marker compound contents in association with bioactivity. The study results are a prelude to further studies involving the active fractions and provide a comprehensive insight into the functional products of this plant against oxidative diseases.
Species of Podocarpus are used traditionally in their native areas for the treatment of fevers, asthma, coughs, cholera, chest pain, arthritis, rheumatism, and sexually transmitted diseases. To identify natural products having efficacy against inflammatory bowel disease (IBD), we identified a new, 16-hydroxy-4β-carboxy-O-β-D-glucopyranosyl-19-nor-totarol (4) together with three known diterpenoids from P. macrophyllus. Furthermore, all the extracts, fractions, and isolates 1-4 were investigated for their anti-inflammatory effects by assessing the expression on nitric oxide (NO) production and proinflammatory cytokines in lipopolysaccharide (LPS)-stimulated RAW 264.7 and HT-29 cells. Among them, nagilactone B (2) exhibited a potent anti-inflammatory effect against NO production on RAW 264.7 cells; therefore, nagilactone B was further assessed for anti-inflammatory activity. Western blot analysis revealed that nagilactone B significantly decreased the expression of LPS-stimulated protein, inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, and phosphorylated extracellular regulated kinase (pERK)1/2. In addition, nagilactone B downregulated tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-8 levels in LPS-induced macrophages and colonic epithelial cells. To our best knowledge, this is the first report on the inhibitory effect of nagilactone B (pure state) and rakanmakilactone G against NO production in LPS-stimulated RAW 264.7 cells. Thus, diterpenoids isolated from P. macrophyllus could be employed as potential therapeutic phytochemicals for IBD.
Ilex rotunda Thunb., has been used to treat common cold, tonsillitis, and eczema. It is also a source of antioxidants. However, information regarding its antioxidative phytochemical composition is still incomplete and limited. In this present study, we initially determined DPPH radical scavenging activity of the extracts of I. rotunda fruits, twigs, and leaves. Among them, the twig extract exhibited a potential of antioxidant capacity. Based on antioxidant effect guided experiments, extraction condition using 80% EtOH was then optimized. DPPH and ABTS radical scavenging assays were also performed for fractions. The n-butanol fraction showed the highest antioxidant effect. Using chromatographic methods, eight marker compounds (1–8) were further isolated. Their structures were determined by spectroscopic and mass data. Method validation was employed to quantitate contents of these eight marker compounds. Subsequently, the HPLC-DPPH method was used to evaluate the contribution of certain compounds to total antioxidant activity of the extract. Lastly, parallel artificial membrane permeability assay for blood–brain barrier (PAMPA-BBB) was applied to investigate brain-penetrable antioxidants from I. rotunda extract. As a result, compound 7 (4,5-dicaffeoylquinic acid) showed significant antioxidant activity and penetration across the BBB via transcellular passive diffusion. Our findings suggested that compound 7 can be used as a therapeutic potential candidate in natural product-based central nervous system (CNS) drug discovery.
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