Diarylheptanoid Glycosides of Morella salicifolia Bark.

A methanolic extract of Morella salicifolia bark was fractionated by various chromatographic techniques yielding six previously unknown cyclic diarylheptanoids, namely, 7-hydroxymyricanol 5-O-ß-d-glucopyranoside (1), juglanin B 3-O-ß-d-glucopyranoside (2), 16-hydroxyjuglanin B 17-O-ß-d-glucopyranoside (3), myricanone 5-O-ß-d-gluco-pranosyl-(1→6)-ß-d-glucopyranoside (4), neomyricanone 5-O-ß-d-glucopranosyl-(1→6)-ß-d-glucopyranoside (5), and myricanone 17-O-α-l-arabino-furanosyl-(1→6)-ß-d-glucopyranoside (6), respectively, together with 10 known cyclic diarylheptanoids. The structural diversity of the diarylheptanoid pattern in M. salicifolia resulted from varying glycosidation at C-3, C-5, and C-17 as well as from substitution at C-11 with hydroxy, carbonyl or sulfate groups, respectively. Structure elucidation of the isolated compounds was achieved on the basis of one- and two-dimensional nuclear magnetic resonance (NMR) as well as high-resolution electrospray ionisation mass spectrometry (HR-ESI-MS) analyses. The absolute configuration of the glycosides was confirmed after hydrolysis and synthesis of O-(S)-methyl butyrated (SMB) sugar derivatives by comparison of their ¹H-NMR data with those of reference sugars. Additionally, absolute configuration of diarylheptanoid aglycones at C-11 was determined by electronic circular dichroism (ECD) spectra simulation and comparison with experimental CD spectra after hydrolysis.

Revisited anti-inflammatory activity of matricine in vitro: Comparison with chamazulene.

The proazulene matricine (1) is present in chamomile flower heads and has been proven to exhibit strong in vivo anti-inflammatory activity. In contrast to other secondary metabolites in chamomile preparations like its degradation product chamazulene (2), no plausible targets have been found to explain this activity. Therefore we revisited 1 regarding its in vitro anti-inflammatory activity in cellular and molecular studies. Using ICAM-1 as a marker for NF-κB activation, it was shown that ICAM-1 protein expression induced by TNF-α and LPS, but not by IFN-γ, was remarkably inhibited by 1 in endothelial cells (HMEC-1). Inhibition was concentration-dependent in a micromolar range (10-75 µM) and did not involve cytotoxic effects. At 75 µM expression of the adhesion molecule ICAM-1 was down to 52.7 ± 3.3% and 20.4 ± 1.8% of control in TNF-α and LPS-stimulated HMEC-1, respectively. In contrast, 2 showed no activity. Quantitative RT-PCR experiments revealed that TNF-α-induced expression of the ICAM-1 gene was also reduced by 1 in a concentration-dependent manner, reaching 32.3 ± 6.2% of control at 100 µM matricine. Additional functional assays (NF-κB promotor activity and cytoplasm to nucleus translocation) confirmed the inhibitory effect of 1 on NF-κB signaling. Despite the fact that 1 lacks an α,ß-unsaturated carbonyl and is thus not able to act via a Michael reaction with electron rich SH groups of functional biological molecules, data gave strong evidence that 1 inhibits NF-κB transcriptional activity in endothelial cells by an hitherto unknown mechanism and this may contribute to its well-known anti-inflammatory activity in vivo.

Biotransformation of Flavokawains A, B, and C, Chalcones from Kava (Piper methysticum), by Human Liver Microsomes.

The in vitro metabolism of flavokawains A, B, and C (FKA, FKB, FKC), methoxylated chalcones from Piper methysticum, was examined using human liver microsomes. Phase I metabolism and phase II metabolism (glucuronidation) as well as combined phase I+II metabolism were studied. For identification and structure elucidation of microsomal metabolites, LC-HRESIMS and NMR techniques were applied. Major phase I metabolites were generated by demethylation in position C-4 or C-4' and hydroxylation predominantly in position C-4, yielding FKC as phase I metabolite of FKA and FKB, helichrysetin as metabolite of FKA and FKC, and cardamonin as metabolite of FKC. To an even greater extent, flavokawains were metabolized in the presence of uridine diphosphate (UDP) glucuronic acid by microsomal UDP-glucuronosyl transferases. For all flavokawains, monoglucuronides (FKA-2'-O-glucuronide, FKB-2'-O-glucuronide, FKC-2'-O-glucuronide, FKC-4-O-glucuronide) were found as major phase II metabolites. The dominance of generated glucuronides suggests a role of conjugated chalcones as potential active compounds in vivo.

Aqueous extracts of the marine brown alga Lobophora variegata inhibit HIV-1 infection at the level of virus entry into cells.

In recent years, marine algae have emerged as a rich and promising source of molecules with potent activities against various human pathogens. The widely distributed brown alga Lobophora variegata that is often associated with tropical coral reefs exerts strong antibacterial and antiprotozoal effects, but so far has not been associated with specific anti-viral activities. This study investigated potential HIV-1 inhibitory activity of L. variegata collected from different geographical regions, using a cell-based full replication HIV-1 reporter assay. Aqueous L. variegata extracts showed strong inhibitory effects on several HIV-1 strains, including drug-resistant and primary HIV-1 isolates, and protected even primary cells (PBMC) from HIV-1-infection. Anti-viral potency was related to ecological factors and showed clear differences depending on light exposition or epiphyte growth. Assays addressing early events of the HIV-1 replication cycle indicated that L. variegata extracts inhibited entry of HIV-1 into cells at a pre-fusion step possibly by impeding mobility of virus particles. Further characterization of the aqueous extract demonstrated that even high doses had only moderate effects on viability of cultured and primary cells (PBMCs). Imaging-based techniques revealed extract effects on the plasma membrane and actin filaments as well as induction of apoptosis at concentrations exceeding EC50 of anti-HIV-1 activity by more than 400 fold. In summary, we show for the first time that L. variegata extracts inhibit HIV-1 entry, thereby suggesting this alga as promising source for the development of novel HIV-1 inhibitors.

Hyperforin is a modulator of inducible nitric oxide synthase and phagocytosis in microglia and macrophages.

Upon activation, microglia, the immunocompetent cells in the brain, get highly phagocytic and release pro-inflammatory mediators like nitric oxide (NO). Excessive NO production is pivotal in neurodegenerative disorders, and there is evidence that abnormalities in NO production and inflammatory responses may at least support a range of neuropsychiatric disorders, including depression. Although extracts of St. John's wort (Hypericum perforatum L.) have been used for centuries in traditional medicine, notably for the treatment of depression, there is still considerable lack in scientific knowledge about the impact on microglia. We used N11 and BV2 mouse microglia, as well as RAW 264.7 macrophages to investigate the effects of St. John's wort extract and constituents thereof on NO production Moreover, flow cytometry and fluorescence microscopy were employed to analyze the influence on phagocytosis, transcription factor activation states, and cell motility. We found that extracts of St. John's wort efficiently suppress lipopolysaccharide-induced NO release and identified hyperforin as the responsible compound, being effective at concentrations between 0.25 and 0.75 microM. The reduced NO production was mediated by diminished inducible nitric oxide synthase expression on the mRNA and protein level. In addition, at similar concentrations, hyperforin reduced zymosan phygocytosis to 20-40% and putatively acted by downregulating the CD206 macrophage mannose receptor and modulation of cell motility. We found that the observed effects correlate with a suppression of the activated state of Nf-kappaB and phospho-CREB, while c-JUN, STAT1, and HIF-1alpha activity and cyclooxygenase-2 expression remained unaffected by hyperforin. These results reveal that hyperforin influences pro-inflammatory and immunological responses of microglia that are involved in the progression of neuropathologic disorders.

Xanthohumol, a chalcon derived from hops, inhibits hepatic inflammation and fibrosis.

Xanthohumol (XN) is a major prenylated chalcone found in hops, which is used to add bitterness and flavor to beer. In this study, we first investigated the effects of XN on hepatocytes and hepatic stellate cells (HSC), the central mediators of liver fibrogenesis. XN inhibited the activation of primary human HSC and induced apoptosis in activated HSC in vitro in a dose dependent manner (0-20 microM). In contrast, XN doses as high as 50 microM did not impair viability of primary human hepatocytes. However, in both cell types XN inhibited activation of the transcription factor NFkappaB and expression of NFkappaB dependent proinflammatory genes. In vivo, feeding of XN reduced hepatic inflammation and expression of profibrogenic genes in a murine model of non-alcoholic steatohepatitis. These data indicate that XN has the potential as functional nutrient for the prevention or treatment of non-alcoholic steatohepatitis or other chronic liver disease.

Influence of Hypericum perforatum extract and its single compounds on amyloid-beta mediated toxicity in microglial cells.

As immunocompetent cells of the brain, microglia are able to counteract the damaging effects of amyloid-beta in Alzheimer's disease by phagocytosis-mediated clearance of protein aggregates. The survival and health of microglia are therefore critical for attenuating and preventing neurodegenerative diseases. In a microglial cell line pretreated with St. John's wort (Hypericum perforatum L.) extract (HPE), the cell death evoked by treatment with amyloid-beta (25-35) and (1-40) was attenuated significantly in a dose-dependent manner. Investigation of the single compounds in the extract revealed that the flavanols (+)-catechin and (-)-epicatechin increase cell viability slightly, whereas the flavonol quercetin and its glycosides rutin, hyperosid and quercitrin showed no effect on cell viability. In contrast, at the same concentration, the flavonoids reduced the formation of amyloid-induced reactive oxygen species in microglia, indicating that improvement of cell viability by the catechins is not correlated to the antioxidant activity. No influence of HPE on the capacity of microglia to phagocytose sub-toxic concentrations of fibrillar amyloid-beta (1-40) was observed. Other experiments showed that HPE, (+)-catechin and (-)-epicatechin can alter cellular membrane fluidity and thereby may have a beneficial effect on cell health. Our findings provide in vitro evidence that treatment especially with the complex plant extract HPE may restore or improve microglial viability and thereby attenuate amyloid-beta mediated toxicity in Alzheimer's disease.