Ulapualides C–E Isolated from a Hawaiian <i>Hexabranchus sanguineus</i> Egg Mass

Three new ulapualides (<b>3</b>–<b>5</b>) were isolated from egg masses of the nudibranch <i>Hexabranchus sanguineus</i>. The structures of <b>3</b>–<b>5</b> were deduced by analyses of physical and spectroscopic data in comparisons with ulapualides A (<b>1</b>) and B (<b>2</b>). Ulapualide C demonstrated submicromolar cytotoxicity against select NCI cell lines (768-0, DU-145, MDA-MB-231, and A549) with the most potent activity against MDA-MB-231 cells (IC₅₀ 0.58 μM). Ulapualides A (<b>1</b>) and B (<b>2</b>) were 2- to 4-fold more potent than <b>3</b>

Molecular structures of tricin and 7-O-methyl-tricin (7 MT), two compounds isolated from BEX.

<p>Molecular structures of tricin and 7-O-methyl-tricin (7 MT), two compounds isolated from BEX.</p

Effect of BEX on lipotoxic MCP-1 production.

<p>Production of MCP-1 was measured in the cell media supernatant (A–C) and cytosolic fraction (D–F) of 3T3-L1 (A, D), C2C12 (B, E), and Hepa6 (C, F) murine cells treated with palmitic acid (PA, 0.4 mM) in combination with BEX (125 µg/ml or 0.5%, v/v) or ethanol vehicle (0.5%, v/v). Concentrations were determined by cytometric bead array immunodetection against a reconstituted MCP-1 standard. Mean ± standard error over 3 trials, n ≥300 beads in any trial. Mean results were compared between treatments of the same cell type and at the same time point. Differences between means are statistically significant if columns do not share any common letters (p<0.05, one-way ANOVA with Bonferroni’s post hoc test).</p

Effect of BEX on MCP-1 mRNA expression.

<p>Levels of MCP-1 mRNA expression in 3T3-L1 (A), C2C12 (B), and Hepa6 (C) murine cells in response to palmitic acid (PA, 0.4 mM) in combination with BEX (125 µg/ml or 0.5%, v/v) or ethanol vehicle (0.5%, v/v) at different time points and differentiation states (3T3-L1 and C2C12). Fold change calculated via comparative cycle threshold (−ΔΔCt) normalized to beta-glucuronidase (GUSβ). Other housekeeping genes such as 18 S ribosomal RNA (18 S), hypoxanthine-guanine phosphoribosyltransferase (HPRT), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were also used and yielded similar results as GUSβ. Mean ± standard deviation of at least 4 qRT-PCRs per cell type. Mean results were compared between treatments of the same cell type and at the same time point. Differences between means are statistically significant if columns do not share any common letters (p<0.05, one-way ANOVA with Tukey’s multiple comparison test).</p

Dose-dependent effects of tricin, 7-O-methyl-tricin (7 MT), Fraction K, and BEX on lipotoxic MCP-1 production and cell viability.

<p>(A) MCP-1 concentrations in cell culture media from 3T3-L1 cells cultured in 96 well plates and treated for 20 hours with PA (0.4 mM) and one of the following: tricin (black triangles), 7 MT (light grey hollow squares), Fraction K (grey diamonds), or BEX (dark grey crosses). Concentrations were determined by cytometric bead array immunodetection against a reconstituted MCP-1 standard. Mean ± standard error of 3 samples per dose, n ≥300 beads per sample. (B) Viability of 3T3-L1 cells treated with tricin (black triangles), 7 MT (light grey hollow squares), Fraction K (grey diamonds), or BEX (dark grey crosses) added to maintenance medium. All values are normalized to 3T3-L1 cells from the same plate treated with maintenance medium without flavonoids or PA (normalized average represented by a red dashed line). Mean ± standard error of three trials, n ≥3 per trial. (C) Viability of 3T3-L1 cells treated with ethanol vehicle (outlined white), Fraction K (grey), or BEX (black). Mean ± standard error of three trials, n ≥4 per trial. All values normalized to 3T3-L1 cells from the same plate treated with only maintenance medium (normalized average represented by a red dashed line). <i>p</i> values were obtained by performing a Student’s <i>t</i>-test between doses.</p

Protective effect of BEX on palmitic acid (PA)-induced lipotoxicity.

<p>3T3-L1 (A), and Hepa6 cells (B) were grown to confluency in 96-well plates and treated with 0.4 mM PA for 24 hours in combination with BEX (125 µg/ml or 0.5%, v/v) or ethanol (0.5%, v/v) as a solvent control. Cell viability was measured using a MTS assay. Mean ± standard deviation over 5 trials, n ≥3 wells per trial. Differences between means are statistically significant if the columns do not share any common letters (p<0.001, one-way ANOVA, Tukey’s multiple comparison test).</p

Clustalw alignment for phylogenetic analysis of HT-58-2Cyano

Phylip formatted output of ClustalW aligned 16S rRNA of HT-58-2Cyano and other cyanobacteria. Supporting data for the manuscript AEM01068-17 submitted to Applied and Environmental Microbiology

ClustalW alignment for phylogenetic analysis of HT-58-2Cyano

Philip formatted output of ClustalW aligned 16S rRNA of HT-58-2Cyano and other cyanobacteria. Supporting data for the manuscript AEM01068-17 submitted to Applied and Environmental Microbiology

Neopetrocyclamines A and B, Polycyclic Diamine Alkaloids from the Sponge <i>Neopetrosia</i> cf <i>exigua</i>

Two new polycyclic alkaloids, neopetrocyclamines A and B (<b>1</b> and <b>2</b>), along with the known metabolites papuamine (<b>3</b>) and haliclonadiamine (<b>4</b>), were isolated from the Indonesian sponge <i>Neopetrosia</i> cf <i>exigua</i>. Neopetrocyclamine A contains a formamidinium moiety, a rare functional group. While these compounds share the same basic biosynthetic building blocks, the size of the ring system differs in <b>1</b> and <b>2</b> because of the formamidinium moiety. Biological evaluations of <b>1</b>–<b>4</b> revealed that papuamine is cytotoxic against glioblastoma SF-295 cells (GI₅₀ = 0.8 μM)

Spongiapyridine and Related Spongians Isolated from an Indonesian <i>Spongia</i> sp.

New compounds 18-nor-3,17-dihydroxyspongia-3,13(16),14-trien-2-one (<b>1</b>), 18-nor-3,5,17-trihydroxyspongia-3,13(16),14-trien-2-one (<b>2</b>), and spongiapyridine (<b>3</b>) and the known compound 17-hydroxy-4-<i>epi</i>-spongialactone A (<b>4</b>) were isolated from an Indonesian sponge of the genus <i>Spongia</i>. The structures of <b>1</b>–<b>3</b> were deduced by analyses of physical and spectroscopic data. Diterpene <b>3</b> is unusual, as the D-ring is a pyridyl ring system rather than the standard δ-lactone. The structure elucidation of this compound was complicated by facile exchange of the axial proton at the C-11 methylene with deuterium from methanol-<i>d</i>₄. The isolated compounds were tested for biological activity in a battery of in vitro assays (TNF-α-induced NFκB, LPS-induced iNOS, RXR stimulation, quinone reductase 1 induction, aromatase inhibition, TRPM7 ion channels, and aspartic protease BACE1 inhibition). Norditerpene <b>2</b> modestly inhibited aromatase with an IC₅₀ of 34 μM and induced quinone reductase 1 activity with a CD (the concentration needed to double the enzymatic response) of 11.2 μM. The remaining isolates were inactive