Antimicrobial activity in host-pathogen co-culture assay.

<p>The assay was based on <i>S. aureus</i> infected HL cell line. Dose-response results for a) ciprofloxacin) and b) 28-<i>O</i>-(<i>N</i>-acetylanthraniloyl)betulin (<b>5</b>).</p

Cytotoxic effect of selected betulins on mammalian cells (Huh-7) after 24 h exposure at different concentrations.

<p>A) betulinic aldehyde (<b>18</b>), B) betulin-28-oxime (<b>31</b>) and C) hetero cycloadduct (<b>43</b>).</p

Antimicrobial and cytotoxic effects of the most active betulin derivatives at 50 µM concentration (thresholds: antimicrobial activity >70%, cytotoxicity >50%).

<p>Values represent the mean ± SD of 3–6 replicates. Inhibitory effects of the most active samples are in bold. Primary screening results for all tested compounds are available in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102696#pone.0102696.s002" target="_blank">Tables S1</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102696#pone.0102696.s003" target="_blank">S2</a>.</p

Chemical structures of betulin and the most active betulin derivatives.

<p>Betulin (<b>1</b>), betulinyl 28-carboxymethoxycarvacrolate (<b>4</b>), 28-<i>O</i>-(<i>N</i>-acetylanthraniloyl)betulin (<b>5</b>), betulinic aldehyde (<b>18</b>), betulonic acid (<b>23</b>), betulin-28-oxime (<b>31</b>), and heterocyclic derivatives with 1,3-dioxol-5-yl (<b>35</b>), 3-nitrophenyl (<b>38</b>) and ethyl (<b>43</b>) attached to the nitrogen atom in the triazolo ring.</p

Suberin Fatty Acids from Outer Birch Bark: Isolation and Physical Material Characterization

The isolation and physical material properties of suberin fatty acids (SFAs) were investigated with special reference to their potential applications as novel pharmaceutical excipients. SFAs were isolated from outer birch bark (OBB) with a new extractive hydrolysis method. The present simplified isolation process resulted in a moderate batch yield and chemical purity of SFAs, but further development is needed for establishing batch-to-batch variation. Cryogenic milling was the method of choice for the particle size reduction of SFAs powder. The cryogenically milled SFAs powder exhibited a semicrystalline structure with apparent microcrystalline domains within an amorphous fatty acids matrix. The thermogravimetric analysis (TGA) of SFAs samples showed a good thermal stability up to 200 °C, followed by a progressive weight loss, reaching a plateau at about 95% volatilization at about 470 °C. The binary blends of SFAs and microcrystalline cellulose (MCC; Avicel PH 101) in a ratio of 25:75 (w/w) displayed good powder flow and tablet compression properties. The corresponding theophylline-containing tablets showed sustained or prolonged-release characteristics. The physicochemical and bulk powder properties of SFAs isolated from OBB are auspicious in terms of potential pharmaceutical excipient applications

Abietane-Type Diterpenoid Amides with Highly Potent and Selective Activity against <i>Leishmania donovani</i> and <i>Trypanosoma cruzi</i>

Dehydroabietylamine (<b>1</b>) was used as a starting material to synthesize a small library of dehydroabietyl amides by simple and facile methods, and their activities against two disease-causing trypanosomatids, namely, <i>Leishmania donovani</i> and <i>Trypanosoma cruzi,</i> were assayed. The most potent compound, <b>10</b>, an amide of dehydroabietylamine and acrylic acid, was found to be highly potent against these parasites, displaying an IC₅₀ value of 0.37 μM against <i>L. donovani</i> axenic amastigotes and an outstanding selectivity index of 63. Moreover, compound <b>10</b> fully inhibited the growth of intracellular amastigotes in <i>Leishmania donovani</i>-infected human macrophages with a low IC₅₀ value of 0.06 μM. This compound was also highly effective against <i>T. cruzi</i> amastigotes residing in L6 cells with an IC₅₀ value of 0.6 μM and high selectivity index of 58, being 3.5 times more potent than the reference compound benznidazole. The potent activity of this compound and its relatively low cytotoxicity make it attractive for further development in pursuit of better drugs for patients suffering from leishmaniasis and Chagas disease

Primary 3D screen.

<p>PC-3 cells were cultured in 3D Matrigel ECM for 4 days and treated for 6 days with 25 betulin derivatives (from 2D high throughput screens), DMSO/vehicle control, and three reference compounds. A) Representative maximum intensity projections of confocal microscope stack images for selected compound treatments at 300 nM concentration (5× objective, scale 100 μm). B) Three graphs showing the relative impact on three morphometric parameters for 7 betulin derivatives, DMSO control, and one control compound (paclitaxel). Data scaling: displays the relative difference between median of Area/Complexity/Area Ratio, to DMSO control. Paclitaxel treatments and DMSO controls have been assigned values of -100 and 0, respectively. C) Hierarchical clustering was done using three morphological parameters derived from PC3 organoids: spheroid size (area), complexity and the number of dead cells. D) Wound healing curves of the two betulin derivatives <b>4</b> and <b>20</b>, highlighting the 50% cut-off level (orange dashed line).</p

Collection of the features used to define the pharmacophoric model for the binding of the triterpenes to the ABHD12.

<p>Color coding of pharmacophoric features: green, hydrophobic site; magenta, acceptor site; cyan, donor site; gray cage, ligand shape constraint. <b>B</b>. The best solution for the alignment of the most active compound (<b>33</b>) to pharmacophore model (the same color coding as above except the ligand surface omitted for the clarity).</p

Inhibitor potency in hABHD12-HEK293 lysates for compounds 24–30 as well as their calculated lipophilicity values (logD).

<p>Data are mean ± SEM from three independent experiments.</p

Cytotoxicity tests performed in 2D monolayer culture.

<p><b>A</b>) Cell proliferation/cell number, <b>B</b>) programmed cell death (apoptosis), and <b>C</b>) number of dead cells were assessed by conventional assays, combined with high-content microscopy (Operetta). Cells were treated with the five most effective betulin derivatives, including paclitaxel control for 72h. Proliferation was measured as the total number of nuclei (= cells), apoptosis as ratio of caspase-3 positive versus all cells, and cell death as ethidium homodimer-2 positive nuclei versus all cells.</p