Cyclic enaminones and a 4-quinazolinone from an unidentified actinomycete of the family Micromonosporaceae.

Four novel cyclic enaminones, designated RD4123A-D (1-4), and a new 4-quinazolinone metabolite, RD4123E (5), were isolated from the culture extract of an unidentified actinomycete strain RD004123, which belongs to the family Micromonosporaceae. Structures of 1-5 were determined by spectroscopic analyses using NMR, MS, and electronic circular dichroism (ECD), combined with quantum chemical calculations of ECD and NMR chemical shifts and biosynthetic consideration. Compounds 1-5 showed weak to modest cytotoxicity against murine leukemia P388 cells, while being inactive against bacteria and fungi.

Pithohirolide, an antimicrobial tetradepsipeptide from a fungus Pithomyces chartarum.

Pithohirolide (1), a new depsipeptide, was isolated from an ascomycetous fungus Pithomyces chartarum TAMA 581. The planar structure of 1 was elucidated on the basis of NMR and MS analyses and the absolute configuration was determined by the advanced Marfey's analysis, chiral-phase HPLC analysis, and synthesis of degradation product. Compound 1 possesses a cyclic structure comprising (S)-2-hydroxy-3-phenylpropanoic acid, (S)-3-hydroxy-3-phenylpropanoic acid, (S)-2-hydroxyisovaleric acid, and N-methyl-L-alanine, connected via three ester and one amide linkages. Compound 1 exhibited antimicrobial activity against Staphylococcus aureus and Saccharomyces cerevisiae at MIC 3.1 µg ml-1.

TMKS8A, an antibacterial and cytotoxic chlorinated α-lapachone, from a sea slug-derived actinomycete of the genus Streptomyces.

TMKS8A (1), a new chlorinated α-lapachone derivative, along with five known related metabolites, A80915 C (2), SF2415B1 (3), chlorinated dihydroquinone 3 (4), SF2415B3 (5), and A80915 C (6), were identified from the culture extract of Streptomyces sp. TMKS8, which was isolated from a sea slug, Paromoionchis tumidus. The structure of 1 was determined by the analysis of NMR and MS spectral data, assisted by NMR chemical shift prediction using DFT-based calculation. The absolute configuration was determined to be R by comparison of experimental and calculated ECD spectra. Compound 1 displayed antimicrobial activity against Gram-positive bacteria with MIC values ranging from 6.25 to 12.5 µg ml-1 and cytotoxicity against murine leukemia P388 cells with IC50 9.8 µM.

Potent and broad anticancer activities of leaf extracts from Melia azedarach L. of the subtropical Okinawa islands.

Plant extracts have been traditionally used for various therapeutic applications. By conducting an initial screening of several subtropical plants, in this study, we evaluated the anticancer activities of Melia azedarach L. The extract from Melia azedarach L. leaves (MLE) show high cytotoxic effects on cancer cells and in vivo mouse and dog tumor models. During the initial screening, MLE showed strong antiproliferative activity against HT-29 colon, A549 lung, and MKN1 gastric cancer cells. In subsequent tests, using 39 human tumor cell lines, we confirmed the potent anticancer activities of MLE. The anticancer activity of MLE was also confirmed in vivo. MLE markedly inhibited the growth of transplanted gastric MKN1 cancer xenografts in mice. To elucidate the mechanism underlying the anticancer effects of MLE, MLE-treated MKN1 cells were observed using an electron microscope; MLE treatment induced autophagy. Furthermore, western blot analysis of proteins in lysates of MLE-treated cells revealed induction of light chain 3 (LC3)-II autophagosomal proteins. Thus, MLE appeared to suppress MKN1 cell proliferation by inducing autophagy. In addition, in the mouse macrophage cell line J774A.1, MLE treatment induced TNF-α production, which might play a role in tumor growth suppression in vivo. We also performed a preclinical evaluation of MLE treatment on dogs with various cancers in veterinary hospitals. Dogs with various types of cancers showed a mean recovery of 76% when treated with MLE. Finally, we tried to identify the active substances present in MLE. All the active fractions obtained by reverse-phase chromatography contained azedarachin B-related moieties, such as 3-deacetyl-12-hydroxy-amoorastatin, 12-hydroxy-amoorastatin, and 12-hydroxyamoorastaton. In conclusion, MLE contains substances with promising anticancer effects, suggesting their future use as safe and effective anticancer agents.

Functional growth inhibition of influenza A and B viruses by liquid and powder components of leaves from the subtropical plant Melia azedarach L.

We evaluated the anti-influenza-virus effects of Melia components and discuss the utility of these components. The effects of leaf components of Melia azedarach L. on viruses were examined, and plaque inhibition tests were performed. The in vivo efficacy of M. azedarach L. was tested in a mouse model. Leaf components of Melia azedarach L. markedly inhibited the growth of various influenza viruses. In an initial screening, multiplication and haemagglutination (HA) activities of H1N1, H3N2, H5, and B influenza viruses were inactivated by the liquid extract of leaves of M. azedarach L. (MLE). Furthermore, plaque inhibition titres of H1N1, H3N2, and B influenza viruses treated with MLE ranged from 103.7 to 104.2. MLE possessed high plaque-inhibitory activity against pandemic avian H5N1, H7N9, and H9N2 vaccine candidate strains, with a plaque inhibition titre of more than 104.2. Notably, the buoyant density decreased from 1.175 to 1.137 g/cm3, and spikeless particles appeared. We identified four anti-influenza virus substances: pheophorbide b, pheophorbide a, pyropheophorbide a, and pheophytin a. Photomorphogenesis inside the envelope may lead to removal of HA and neuraminidase spikes from viruses. Thus, MLE could efficiently remove floating influenza virus in the air space without toxicity. Consistent with this finding, intranasal administration of MLE in mice significantly decreased the occurrence of pneumonia. Additionally, leaf powder of Melia (MLP) inactivated influenza viruses and viruses in the intestines of chickens. MLE and MLP may have applications as novel, safe biological disinfectants for use in humans and poultry.

Bipolamides A and B, triene amides isolated from the endophytic fungus Bipolaris sp. MU34.

As a result of the continued screening for new metabolites produced by endophytic fungi from Thai medicinal plants, two new triene fatty acid amides, bipolamides A (1) and B (2), were discovered from the endophytic fungus Bipolaris sp. MU34. The structures of all of the isolated compounds were elucidated on the basis of the spectroscopic data of NMR and MS. An antimicrobial assay revealed that bipolamide B (2) had moderate antifungal activity against Cladosporium cladosporioides FERMS-9, Cladosporium cucumerinum NBRC 6370, Saccharomyces cerevisiae ATCC 9804, Aspergillus niger ATCC 6275 and Rhisopus oryzae ATCC 10404, with Minimum inhibitory concentration (MIC) values of 16, 32, 32, 64 and 64 µg ml(-1), respectively.

High-throughput screening of inhibitors targeting Agr/Fsr quorum sensing in Staphylococcus aureus and Enterococcus faecalis.

Staphylococcus aureus and Enterococcus faecalis employ cyclic peptide-mediated quorum sensing (QS) systems, termed agr and fsr respectively, to regulate the expression of a series of virulence genes. To identify quorum sensing inhibitors (QSIs) that target agr/fsr systems, an efficient screening system was established. In addition to the gelatinase-induction assay to examine E. faecalis fsr QS, the use of an S. aureus agr reporter strain that carries luciferase and green fluorescence protein genes under the agr P3 promoter facilitated the development of a high-throughput screen (HTS) for QSIs. As a result of screening of 906 actinomycetes culture extracts, four showed QSI activity against the agr and fsr systems without growth inhibitory activity. The extracts were purified on a small scale, and three HPLC peaks were obtained with obvious QSI activity. In sum, the established HTS system is a promising strategy for the discovery of anti-pathogenic agents targeting cyclic peptide-mediated QS in Gram-positive pathogens.

Brartemicin, an inhibitor of tumor cell invasion from the actinomycete Nonomuraea sp.

Brartemicin (1), a new trehalose-derived metabolite, was isolated from the culture broth of the actinomycete of the genus Nonomuraea. Its structure and absolute configuration were determined by spectroscopic analyses. The new compound inhibited the invasion of murine colon carcinoma 26-L5 cells with an IC(50) value of 0.39 microM in a concentration-dependent manner without showing cytotoxic effects.

Cremastrine, a pyrrolizidine alkaloid from Cremastra appendiculata.

A new pyrrolizidine alkaloid, cremastrine (1), was isolated from the bulbs of Cremastra appendiculata. Its configuration was determined by spectroscopic and chemical analyses. Compound 1 inhibited the binding of tritium-labeled N-methylscopolamine to the muscarinic M3 receptor with a K(i) value of 126 nM.