Takeshi Kodama

Isopimarane diterpenoids from Kaempferia pulchra rhizomes collected in Myanmar and their Vpr inhibitory activity

Viral protein R (Vpr), an accessory gene of HIV-1, plays important roles in viral pathogenesis. Screening of Myanmar medicinal plants that are popular as primary treatments for HIV/AIDS and for HIV-related problems revealed the potent anti-Vpr activity of the CHCl3-soluble extract of Kaempferia pulchra rhizomes, in comparison with that of the positive control, damnacanthal. Fractionation of the active CHCl3-soluble extract led to the identification of 30 isopimarane diterpenoids, including kaempulchraols A-W (1-23). All isolates were assayed for anti-Vpr activity against TREx-HeLa-Vpr cells, in which Vpr expression is tightly regulated by tetracycline. Kaempulchraols B (2), D (4), G (7), Q (17), T (20), U (21), and W (23) exhibited potent anti-Vpr activity, at concentrations ranging from 1.56 to 6.25μM. The structure-activity relationships of the active kaempulchraols suggested that the presence of a hydroxy group at C-14 in an isopimara-8(9),15-diene skeleton and the presence of an acetoxy group at C-1 or C-7 in an isopimara-8(14),15-diene skeleton are the critical factors for the inhibitory effects against TREx-HeLa-Vpr cells.

Kaempulchraols P-T, Diterpenoids from Kaempferia pulchra Rhizomes Collected in Myanmar.

The isolation of the oily fraction obtained from the CHCl3-soluble extract of the rhizomes of Kaempferia pulchra afforded five new isopimarane diterpenoids, kaempulchraols P-T (1-5), along with two known analogues. The structures were elucidated using spectroscopic techniques, including 2D NMR spectroscopy.

Structural basis for olivetolic acid formation by a polyketide cyclase from Cannabis sativa

In polyketide biosynthesis, ring formation is one of the key diversification steps. Olivetolic acid cyclase (OAC) from Cannabis sativa, involved in cannabinoid biosynthesis, is the only known plant polyketide cyclase. In addition, it is the only functionally characterized plant α+β barrel (DABB) protein that catalyzes the C2–C7 aldol cyclization of the linear pentyl tetra‐β‐ketide CoA as the substrate, to generate olivetolic acid (OA). Herein, we solved the OAC apo and OAC–OA complex binary crystal structures at 1.32 and 1.70 Å resolutions, respectively. The crystal structures revealed that the enzyme indeed belongs to the DABB superfamily, as previously proposed, and possesses a unique active‐site cavity containing the pentyl‐binding hydrophobic pocket and the polyketide binding site, which have never been observed among the functionally and structurally characterized bacterial polyketide cyclases. Furthermore, site‐directed mutagenesis studies indicated that Tyr72 and His78 function as acid/base catalysts at the catalytic center. Structural and/or functional studies of OAC suggested that the enzyme lacks thioesterase and aromatase activities. These observations demonstrated that OAC employs unique catalytic machinery utilizing acid/base catalytic chemistry for the formation of the precursor of OA. The structural and functional insights obtained in this work thus provide the foundation for analyses of the plant polyketide cyclases that will be discovered in the future.

New cytotoxic phloroglucinols, baeckenones D–F, from the leaves of Indonesian Baeckea frutescens

Three new phloroglucinols, baeckenones D-F (1-3), were obtained from the leaves of Indonesian Baeckea frutescens, along with the known unusual endoperoxide, phloroglucinol (4). The structures of the isolated compounds were elucidated by 1D and 2D NMR and HREIMS spectra. Furthermore, the stereochemistry of baeckenone D (1) was established by an X-ray diffraction analysis. Among the isolated compounds 1-4, baeckenone F (3) showed moderate cytotoxic activities against human pancreatic (PSN-1), lung (A549), and breast (MDA-MB-231) cancer cell lines, with IC50 values of 33.3 μM, 34 μM, and 39.3 μM, respectively.

New sesquiterpene lactones, vernonilides A and B, from the seeds of Vernonia anthelmintica in Uyghur and their antiproliferative activities.

A new guaianolide sesquiterpene lactone, vernonilide A (1), and a new elemanolide sesquiterpene lactone, vernonilide B (2), were isolated from the seeds of Vernonia anthelmintica, together with three known elemanolide sesquiterpene lactones (3-5). The structures of the isolated compounds were elucidated on the basis of physicochemical evidences. Compounds 1, 3, and 4 showed strong antiproliferative activities against three human cancer cell lines (A549, HeLa, and MDA-MB-231), with IC50 values ranging from 0.10 to 1.00μM. In addition, 5 exhibited significant antiproliferative activities against HeLa and MDA-MB-231 cells, with IC50 values ranging from 1.90 to 2.20μM. The antiproliferative activities of the acetyl derivatives 6 and 7 prepared from 4 and 3, respectively, against the three cell lines were 4-10-fold weaker than the original activities.

Picrajavanicins A–G, Quassinoids from Picrasma javanica Collected in Myanmar

Seven new tetracyclic quassinoids, picrajavanicins A-G (1-7), along with three known analogues, were isolated from a CHCl3-soluble extract of the bark of Picrasma javanica collected in Myanmar. The structures of these compounds were elucidated using spectroscopic techniques, including 1D and 2D NMR. The absolute configuration at C-2 of 2 was determined to be S by the modified Mosher method. All the isolates were tested for their antiproliferative activities against a small panel of five human cancer cell lines. However, none of the isolated compounds exhibited inhibitory activity against any of the cancer cells used (IC50 values >10 μM).

Quassinoids: Viral protein R inhibitors from Picrasma javanica bark collected in Myanmar for HIV infection

Viral protein R (Vpr) is an accessory protein that plays important roles in the viral pathogenesis of Human Immunodeficiency Virus-1 (HIV-1). An assay for anti-Vpr activity, using TREx-HeLa-Vpr cells, is a promising strategy to discover Vpr inhibitors. The anti-Vpr assay revealed that the CHCl3-soluble extract of Picrasma javanica bark possesses potent anti-Vpr activity. Furthermore, studies of quassinoids (1-15) previously isolated from the extract demonstrated that all of the tested quassinoids exhibit anti-Vpr activity. Among the tested compounds, javanicin I (15) exhibited the most potent anti-Vpr activity ((***)p <0.001) in comparing with that of the positive control, damnacanthal. The structure-activity relationships of the active quassinoids suggested that the presence of a methyl group at C-13 in the 2,12,14-triene-1,11,16-trione-2,12-dimethoxy-18-norpicrasane quassinoids is the important factor for the potent inhibitory effect in TREx-HeLa-Vpr cells.

Two new cyclopentenones and a new furanone from Baeckea frutescens and their cytotoxicities

Two new cyclopentenones, frutescencenones A (1) and B (2), and a new furanone derivative, frutescencenone C (3), together with two known cyclopentenones (4 and 5), were isolated from the leaves of Baeckea frutescens. Their structures were deduced by comprehensive spectroscopic analyses, including 1D and 2D NMR, and HREIMS data. Frutescencenone A (1) showed moderate growth inhibitory activity against human lung A549, pancreatic PSN-1, and breast MDA-MB-231 cancer cell lines, with IC50 values of 36.3μM, 38.2μM, and 29.3μM, respectively. In contrast, frutescencenone C (3) showed selective cytotoxic activity against PSN-1, with an IC50 value of 20.1μM.

Identification and Characterization of Daurichromenic Acid Synthase Active in Anti-HIV Biosynthesis

Daurichromenic acid, an anti-HIV meroterpenoid, is biosynthesized by a novel flavoprotein oxidase localized in the specialized epidermal organ, the glandular scales of Rhododendron dauricum. Daurichromenic acid (DCA) synthase catalyzes the oxidative cyclization of grifolic acid to produce DCA, an anti-HIV meroterpenoid isolated from Rhododendron dauricum. We identified a novel cDNA encoding DCA synthase by transcriptome-based screening from young leaves of R. dauricum. The gene coded for a 533-amino acid polypeptide with moderate homologies to flavin adenine dinucleotide oxidases from other plants. The primary structure contained an amino-terminal signal peptide and conserved amino acid residues to form bicovalent linkage to the flavin adenine dinucleotide isoalloxazine ring at histidine-112 and cysteine-175. In addition, the recombinant DCA synthase, purified from the culture supernatant of transgenic Pichia pastoris, exhibited structural and functional properties as a flavoprotein. The reaction mechanism of DCA synthase characterized herein partly shares a similarity with those of cannabinoid synthases from Cannabis sativa, whereas DCA synthase catalyzes a novel cyclization reaction of the farnesyl moiety of a meroterpenoid natural product of plant origin. Moreover, in this study, we present evidence that DCA is biosynthesized and accumulated specifically in the glandular scales, on the surface of R. dauricum plants, based on various analytical studies at the chemical, biochemical, and molecular levels. The extracellular localization of DCA also was confirmed by a confocal microscopic analysis of its autofluorescence. These data highlight the unique feature of DCA: the final step of biosynthesis is completed in apoplastic space, and it is highly accumulated outside the scale cells.

Antibacterial activities of chemical constituents from the aerial parts of Hedyotis pilulifera

Abstract Context: Hedyotis pilulifera (Pit.) T.N. Ninh (Rubiaceae) has been used in Vietnamese ethnomedicine; the methanol extract exhibited antibacterial activity in our preliminary screening. Objectives: In this study, compounds from H. pilulifera were isolated and their antibacterial activity in vitro was evaluated. Materials and methods: The aerial parts of H. pilulifera (1.4 kg) were extracted with MeOH, suspended in water and ethyl acetate extract was chromatographed on a silica gel column. The structures of isolated compounds were elucidated by the combination analyses of spectroscopy including 1D-, 2D-NMR, HRMS and in comparison with the reported NMR data in the literature. All isolated compounds were evaluated for inhibitory effect using the microdilution method toward Staphylococcus aureus, Bacillus subtilis and Mycobacterium smegmatis, and MIC values were determined. Results: Twenty compounds were isolated, including five triterpenoids, two steroids, two aromatic compounds, three fatty acids, one quinone derivative, one lignan glycoside, one ceramide and five glycolipids. Among these, oleanolic acid showed significant antibacterial activity against M. smegmatis with the MIC value of 2.5 μg/mL. Remarkably, rotungenic acid showed strong activity against S. aureus, B. subtilis, M. smegmatis with MIC values of 2.5, 2.5 and 1.25 μg/mL, respectively. Rotundic acid exhibited significant antibacterial activity against B. subtilis with the MIC value of 5 μg/mL. To the best of our knowledge, the antibacterial activity of rotungenic acid, stigmast-4-ene-3,6-dione and (2S,3S,4R,2′R)-2-(2′-hydroxytetracosanoylamino) octadecane-1,3,4-triol was reported for the first time. Conclusions: Oleanolic acid, rotungenic acid, and rotundic acid were considered to be useful for developing new antimicrobial therapeutic agents for human.