Anti-Vpr activities of sesqui- and diterpenoids from the roots and rhizomes of Kaempferia candida.

New copaene-type and nerolidol-type sesquiterpenoids, 7-hydroxymustakone (1) and 15-hydroxynerolidol (2), and a 15-norlabdane diterpenoid, kaempcandiol (3), together with four known compounds (4-7) were isolated from the chloroform extract of Kaempferia candida roots and rhizomes. The structures of the new compounds 1-3 were elucidated based on 1D and 2D NMR and HRESIMS spectroscopic analyses. The extract of the K. candida roots and rhizomes and all isolated compounds 1-7 possessed HIV-1 viral protein R (Vpr) inhibitory activities on the TREx-HeLa-Vpr cell line at a 5 µM concentration, without detectable cytotoxicity.

Shanpanootols A-F, diterpenoids from Kaempferia pulchra rhizomes collected in Myanmar and their Vpr inhibitory activities.

Six new isopimarane diterpenoids, shanpanootols A-F (1-6), along with two known analogues, were isolated from the ethyl acetate-soluble extract of Kaempferia pulchra rhizomes collected in Myanmar. The structures of these compounds were elucidated by extensive spectroscopic techniques such as 1D and 2D NMR and HRESIMS. The absolute configuration of 1 was determined by the modified Mosher method. The new isolates (1-6) were tested for their Vpr inhibitory activities against TREx-HeLa-Vpr cells. Shanpanootols C (3) and E (5) inhibited Vpr at doses of 2.5 and 5 µM, respectively.

Anti-Vpr activities of homodrimane sesquiterpenoids and labdane diterpenoids from Globba sherwoodiana rhizomes.

Two new homodrimane sesquiterpenoids, globbatones A and B (1 and 2), and one 16-norlabdane diterpenoid, globbatone C (3), together with two new naturally occurring, (E)-labda-8(17),12-diene-15,16-olide (4) and γ-bicyclohomofarnesen-12-ol (5), and one known homodrimane sesquiterpenoid (6), nine known labdane diterpenoids (7-15), and one isospongian diterpenoid (16), were isolated from the chloroform extract of Globba sherwoodiana rhizomes. The structures of the new compounds 1-3 were elucidated based on 1D and 2D NMR and HRESIMS spectroscopic analyses. The chloroform extract of G. sherwoodiana rhizomes and 10 µM concentrations of some of its constituents 1, 3, 4, 8, 9, 12, and 14 showed the moderate anti-Vpr activities, without cytotoxic effects on the TREx-HeLa-Vpr cell line.

Correction to: Biosynthesis of medicinally important plant metabolites by unusual type III polyketide synthases.

The article Biosynthesis of medicinally important plant metabolites by unusual.

Biosynthesis of medicinally important plant metabolites by unusual type III polyketide synthases.

Recent research progress on the "second generation" type III polyketide synthases is summarized. This class of enzymes catalyzes unusual condensation chemistries of CoA thioesters to generate various core structures of medicinally important plant secondary metabolites, including the R1-C-R2 scaffold of alkyl quinolones, curcuminoids, as well as the 8-azabicyclo[3.2.1]octane ring of tropane alkaloids. The discovery of this fascinating enzyme superfamily provides excellent opportunities for the manipulation of the enzyme reactions to expand the supply of natural and unnatural molecules for future drug development.

Three new quassinoids isolated from the wood of Picrasma javanica and their anti-Vpr activities.

Three new quassinoids, javanicinols A and B (1 and 2) and 4-keto-(16S)-methoxyjavanicin B (3), together with three known quassinoids (4-6) were isolated from the chloroform-soluble fraction of the methanol extract of the Picrasma javanica wood. The structures of 1-3 were determined by spectroscopic analyses, including 1D and 2D NMR, HRESIMS, and CD. The anti-HIV-1 viral protein R (Vpr) assay revealed that 1 and 2 exhibited potent anti-Vpr activities at 1.25 µM. Furthermore, the assay also revealed the potent anti-Vpr activities of (16R)-methoxyjavanicin B (7) and (16S)-methoxyjavanicin B (8), which were previously isolated from the Picrasma javanica wood.

Identification and characterization of N9-methyltransferase involved in converting caffeine into non-stimulatory theacrine in tea.

Caffeine is a major component of xanthine alkaloids and commonly consumed in many popular beverages. Due to its occasional side effects, reduction of caffeine in a natural way is of great importance and economic significance. Recent studies reveal that caffeine can be converted into non-stimulatory theacrine in the rare tea plant Camellia assamica var. kucha (Kucha), which involves oxidation at the C8 and methylation at the N9 positions of caffeine. However, the underlying molecular mechanism remains unclear. Here, we identify the theacrine synthase CkTcS from Kucha, which possesses novel N9-methyltransferase activity using 1,3,7-trimethyluric acid but not caffeine as a substrate, confirming that C8 oxidation takes place prior to N9-methylation. The crystal structure of the CkTcS complex reveals the key residues that are required for the N9-methylation, providing insights into how caffeine N-methyltransferases in tea plants have evolved to catalyze regioselective N-methylation through fine tuning of their active sites. These results may guide the future development of decaffeinated drinks.

Bis-iridoid and iridoid glycosides: Viral protein R inhibitors from Picrorhiza kurroa collected in Myanmar.

Four new bis-iridoid glycosides, saungmaygaosides A-D (1-4), and six known iridoid glycosides (5-10) were isolated from the n-butanol extract of the stems of Picrorhiza kurroa collected in Myanmar. Their structures were elucidated by extensive spectroscopic techniques. All of the isolates were assayed for anti-Vpr activity, using TREx-HeLa-Vpr cells. Among the isolates, saungmaygaoside D (4), sylvestroside IV dimethyl acetal (7), and sweroside (8) were the most potent inhibitors with effective doses of 5 and 10 µM, respectively, without showing any notable cytotoxicities.

Dinorcassane Diterpenoid from Boesenbergia rotunda Rhizomes Collected in Lower Myanmar.

A new dinorcassane diterpenoid, seikphoochinal A (1), and four known compounds, pinostrobin (2), 4',7-dimethylkaempferol (3), and galanals A (4) and B (5), were isolated from the chloroform-soluble crude extract of wild type Boesenbergia rotunda rhizomes collected in Lower Myanmar. The chemical structures of these compounds were identified, using a combination of spectroscopic methods. The presence of the diterpenoids 1, 4, and 5 demonstrated the structural diversity of wild type B. rotunda. Among the isolates, compounds 4 and 5 exhibited significant antiproliferative activities against a small panel of human cancer cell lines, including lung (LK-2, A549), stomach (ECC4), breast (MCF7), cervix (HeLa), and prostate (DU145).

Lignans with melanogenesis effects from Premna serratifolia wood.

Three new lignoids, premnan A (1), premnan B (2), and tauntangyiol C (3), were isolated from Premna serratifolia wood, a traditional cosmetic plant in Myanmar, together with a new lignoid, premnan C (4) assumed to be an artifact, one natural new lignoid (5), and three known lignoids (6-8). The structures of the new compounds 1-4 were elucidated based on 1D and 2D NMR, IR spectroscopy, and HRESIMS. The absolute configurations of 1-4 were also determined by optical rotation, circular dichroism (CD) data analyses, and comparisons with the reported literature. All isolated compounds were tested for their melanogenesis activities against the B16-F10 mouse melanoma cell line. Compounds 1 and 4 showed melanogenesis enhancing activities of 31% and 50%, respectively, at a 50 µM concentration. Compounds 2, 3, and 6 increased melanin production by 67%, 30%, and 45%, respectively, at a 100 µM concentration, without any cytotoxicity.

Two new pyrrolo-2-aminoimidazoles from a Myanmarese marine sponge, Clathria prolifera.

Marine organisms such as marine sponges and soft corals are valuable sources of pharmacologically active secondary metabolites. In our ongoing research on the discovery of new secondary metabolites from marine organisms, two new pyrrolo-2-aminoimidazoles, clathriroles A (1) and B (2), were isolated from the water-soluble portion prepared from the methanol and acetone (2:1) extract of the marine sponge, Clathria prolifera, collected in Myanmar. The chemical structures of the isolated compounds were determined using extensive spectroscopic techniques, including NMR, HRESIMS, IR, and optical rotation, and comparisons with the reported literature. The spectroscopic analyses of 1 and 2 suggested that 1 is an enantiomer of antifungal N-methylmanzacidin C isolated from the marine sponge Axinella brevistyla, whereas 2 is a diastereomer of manzacidin D at C-11 isolated from the marine sponge Astrosclera willeyana. To the best of our knowledge, this is the first report of the isolation of the pyrrolo-2-aminoimidazole compounds from C. prolifera. Furthermore, in contrast to the potency of N-methylmanzacidin C against Saccharomyces cerevisiae, the antifungal assay revealed that 1 and 2 lack any activity against this strain. Thus, these observations may suggest that the absolute configurations at both C-9 and C-11 play an important role in controlling the antifungal activity of this type of compound.

Tetrahydrofuran lignans: Melanogenesis inhibitors from Premna integrifolia wood collected in Myanmar.

Two new tetrahydrofuran lignans, taungtangyiols A (1) and B (2), and eight known furofuran lignans (3-10), were isolated from the chloroform extract of Premna integrifolia wood collected in Myanmar. Their structures were elucidated by extensive spectroscopic techniques. The X-ray crystal structure of 1 clearly indicated its relative configuration. Taungtangyiols A (1) and B (2) inhibited the deposition of melanin in B16F10 mouse melanoma cells, with IC50 values of 50.7 and 40.9 µM, respectively, without notable cytotoxicity. An SAR study demonstrated that the furofuran and dioxymethylene moieties of the lignans play a vital role in inhibiting melanogenesis.

3,5-Dimethylorsellinic Acid Derived Meroterpenoids from Penicillium chrysogenum MT-12, an Endophytic Fungus Isolated from Huperzia serrata.

Eight new chrysogenolides (A-H (1-8)) and seven known (9-15) 3,5-dimethylorsellinic acid derived meroterpenoids were isolated from the solid substrate fermentation cultures of a Huperzia serrata endophytic fungus, Penicillium chrysogenum MT-12. The structures of the new compounds were elucidated by interpretation of spectroscopic and spectrometric data (1D and 2D NMR, IR, and HRESIMS). The absolute configurations of 1-4 were determined by single-crystal X-ray crystallographic analysis, and those of 5-8 were assigned on the basis of experimental and calculated electronic circular dichroism spectra. Compounds 3, 4, 6, 11, and 12 showed inhibition of nitric oxide production in lipopolysaccharide-activated RAW 264.7 macrophage cells with IC50 values in the range of 4.3-78.2 µM (positive control, indomethacin, IC50 = 33.6 ± 1.4 µM).

Labdane diterpenoids from Curcuma amada rhizomes collected in Myanmar and their antiproliferative activities.

Four new labdane diterpenoids, 12ß-hydroxy-15-norlabda-8(17),13(14)-dien-16-oic acid (1), (E)-15-ethoxy-15-methoxylabda-8(17),12-dien-16-al (2), (E)-15α-ethoxy-14α-hydroxylabda-8(17),12-dien-16-olide (3), and 15-ethoxy-12ß-hydroxylabda-8(17),13(14)-dien-16,15-olide (4) were isolated from the methanol extract of Curcuma amada rhizomes collected in Myanmar, together with 13 known analogs. Their structures were elucidated by extensive spectroscopic techniques. All of the isolates were evaluated for their antiproliferative activities against a small panel of five different human cancer cell lines (A549, human lung cancer; HeLa, human cervical cancer; MCF7, human breast cancer; PANC-1 and PSN-1, human pancreatic cancer). Among the isolates, compounds 2-4, 7, 8, 12, and 17 showed mild antiproliferative activities with IC50 values ranging from 19.7 to 96.1µM. (E)-14-Hydroxy-15-norlabda-8(17),12-dien-16-al (11) exhibited strong antiproliferative activities selectively against HeLa, PANC-1, and PSN-1 cells, with IC50 values of 5.88, 1.00, and 3.98µM, respectively. These potencies were comparable to those of the positive control, 5-fluorouracil.

Naturally occurring Vpr inhibitors from medicinal plants of Myanmar.

Human immunodeficiency virus type-1 (HIV-1) is a lentiviral family member that encodes the retroviral Gag, Pol, and Env proteins, along with six additional accessory proteins, Tat, Rev, Vpu, Vif, Nef, and Vpr. The currently approved anti-HIV drugs target the Pol and Env encoded proteins. However, these drugs are only effective in reducing viral replication. Furthermore, the drugs' toxicities and the emergence of drug-resistant strains have become serious worldwide problems. Resistance eventually arises to all of the approved anti-HIV drugs, including the newly approved drugs that target HIV integrase (IN). Drug resistance likely emerges because of spontaneous mutations that occur during viral replication. Therefore, new drugs that effectively block other viral components must be developed to reduce the rate of resistance and suppress viral replication with little or no long-term toxicity. The accessory proteins may expand treatment options. Viral protein R (Vpr) is one of the promising drug targets among the HIV accessory proteins. However, the search for inhibitors continues in anti-HIV drug discovery. In this review, we summarize the naturally occurring compounds discovered from two Myanmar medicinal plants as well as their structure-activity relationships. A total of 49 secondary metabolites were isolated from Kaempferia pulchra rhizomes and Picrasama javanica bark, and the types of compounds were identified as isopimarane diterpenoids and picrasane quassinoids, respectively. Among the isolates, 7 diterpenoids and 15 quassinoids were found to be Vpr inhibitors lacking detectable toxicity, and their potencies varied according to their respective functionalities.

2-Alkylquinolone alkaloid biosynthesis in the medicinal plant Evodia rutaecarpa involves collaboration of two novel type III polyketide synthases.

2-Alkylquinolone (2AQ) alkaloids are pharmaceutically and biologically important natural products produced by both bacteria and plants, with a wide range of biological effects, including antibacterial, cytotoxic, anticholinesterase, and quorum-sensing signaling activities. These diverse activities and 2AQ occurrence in vastly different phyla have raised much interest in the biosynthesis pathways leading to their production. Previous studies in plants have suggested that type III polyketide synthases (PKSs) might be involved in 2AQ biosynthesis, but this hypothesis is untested. To this end, we cloned two novel type III PKSs, alkyldiketide-CoA synthase (ADS) and alkylquinolone synthase (AQS), from the 2AQ-producing medicinal plant, Evodia rutaecarpa (Rutaceae). Functional analyses revealed that collaboration of ADS and AQS produces 2AQ via condensations of N-methylanthraniloyl-CoA, a fatty acyl-CoA, with malonyl-CoA. We show that ADS efficiently catalyzes the decarboxylative condensation of malonyl-CoA with a fatty acyl-CoA to produce an alkyldiketide-CoA, whereas AQS specifically catalyzes the decarboxylative condensation of an alkyldiketide acid with N-methylanthraniloyl-CoA to generate the 2AQ scaffold via C-C/C-N bond formations. Remarkably, the ADS and AQS crystal structures at 1.80 and 2.20 Å resolutions, respectively, indicated that the unique active-site architecture with Trp-332 and Cys-191 and the novel CoA-binding tunnel with Tyr-215 principally control the substrate and product specificities of ADS and AQS, respectively. These results provide additional insights into the catalytic versatility of the type III PKSs and their functional and evolutionary implications for 2AQ biosynthesis in plants and bacteria.

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.

Effective Production of Aromatic Polyketides in Streptomyces using a Combined-Culture Method.

Combinedculture is a fermentation method which efficiently induces secondary metabolite production in Streptomyces by co-culturing them with mycolic acid-containing bacteria. As a result of combined-culture screening -of our terrestrial Streptomyces collection using UV-HPLC, one of the tested strains, Streptoinyces. sp. TAKO-2, produced two known aromatic polyketides, julichrome Q6 (1) and julichrome Q8.8 (2), when co-cultured with the mycolic acid- containing bacterium Tsukamurella pulmonis TP-B0596. The structures of 1 and 2 were confirmed by spectroscopic analysis and literature data.

Induced production of mycotoxins in an endophytic fungus from the medicinal plant Datura stramonium L.

Epigenetic modifiers, including DNA methyltransferase (DNMT) or histone deacetylase (HDAC) inhibitors, are useful to induce the expression of otherwise dormant biosynthetic genes under standard laboratory conditions. We isolated several endophytic fungi from the medicinal plant Datura stramonium L., which produces pharmaceutically important tropane alkaloids, including scopolamine and hyoscyamine. Although none of the endophytic fungi produced the tropane alkaloids, supplementation of a DNMT inhibitor, 5-azacytidine, and/or a HDAC inhibitor, suberoylanilide hydroxamic acid, to the culture medium induced the production of mycotoxins, including alternariol, alternariol-5-O-methyl ether, 3'-hydroxyalternariol-5-O-methyl ether, altenusin, tenuazonic acid, and altertoxin II, by the endophytic fungus Alternaria sp. This is the first report of a mycotoxin-producing endophytic fungus from the medicinal plant D. stramonium L. This work demonstrates that treatments with epigenetic modifiers induce the production of mycotoxins, thus providing a useful tool to explore the biosynthetic potential of the microorganisms.