Alcyonacean metabolites VII--chemical constituents of Lobophytum denticulatum and Lobophytum strictum of the Indian Ocean

A rare cembranoid diterpene, (7E,11E.1 R,2S,3R,4R,l4S)-14-acetoxy-3,4-epoxycembra-7,11,15-triene-17,2-olide (1), was isolated from Lobophytum denticulatum and a new polyhydroxysterol, 7-hydroxyandamansterol (9) has been identified as peracetyl derivative from Lobophytum strictum. Several known polyhydroxysterols have also been isolated from these organisms. 1 exhibited moderate antibacterial activity.     

昆明山海棠化学成分的研究

从昆明山海棠(Tripterygium hypoglaucum (Lévl) Hutch)茎中分得一单体,经光谱UV,IR,MS,¹H-NMR,¹³C-NMR和2D-NMR)分析,推定为一新成分,命名为雷酚二萜酸(triptonoditerpenic acid).     

A new acyclic diterpene acid and bioactive compounds from <Emphasis Type="Italic">Knema glauca</Emphasis>

Investigation of the chemical constituents of the fruits of Knema glauca (Myristicaceae) yielded a new acyclic diterpene acid, named glaucaic acid 4, together with four acylphenols, including 1-(2,6-dihydroxyphenyl) tetradecan-1-one 1, malabaricone A 6, dodecanoylphloroglucinol 7 and 1-(2,4,6-trihydroxyphenyl)-9-phenylnonan-1-one 8, two lignans sesamin 2 and asarinin 3, and a flavan, myristinin D 5. In addition, myristinin A 9 and (±)-7,4′-dihydroxy-3′-methoxyflavan 10 were isolated from its leaves and stems, respectively. When tested against small-cell lung cancer (NCI-H187), epidermoid carcinoma (KB) and breast cancer (BC) cell lines, compounds 1, 6–8 and 10 displayed weak to moderate cytotoxicity. The acylphenols 6–8 displayed antituberculosis activity against the microbe Mycobacterium tuberculosis with MIC values of 25, 50 and 100 μg/mL, respectively, and antiviral activity against herpes simplex virus type 1, with 7 as the most active compound (IC₅₀ = 3.05 μg/mL). Malabaricone A 6 was also active against the malarial parasite Plasmodium falciparum with an IC₅₀ value of 2.78 μg/mL.     

前益母草素(prehispanolone)对小鼠T,B淋巴细胞的影响

用[³H]-thymidine参入法表明:益母草半日花烷型双环二萜——前益母草素(prehi-spanolone,LC-5504)对由Con A活化的T淋巴细胞有较强的促进增殖作用。其作用是单独使用Con A的5～8倍。实验研究提示,前益母草素能够增强机体的细胞免疫功能。     

Novel diterpenes with cytotoxic, anti-malarial and anti-tuberculosis activities from a brown alga Dictyota sp

Two new diterpenes, (1E,2R*,3R*,4S*,6E,18S*)-4,18-dihydroxydictyolactone (1) and 8alpha,11-dihydroxypachydictyol A (2), together with fucoxanthin (3) and a known diterpene 4alpha-hydroxycrenulatane (4) were isolated from the methanol extract of the brown alga Dictyota sp. collected from Bangsaen Beach, Thailand. Their structures were determined by spectroscopic data. The relative stereochemistry of 1 was established by NOE difference and NOESY experiments. Compound 1 showed weak anti-tuberculosis activity while 2 displayed strong cytotoxicity against the NCI-H187 cell line, and potent anti-malarial activity. In addition, 3 was active against HSV-1 and malarial parasites.     

Anti-inflammatory effect of kaurenoic acid, a diterpene from Copaifera langsdorffii on acetic acid-induced colitis in rats

Kaurenoic acid, a diterpene from Copaifera langsdorffii (Leguminaceae), was evaluated on rat colitis induced by acetic acid. Rats were pretreated orally (15 and 2 h before) or rectally 2 h before induction of colitis with kaurenoic acid (50 and 100 mg/kg) or vehicle (1 ml, 3% DMSO). Colitis was induced by intracolonic instillation of a 2 ml of 4% (v/v) acetic acid solution and, 24 h later, the colonic mucosal damage was analysed macroscopically for the severity of mucosal damage, the myeloperoxidase (MPO) activity and the malondialdehyde (MDA) levels in the colon segments. A marked reduction in gross damage score (52% and 42%) and wet weight of damaged colon tissue (39% and 32%) were observed in rats that received 100 mg/kg kaurenoic acid, respectively, by rectal and oral routes. This effect was confirmed biochemically by a two- to three-fold reduction of colitis associated increase in MPO activity, the marker of neutrophilic infiltration and by a marked decrease in MDA level, an indicator of lipoperoxidation in colon tissue. Furthermore, light microscopy revealed the marked diminution of inflammatory cell infiltration and submucosal edema formation in the colon segments of rats treated with the test compound. These findings indicate the anti-inflammatory potential of kaurenoic acid in acetic acid-induced colitis.     

In vitro study of the effects of plaunotol on oral cell proliferation and wound healing

Plaunotol is an acyclic diterpene alcohol extracted from a medicinal plant called plau-noi, Croton stellatopilosus Ohba, and has been widely used for the treatment of gastric ulcers in Japan. The aim of this study was to examine the effects of plaunotol on human gingival fibroblasts (HGFs) and human oral keratinocytes (HOKs). To assess the cytotoxic effect, HGFs and HOKs were treated with plaunotol. Subsequently, the morphology of cells was recorded and cells were subjected to MTT assay. To investigate cell proliferation effect, cells were treated with plaunotol and counted with a haemocytometer. To determine wound healing effect, the number of cells repopulated into the wounded areas in monolayer culture and in fibroblast-populated collagen lattice (FPCL) was measured. The results showed that 10 and 1?μg/ml (33 and 3.3?μmol/l) plaunotol induced toxicity in HGFs and HOKs, respectively. However, 0.1?μg/ml (0.33?μmol/l) plaunotol promoted HGF proliferation and wound healing in monolayer and FPCL models. In contrast, 0.1?μg/ml plaunotol could not induce HOK proliferation nor in vitro wound healing using monolayer culture, but it induced wound healing in a modified FPCL model. Our data suggested that plaunotol could promote oral cell proliferation and wound healing in vitro and may have an implication on oral wound healing.     

Pubescenes, jatrophane diterpenes, from Euphorbia pubescens, with multidrug resistance reversing activity on mouse lymphoma cells

The macrocyclic jatrophane diterpene polyesters, pubescenes A - D ( 1 - 4) were isolated from the whole dried plant of Euphorbia pubescens, and evaluated for multidrug resistance (MDR) reversing activity on mouse lymphoma cells. All the compounds displayed very strong activity compared with the positive control verapamil. Pubescene D ( 4) is a new compound, whose structure was established as 3beta,9alpha,-diacetoxy-7beta-benzoyloxy-15beta-hydroxy-14-oxo-2beta H-jatropha-5 E,12 E-diene by spectroscopic methods, including (1)H- (1)H COSY, HMQC, HMBC and NOESY.     

The induction of senescence-like growth arrest by protein kinase C-activating diterpene esters in solid tumor cells

We previously identified the induction of senescence in melanoma cell lines sensitive to diterpene esters, indicating a therapeutic potential. Here we compared the cytostatic effects of two diterpene esters: the prototypic PKC-activating drug TPA (12-O-tetradecanoylphorbol-13-acetate), and the novel compound PEP008 (20-O-acetyl-ingenol-3-angelate) in cell lines derived from melanoma, breast cancer and colon cancer. The diterpene esters induced permanent growth arrest with characteristics of senescence in a subset of cell lines in all three solid tumor models at 100–1000 ng/ml. Use of the PKC inhibitor bisindolylmaleimide-l demonstrated that activation of PKC was required for growth arrest. Full genome expression profiling identified pivotal genes involved in DNA synthesis and cell cycle control down-regulated by treatment in all three sensitive tumor models. At the protein level, prolonged down-regulation of E2F-1 and proliferating cell nuclear antigen (PCNA), sustained expression of p21^WAF1/CIP1 and dephosphorylation of retinoblastoma (Rb) occurred in the sensitive cells. Additionally, the type II tumor suppressor HRASLS3, which has a role in mitogen-activated protein kinase (MAPK) pathway suppression, was constitutively elevated in cell lines resistant to the senescence effects compared to their sensitive counterparts. Together, these results demonstrate that both TPA and the novel PKC-activating drug PEP008 induce growth arrest with characteristics of senescence in solid tumor cell lines derived from a variety of tissue types, and by a similar mechanism. PKC-activating diterpene esters may therefore have therapeutic potential in a subset of breast cancer, colon cancer and melanoma tumors.     

Skin Irritant Ingenol Esters from Euphorbia esula.

An ethyl acetate fraction of aerial parts of E. ESULA L. collected in Germany was shown to exhibit irritant and weak tumor promoting but not solitary carcinogenic activity. The diterpene moiety of the irritants and tumor promotors contained in E. esula was identified to be ingenol (I). From aerial parts the irritant 3-0-(2E,4Z)-decadienoylingenol (III, Euphorbia factor E (1)) and a mixture E (M) of irritant Euphorbia factors shown to contain Euphorbia factor E (1), 3-0-(2,4,6)-decatrienoylingenol (V, Euphorbia factor E (2)) and 3-0-(2,4,6,8)-dodecatetraenoylingenol (VI, Euphorbia factor E (3)) were isolated. The antileukemic ingenol-3,20-dibenzoate (IX) was not found in the plant material investigated here. IX, a strong irritant, prepared by partial synthesis from ingenol (I) proved to be a weak tumor promoter as well.     

Alcyonacean Metabolites VII – Chemical Constituents of Lobophytum denticulatum and Lobophytum strictum of the Indian Ocean

Abstract

A rare cembranoid diterpene, (7E,11E,1R,2S,3R,4R,14S)-14-acetoxy-3,4-epoxycembra-7,11,15-triene-17,2-olide (1), was isolated from Lobophytum denticulatum and a new polyhydroxysterol, 7α-hydroxyandamansterol (9) has been identified as peracetyl derivative from Lobophytum strictum. Several known polyhydroxysterols have also been isolated from these organisms. 1 exhibited moderate antibacterial activity.     

Diterpene glycosides from Aster homochlamydeus

A new diterpene glycoside 1 was isolated from the whole plant of Aster homochlamydeus Hand-mazz (Compositae), along with four known diterpene glycosides 2-5. Their structures were elucidated by spectroscopic methods, MS, IR, NMR and X-raycrystallographic analyisis. Antibacterial activity of compounds 1-5 was observed.     

Terpenoids from the tuber of Cremastra appendiculata

Two new terpenoids including a cadinane sesquiterpene (1), and an ent-kaurane diterpene diglycoside (2), together with a known triterpene containing 32 carbons (3), have been isolated from the ethanolic extract of Cremastra appendiculata. Their structures were established by the spectroscopic methods including the IR, MS, 1D-, and 2D-NMR experiments as ( - )-cadin-4,10(15)-dien-11-oic acid (1), ( - )-ent-12beta-hydroxykaur-16-en-19-oic acid, 19-O-beta-D-xylopyranosyl-(1 --> 6)-O-beta-D-glucopyranoside (2), and (+)-24,24-dimethyl-25,32-cyclo-5alpha-lanosta-9(11)-en-3beta-ol (3). Compounds 1-3 were evaluated against several human cancer cell lines. Compound 3 showed in vitro-selective cytotoxicity against human breast cancer cell lines (MCF-7) with an IC50 of 3.18 microM, but 1 and 2 were inactive (IC50>10 microg/ml).     

Terpenoids from the tuber of Cremastra appendiculata

Two new terpenoids including a cadinane sesquiterpene (1), and an ent-kaurane diterpene diglycoside (2), together with a known triterpene containing 32 carbons (3), have been isolated from the ethanolic extract of Cremastra appendiculata. Their structures were established by the spectroscopic methods including the IR, MS, 1D-, and 2D-NMR experiments as ( - )-cadin-4,10(15)-dien-11-oic acid (1), ( - )-ent-12beta-hydroxykaur-16-en-19-oic acid, 19-O-beta-d-xylopyranosyl-(1 --> 6)-O-beta-d-glucopyranoside (2), and (+)-24,24-dimethyl-25,32-cyclo-5alpha-lanosta-9(11)-en-3beta-ol (3). Compounds 1-3 were evaluated against several human cancer cell lines. Compound 3 showed in vitro-selective cytotoxicity against human breast cancer cell lines (MCF-7) with an IC(50) of 3.18 muM, but 1 and 2 were inactive (IC(50)>10 mug/ml).     

A new labdane diterpene from Leonurus heterophyllus

A new labdane diterpene, heteronone B (1), together with a known labdane diterpene, heteronone A (2), have been isolated from the aerial part of Leonurus heterophyllus. Their structures were established mainly by 1D and 2D NMR analysis and the stereochemistry of 2 was confirmed by single-crystal X-ray diffraction analysis.     

Indomethacin abrogates the suppression by cyclosporin A of lectin-dependent cell-mediated cytotoxicity to HEp-2 cells

The effects of cyclosporin A, prostaglandin E1 and indomethacin were studied on lectin-dependent cell-mediated cytotoxicity (LDCC) against adherent HEp-2 human epipharynx carcinoma target cells. LDCC activity by human peripheral blood lymphocytes was evaluated by detachment from the monolayer of [3H]thymidine-prelabelled HEp-2 cells in a 24-h assay at 50:1 effector:target cell ratio in the presence of 25 micrograms/ml concanavalin A. Under these conditions, but without concanavalin A, considerable natural cell-mediated cytotoxicity was not elicited although LDCC was significantly augmented in the presence of concanavalin A. Addition of both cyclosporin A (0.1, 1.0 or 10 micrograms/ml) and prostaglandin E1 (10(-8), 10(-7) or 10(-6) M) dose-dependently suppressed LDCC activity. Indomethacin (0.1, 1.0 or 10 micrograms/ml) did not in itself influence LDCC although suppression of LDCC by cyclosporin A, but not prostaglandin E1, was abrogated in the presence of indomethacin. Similar to indomethacin, acetyl salicylic acid also reversed the inhibition of LDCC by cyclosporin A. In parallel experiments, cyclosporin A elicited a more than two-fold increase of prostaglandin E production under LDCC assay conditions as measured by radioimmunoassay. Contrary to LDCC, depression of concanavalin A induced blastogenesis by cyclosporin A was not influenced by indomethacin, suggesting that the inhibition by cyclosporin A of LDCC and concanavalin A-induced blastogenesis proceed via different mechanisms.     

In vitro susceptibility of four serotypes of enterohaemorrhagic Escherichia coli to antimicrobial agents

We evaluated the in vitro susceptibility of four serotypes of enterohaemorrhagic Escherichia coli (E. coli 026, E. coli O111, E. coli O157, and E. coli O165) with diverse DNA patterns to antimicrobial agents. The minimum inhibitory concentrations (MIC) determined in a total of 83 strains using Mueller-Hinton agar under aerobic and anaerobic conditions were 0.015-0.12 microg/ml for ciprofloxacin, 0.06-1 microg/ml for norfloxacin, 2-64 microg/ml for fosfomycin without glucose-6-phosphate (G-6-P), 0.25-32 microg/ml for fosfomycin with G-6-P, 2- > or = 256 microg/ml for kanamycin, 0.125-2 microg/ml for cefoperazone, and 0.06-1 microg/ml for ceftazidime. The MIC of ciprofloxacin, norfloxacin, cefoperazone, and ceftazidime were low in all strains examined.     

New phenylpropane and anti-inflammatory diterpene derivatives from Amentotaxus formosana

One new diterpene, 8(14),15-sandaracopimaradiene-2alpha,3beta,18-triol (1), two new phenylpropane derivatives, i.e., (E)-methyl 2-(3,4-methylene-dioxyphenyl)-3-methoxypropenoate (2) and (E)-2-(3,4-methylene-dioxyphenyl)-3-methoxypropenoic acid (3), and two known diterpenes, ent-8(14),15-sandaracopimaradiene-2alpha,18-diol (4) and 8(14),15-sandaracopimaradiene-2alpha,18,19-triol (5), were isolated from the heartwoods and barks of Amentotaxus formosana, respectively. The anti-inflammatory activity of the diterpenes 1, 4, and 5 was assessed in vitro by determining their inhibitory effects on the chemical mediators released from mast cells, neutrophils, macrophages, and microglial cells. Compounds 1, 4, and 5 showed significant concentration-dependent inhibitory effects on the release of beta-glucuronidase from rat neutrophils in response to formyl-Met-Leu-Phe/cytochalasin B (fMLP/CB) with IC50 values of 5.5 +/- 1.8, 8.4 +/- 2.9 and 19.2 +/- 3.3 microM, respectively. Compounds 1 and 5 also showed significant concentration-dependent inhibitory effects on superoxide anion generation in rat neutrophils stimulated with fMLP/CB and phorbol 12-myristate 13-acetate (PMA) with IC50 values of 12.6 +/- 1.2 and 9.4 +/- 1.7, and 10.7 +/- 3.3 and 12.9 +/- 0.9 microM, respectively.     

In vitro study of the effects of plaunotol on oral cell proliferation and wound healing

Plaunotol is an acyclic diterpene alcohol extracted from a medicinal plant called plau-noi, Croton stellatopilosus Ohba, and has been widely used for the treatment of gastric ulcers in Japan. The aim of this study was to examine the effects of plaunotol on human gingival fibroblasts (HGFs) and human oral keratinocytes (HOKs). To assess the cytotoxic effect, HGFs and HOKs were treated with plaunotol. Subsequently, the morphology of cells was recorded and cells were subjected to MTT assay. To investigate cell proliferation effect, cells were treated with plaunotol and counted with a haemocytometer. To determine wound healing effect, the number of cells repopulated into the wounded areas in monolayer culture and in fibroblast-populated collagen lattice (FPCL) was measured. The results showed that 10 and 1 μg/ml (33 and 3.3 μmol/l) plaunotol induced toxicity in HGFs and HOKs, respectively. However, 0.1 μg/ml (0.33 μmol/l) plaunotol promoted HGF proliferation and wound healing in monolayer and FPCL models. In contrast, 0.1 μg/ml plaunotol could not induce HOK proliferation nor in vitro wound healing using monolayer culture, but it induced wound healing in a modified FPCL model. Our data suggested that plaunotol could promote oral cell proliferation and wound healing in vitro and may have an implication on oral wound healing.     

Development of sensitive colorimetric capture ELISAs for Clostridium botulinum neurotoxin serotypes E and F.

Sensitive and specific enzyme-linked immunosorbent assays (ELISAs) were developed to detect Clostridium botulinum neurotoxin serotypes E (BoNT E) and F (BoNT F) in assay buffer and human serum. The assay is based upon affinity-purified horse polyclonal antibodies directed against the approximately 50 kD C-fragments of each toxin. Standard curves were linear over 0.5-10 ng/ml (BoNT E) or 2-20 ng/ml (BoNT F). Accurate measurements were achieved at 0.5 ng/ml (BoNT E) or 2 ng/ml (BoNT F) in assay buffer and 10% human serum. Variation between triplicates was typically 5-10%. Less than 1% cross-reactivity occurred between other serotypes A, B, E or F). When tested against toxins complexed to their neurotoxin-associated proteins, interference was absent for BoNT F. However, pure BoNT E and that complexed to associated proteins demonstrated significant quantitative differences. We believe these differences arise from trypsin activation of the toxin. These assays demonstrated sensitivities close to that of the mouse bioassay, without the use of animals, in a much simpler format than other reported assays of similar sensitivity.